Comparison of electrofishing techniques to detect larval lampreys in wadeable streams in the Pacific Northwest

Science.gov (United States)

Dunham, Jason B.; Chelgren, Nathan D.; Heck, Michael P.; Clark, Steven M.

2013-01-01

We evaluated the probability of detecting larval lampreys using different methods of backpack electrofishing in wadeable streams in the U.S. Pacific Northwest. Our primary objective was to compare capture of lampreys using electrofishing with standard settings for salmon and trout to settings specifically adapted for capture of lampreys. Field work consisted of removal sampling by means of backpack electrofishing in 19 sites in streams representing a broad range of conditions in the region. Captures of lampreys at these sites were analyzed with a modified removal-sampling model and Bayesian estimation to measure the relative odds of capture using the lamprey-specific settings compared with the standard salmonid settings. We found that the odds of capture were 2.66 (95% credible interval, 0.87–78.18) times greater for the lamprey-specific settings relative to standard salmonid settings. When estimates of capture probability were applied to estimating the probabilities of detection, we found high (>0.80) detectability when the actual number of lampreys in a site was greater than 10 individuals and effort was at least two passes of electrofishing, regardless of the settings used. Further work is needed to evaluate key assumptions in our approach, including the evaluation of individual-specific capture probabilities and population closure. For now our results suggest comparable results are possible for detection of lampreys by using backpack electrofishing with salmonid- or lamprey-specific settings.

Sequencing of the sea lamprey (Petromyzon marinus) genome provides insights into vertebrate evolution

Science.gov (United States)

Smith, Jeramiah J; Kuraku, Shigehiro; Holt, Carson; Sauka-Spengler, Tatjana; Jiang, Ning; Campbell, Michael S; Yandell, Mark D; Manousaki, Tereza; Meyer, Axel; Bloom, Ona E; Morgan, Jennifer R; Buxbaum, Joseph D; Sachidanandam, Ravi; Sims, Carrie; Garruss, Alexander S; Cook, Malcolm; Krumlauf, Robb; Wiedemann, Leanne M; Sower, Stacia A; Decatur, Wayne A; Hall, Jeffrey A; Amemiya, Chris T; Saha, Nil R; Buckley, Katherine M; Rast, Jonathan P; Das, Sabyasachi; Hirano, Masayuki; McCurley, Nathanael; Guo, Peng; Rohner, Nicolas; Tabin, Clifford J; Piccinelli, Paul; Elgar, Greg; Ruffier, Magali; Aken, Bronwen L; Searle, Stephen MJ; Muffato, Matthieu; Pignatelli, Miguel; Herrero, Javier; Jones, Matthew; Brown, C Titus; Chung-Davidson, Yu-Wen; Nanlohy, Kaben G; Libants, Scot V; Yeh, Chu-Yin; McCauley, David W; Langeland, James A; Pancer, Zeev; Fritzsch, Bernd; de Jong, Pieter J; Zhu, Baoli; Fulton, Lucinda L; Theising, Brenda; Flicek, Paul; Bronner, Marianne E; Warren, Wesley C; Clifton, Sandra W; Wilson, Richard K; Li, Weiming

2013-01-01

Lampreys are representatives of an ancient vertebrate lineage that diverged from our own ~500 million years ago. By virtue of this deeply shared ancestry, the sea lamprey (P. marinus) genome is uniquely poised to provide insight into the ancestry of vertebrate genomes and the underlying principles of vertebrate biology. Here, we present the first lamprey whole-genome sequence and assembly. We note challenges faced owing to its high content of repetitive elements and GC bases, as well as the absence of broad-scale sequence information from closely related species. Analyses of the assembly indicate that two whole-genome duplications likely occurred before the divergence of ancestral lamprey and gnathostome lineages. Moreover, the results help define key evolutionary events within vertebrate lineages, including the origin of myelin-associated proteins and the development of appendages. The lamprey genome provides an important resource for reconstructing vertebrate origins and the evolutionary events that have shaped the genomes of extant organisms. PMID:23435085

Pacific Lamprey Research and Restoration Project, Annual Report 2002.

Energy Technology Data Exchange (ETDEWEB)

Close, David; Aronsuu, Kimmo; Jackson, Aaron

2003-07-01

Pacific lamprey (Lampetra tridentata) has significantly declined along the Oregon coast and in the Columbia River Basin (Downey et al. 1993, Close and Jackson 2001). Declines in adults can be partially attributed to hydroelectric dams, which have impeded passage of adult Pacific lamprey in the Columbia and Snake rivers (Moser et al. 2002), thus effecting larval recruitment in the basin (Moser and Close in press). Adult Pacific lamprey also declined in numbers in the Umatilla River, a tributary of the Columbia River (Close and Jackson 2001). In addition to hydro power dams in the Columbia River, habitat alterations and chemical treatments have been involved in the collapse of Pacific lamprey populations in the Umatilla River (Close 1999). To initiate the restoration effort, CTUIR began developing a restoration plan in 1998. The goal of the lamprey research and restoration project is to restore natural production of Pacific lampreys in the Umatilla River to self-sustaining and harvestable level. This report is summarizing the studies and restoration efforts concluded in 2002.

Fine-scale pathways used by adult sea lampreys during riverine spawning migrations

Science.gov (United States)

Holbrook, Christopher; Bergstedt, Roger A.; Adams, Noah S.; Hatton, Tyson; McLaughlin, Robert L.

2015-01-01

Better knowledge of upstream migratory patterns of spawning Sea Lampreys Petromyzon marinus, an invasive species in the Great Lakes, is needed to improve trapping for population control and assessment. Although trapping of adult Sea Lampreys provides the basis for estimates of lake-wide abundance that are used to evaluate the Sea Lamprey control program, traps have only been operated at dams due to insufficient knowledge of Sea Lamprey behavior in unobstructed channels. Acoustic telemetry and radiotelemetry were used to obtain movement tracks for 23 Sea Lampreys in 2008 and 18 Sea Lampreys in 2009 at two locations in the Mississagi River, Ontario. Cabled hydrophone arrays provided two-dimensional geographic positions from acoustic transmitters at 3-s intervals; depth-encoded radio tag detections provided depths. Upstream movements occurred at dusk or during the night (2015–0318 hours). Sea Lampreys were closely associated with the river bottom and showed some preference to move near banks in shallow glide habitats, suggesting that bottom-oriented gears could selectively target adult Sea Lampreys in some habitats. However, Sea Lampreys were broadly distributed across the river channel, suggesting that the capture efficiency of nets and traps in open channels would depend heavily on the proportion of the channel width covered. Lack of vertical movements into the water column may have reflected lamprey preference for low water velocities, suggesting that energy conservation was more beneficial for lampreys than was vertical searching in rivers. Improved understanding of Sea Lamprey movement will assist in the development of improved capture strategies for their assessment and control in the Great Lakes.

Habituation of adult sea lamprey repeatedly exposed to damage-released alarm and predator cues

Science.gov (United States)

Imre, Istvan; Di Rocco, Richard T.; Brown, Grant E.; Johnson, Nicholas

2016-01-01

Predation is an unforgiving selective pressure affecting the life history, morphology and behaviour of prey organisms. Selection should favour organisms that have the ability to correctly assess the information content of alarm cues. This study investigated whether adult sea lamprey Petromyzon marinus habituate to conspecific damage-released alarm cues (fresh and decayed sea lamprey extract), a heterospecific damage-released alarm cue (white sucker Catostomus commersoniiextract), predator cues (Northern water snake Nerodia sipedon washing, human saliva and 2-phenylethylamine hydrochloride (PEA HCl)) and a conspecific damage-released alarm cue and predator cue combination (fresh sea lamprey extract and human saliva) after they were pre-exposed 4 times or 8 times, respectively, to a given stimulus the previous night. Consistent with our prediction, adult sea lamprey maintained an avoidance response to conspecific damage-released alarm cues (fresh and decayed sea lamprey extract), a predator cue presented at high relative concentration (PEA HCl) and a conspecific damage-released alarm cue and predator cue combination (fresh sea lamprey extract plus human saliva), irrespective of previous exposure level. As expected, adult sea lamprey habituated to a sympatric heterospecific damage-released alarm cue (white sucker extract) and a predator cue presented at lower relative concentration (human saliva). Adult sea lamprey did not show any avoidance of the Northern water snake washing and the Amazon sailfin catfish extract (heterospecific control). This study suggests that conspecific damage-released alarm cues and PEA HCl present the best options as natural repellents in an integrated management program aimed at controlling the abundance of sea lamprey in the Laurentian Great Lakes.

Comparison of synthesis of 15α-hydroxylated steroids in males of four North American lamprey species

Science.gov (United States)

Bryan, Mara B.; Young, Bradley A.; Close, David A.; Semeyn, Jesse; Robinson, T. Craig; Bayer, Jennifer M.; Li, Weiming

2006-01-01

Recent studies have provided evidence that 15α-hydroxytestosterone (15α-T) and 15α-hydroxyprogesterone (15α-P) are produced in vitro and in vivo in adult male sea lampreys (Petromyzonmarinus), and that circulatory levels increase in response to injections with gonadotropin-releasing hormone (GnRH). We examined four species from the Petromyzontidae family including silver lampreys (Ichthyomyzon unicuspis), chestnut lampreys (I. castaneus), American brook lampreys (Lethenteron appendix), and Pacific lampreys (Entosphenus tridentatus) to determine if these unusual steroids were unique to sea lampreys or a common feature in lamprey species. In vitro production was examined through incubations of testis with tritiated precursors, and 15α-T and 15α-P production was confirmed in all species through co-elution with standards on both high performance liquid chromatography (HPLC) and thin layerchromatography. In vivo production was proven by demonstrating that HPLC-fractionated plasma had peaks of immunoreactive 15α-T and 15α-P that co-eluted with standards through using previously developed radioimmunoassays for 15α-T and 15α-P. The possible functionality of 15α-T and 15α-P was further examined in silver and Pacific lampreys by investigating the effect of injection of either type of lamprey GnRH on plasma concentrations of 15α-T and 15α-P. Injections with exogenous GnRH did not affect circulatory levels of either steroid in silver lampreys, and only GnRH III elicited higher levels of both steroids in Pacific lampreys. The 15α-hydroxylase enzyme(s) for steroids appeared to present in adult males of all species examined, but the question of whether 15α-hydroxylated steroids are functional in these lamprey species, and the significance of the 15-hydroxyl group, requires further research.

HPLC and ELISA analyses of larval bile acids from Pacific and western brook lampreys

Science.gov (United States)

Yun, S.-S.; Scott, A.P.; Bayer, J.M.; Seelye, J.G.; Close, D.A.; Li, W.

2003-01-01

Comparative studies were performed on two native lamprey species, Pacific lamprey (Lampetra tridentata) and western brook lamprey (Lampetra richardsoni) from the Pacific coast along with sea lamprey (Petromyzon marinus) from the Great Lakes, to investigate their bile acid production and release. HPLC and ELISA analyses of the gall bladders and liver extract revealed that the major bile acid compound from Pacific and western brook larval lampreys was petromyzonol sulfate (PZS), previously identified as a migratory pheromone in larval sea lamprey. An ELISA for PZS has been developed in a working range of 20pg-10ng per well. The tissue concentrations of PZS in gall bladder were 127.40, 145.86, and 276.96??g/g body mass in sea lamprey, Pacific lamprey, and western brook lamprey, respectively. Releasing rates for PZS in the three species were measured using ELISA to find that western brook and sea lamprey released PZS 20 times higher than Pacific lamprey did. Further studies are required to determine whether PZS is a chemical cue in Pacific and western brook lampreys. ?? 2003 Elsevier Inc. All rights reserved.

Chondrogenesis of the branchial skeleton in embryonic sea lamprey, Petromyzon marinus.

Science.gov (United States)

Morrison, S L; Campbell, C K; Wright, G M

2000-11-01

This study provides concise temporal and spatial characteristics of branchial chondrogenesis in embryonic sea lamprey, Petromyzon marinus, using high resolution light microscopy, transmission electron, and immunoelectron microscopy. Prechondrogenic condensations representing the first branchial arch appeared first in the mid-region of the third pharyngeal arch at 13 days post-fertilization (pf). Cartilage differentiation, defined by the presence of the unique, fibrillar, non-collagenous matrix protein characteristic of branchial cartilage, was first observed at 14 days pf. Development of lamprey branchial cartilage appeared unusual compared to that in jawed fishes, in that precartilage condensations appear as a one-cell wide orderly stack of flattened cells that extend by the addition of one dorsal and one ventral condensation. Development of lamprey gill arches from three condensations that fuse to form a single skeletal element differs from the developing gill arches of jawed fishes, where more than one skeletal element forms from a single condensation. The initial orderly arrangement of cells in the lamprey branchial prechondrogenic condensations remains throughout development. Once chondrification of the condensations begins, the branchial arches start to grow. Initially, growth occurs as a result of matrix secretion and cell migration. Later in development, the arches grow mainly by cell proliferation and enlargement. This study defines the morphology and timing of lamprey branchial chondrogenesis. Studies of lamprey chondrogenesis provide not only insight into the developmental biology of a unique non-collagenous cartilage in a primitive vertebrate but also into the general evolution of the skeletal system in vertebrates. Copyright 2000 Wiley-Liss, Inc.

Ecological and Cultural Importance of a Species at Risk of Extinction, Pacific Lamprey, 1964-2002 Technical Report.

Energy Technology Data Exchange (ETDEWEB)

Close, David A.

2002-07-01

The cultural and ecological values of Pacific lamprey (Lampetra tridentata) have not been understood by Euro-Americans and thus their great decline has almost gone unnoticed except by Native Americans, who elevated the issue and initiated research to restore its populations, at least in the Columbia Basin. They regard Pacific lamprey as a highly valued resource and as a result ksuyas (lamprey) has become one of their cultural icons. Ksuyas are harvested to this day as a subsistence food by various tribes along the Pacific coast and are highly regarded for their cultural value. Interestingly, our review suggests that the Pacific lamprey plays an important role in the food web, may have acted as a buffer for salmon from predators, and may have been an important source of marine nutrients to oligotrophic watersheds. This is very different from the Euro-American perception that lampreys are pests. We suggest that cultural biases affected management policies.

Use of genotyping by sequencing data to develop a high-throughput and multifunctional SNP panel for conservation applications in Pacific lamprey.

Science.gov (United States)

Hess, Jon E; Campbell, Nathan R; Docker, Margaret F; Baker, Cyndi; Jackson, Aaron; Lampman, Ralph; McIlraith, Brian; Moser, Mary L; Statler, David P; Young, William P; Wildbill, Andrew J; Narum, Shawn R

2015-01-01

Next-generation sequencing data can be mined for highly informative single nucleotide polymorphisms (SNPs) to develop high-throughput genomic assays for nonmodel organisms. However, choosing a set of SNPs to address a variety of objectives can be difficult because SNPs are often not equally informative. We developed an optimal combination of 96 high-throughput SNP assays from a total of 4439 SNPs identified in a previous study of Pacific lamprey (Entosphenus tridentatus) and used them to address four disparate objectives: parentage analysis, species identification and characterization of neutral and adaptive variation. Nine of these SNPs are FST outliers, and five of these outliers are localized within genes and significantly associated with geography, run-timing and dwarf life history. Two of the 96 SNPs were diagnostic for two other lamprey species that were morphologically indistinguishable at early larval stages and were sympatric in the Pacific Northwest. The majority (85) of SNPs in the panel were highly informative for parentage analysis, that is, putatively neutral with high minor allele frequency across the species' range. Results from three case studies are presented to demonstrate the broad utility of this panel of SNP markers in this species. As Pacific lamprey populations are undergoing rapid decline, these SNPs provide an important resource to address critical uncertainties associated with the conservation and recovery of this imperiled species. © 2014 John Wiley & Sons Ltd.

Synthesis of juvenile lamprey migration and passage research and monitoring at Columbia and Snake River Dams

Science.gov (United States)

Mesa, Matthew G.; Weiland, Lisa K.; Christiansen, Helena E.

2016-01-01

We compiled and summarized previous sources of data and research results related to the presence, numbers, and migration timing characteristics of juvenile (eyed macropthalmia) and larval (ammocoetes) Pacific lamprey Entosphenus tridentatus, in the Columbia River basin (CRB). Included were data from various screw trap collections, data from historic fyke net studies, catch records of lampreys at JBS facilities, turbine cooling water strainer collections, and information on the occurrence of lampreys in the diets of avian and piscine predators. We identified key data gaps and uncertainties that should be addressed in a juvenile lamprey passage research program. The goal of this work was to summarize information from disparate sources so that managers can use it to prioritize and guide future research and monitoring efforts related to the downstream migration of juvenile Pacific lamprey within the CRB. A common finding in all datasets was the high level of variation observed for CRB lamprey in numbers present, timing and spatial distribution. This will make developing monitoring programs to accurately characterize lamprey migrations and passage more challenging. Primary data gaps centered around our uncertainty on the numbers of juvenile and larval present in the system which affects the ability to assign risk to passage conditions and prioritize management actions. Recommendations include developing standardized monitoring methods, such as at juvenile bypass systems (JBS’s), to better document numbers and timing of lamprey migrations at dams, and use biotelemetry tracking techniques to estimate survival potentials for different migration histories.

Olfactory-mediated stream-finding behavior of migratory adult sea lamprey (Petromyzon marinus)

Science.gov (United States)

Vrieze, L.A.; Bergstedt, R.A.; Sorensen, P.W.

2011-01-01

Stream-finding behavior of adult sea lamprey (Petromyzon marinus), an anadromous fish that relies on pheromones to locate spawning streams, was documented in the vicinity of an important spawning river in the Great Lakes. Untreated and anosmic migrating sea lampreys were implanted with acoustic transmitters and then released outside the Ocqueoc River. Lampreys swam only at night and then actively. When outside of the river plume, lampreys pursued relatively straight bearings parallel to the shoreline while making frequent vertical excursions. In contrast, when within the plume, lampreys made large turns and exhibited a weak bias towards the river mouth, which one-third of them entered. The behavior of anosmic lampreys resembled that of untreated lampreys outside of the plume, except they pursued a more northerly compass bearing. To locate streams, sea lampreys appear to employ a three-phase odor-mediated strategy that involves an initial search along shorelines while casting vertically, followed by river-water-induced turning that brings them close to the river's mouth, which they then enter using rheotaxis. This novel strategy differs from that of salmonids and appears to offer this poor swimmer adaptive flexibility and suggests ways that pheromonal odors might be used to manage this invasive species.

76 FR 12129 - Lake Champlain Sea Lamprey Control Alternatives Workgroup

Science.gov (United States)

2011-03-04

... implementation of sea lamprey control techniques alternative to lampricide that are technically feasible, cost... regarding the implementation of sea lamprey control methods alternative to lampricides, to recommend...

Sea lamprey (Petromyzon marinus) parasite-host interactions in the Great Lakes

Science.gov (United States)

Bence, James R.; Bergstedt, Roger A.; Christie, Gavin C.; Cochran, Phillip A.; Ebener, Mark P.; Koonce, Joseph F.; Rutter, Michael A.; Swink, William D.

2003-01-01

Prediction of how host mortality responds to efforts to control sea lampreys (Petromyzon marinus) is central to the integrated management strategy for sea lamprey (IMSL) in the Great Lakes. A parasite-host submodel is used as part of this strategy, and this includes a type-2 multi-species functional response, a developmental response, but no numerical response. General patterns of host species and size selection are consistent with the model assumptions, but some observations appear to diverge. For example, some patterns in sea lamprey marking on hosts suggest increases in selectivity for less preferred hosts and lower host survival when preferred hosts are scarce. Nevertheless, many of the IMSL assumptions may be adequate under conditions targeted by fish community objectives. Of great concern is the possibility that the survival of young parasites (parasitic-phase sea lampreys) varies substantially among lakes or over time. Joint analysis of abundance estimates for parasites being produced in streams and returning spawners could address this. Data on sea lamprey marks is a critical source of information on sea lamprey activity and potential effects. Theory connecting observed marks to sea lamprey feeding activity and host mortality is reviewed. Uncertainties regarding healing and attachment times, the probability of hosts surviving attacks, and problems in consistent classification of marks have led to widely divergent estimates of damages caused by sea lamprey. Laboratory and field studies are recommended to provide a firmer linkage between host blood loss, host mortality, and observed marks on surviving hosts, so as to improve estimates of damage.

76 FR 43698 - Lake Champlain Sea Lamprey Control Alternatives Workgroup

Science.gov (United States)

2011-07-21

... implementation of sea lamprey control techniques alternative to lampricide that are technically feasible, cost... provide advice regarding the implementation of sea lamprey control methods alternative to lampricides, to...

Conservation challenges and research needs for Pacific lamprey in the Columbia River Basin

Science.gov (United States)

Clemens, Benjamin J.; Beamish, Richard J.; Coates, Kelly C.; Docker, Margaret F.; Dunham, Jason B.; Gray, Ann E.; Hess, Jon E.; Jolley, Jeffrey C.; Lampman, Ralph T.; McIlraith, Brian J.; Moser, Mary L.; Murauskas, Joshua G.; Noakes, David L. G.; Schaller, Howard A.; Schreck, Carl B.; Starcevich, Steven J.; Streif, Bianca; van de Wetering, Stan J.; Wade, Joy; Weitkamp, Laurie A.; Wyss, Lance A.

2017-01-01

The Pacific Lamprey Entosphenus tridentatus, an anadromous fish native to the northern Pacific Ocean and bordering freshwater habitats, has recently experienced steep declines in abundance and range contractions along the West Coast of North America. During the early 1990s, Native American tribes recognized the declining numbers of lamprey and championed their importance. In 2012, 26 entities signed a conservation agreement to coordinate and implement restoration and research for Pacific Lamprey. Regional plans have identified numerous threats, monitoring needs, and strategies to conserve and restore Pacific Lamprey during their freshwater life stages. Prime among these are needs to improve lamprey passage, restore freshwater habitats, educate stakeholders, and implement lamprey-specific research and management protocols. Key unknowns include range-wide trends in status, population dynamics, population delineation, limiting factors, and marine influences. We synthesize these key unknowns, with a focus on the freshwater life stages of lamprey in the Columbia River basin.

Bioinformatic Characterization of Genes and Proteins Involved in Blood Clotting in Lampreys

OpenAIRE

Doolittle, RF

2015-01-01

© 2015, Springer Science+Business Media New York. Lampreys and hagfish are the earliest diverging of extant vertebrates and are obvious targets for investigating the origins of complex biochemical systems found in mammals. Currently, the simplest approach for such inquiries is to search for the presence of relevant genes in whole genome sequence (WGS) assemblies. Unhappily, in the past a high-quality complete genome sequence has not been available for either lampreys or hagfish, precluding th...

Spatial and Ontogenetic Variation in Mercury in Lake Superior Basin Sea Lamprey (Petromyzon marinus).

Science.gov (United States)

Moses, Sara K; Polkinghorne, Christine N; Mattes, William P; Beesley, Kimberly M

2018-01-01

Mercury concentrations were measured in eggs, larvae, and adult spawning-phase sea lampreys (Petromyzon marinus) collected in tributaries of Lake Superior to investigate spatial and ontogenetic variation. There were significant differences in mercury concentrations between all three life stages, with levels highest in adults (mean = 3.01 µg/g), followed by eggs (mean = 0.942 µg/g), and lowest in larvae (mean = 0.455 µg/g). There were no significant differences in mercury concentrations by location for any life stage or by sex in adults. Mercury was not correlated with adult or larval lamprey length or mass. Mercury levels in adult lampreys exceeded U.S. and Canadian federal guidelines for human consumption. Mercury concentrations in all life stages exceeded criteria for the protection of piscivorous wildlife, posing a threat to local fish, birds, and mammals. High mercury levels in adult lampreys combined with their semelparous life history make them a potential source of lake-derived mercury to spawning streams.

PCB concentrations and activity of sea lamprey Petromyzon marinus vary by sex

Science.gov (United States)

Madenjian, Charles P.; Johnson, Nicholas S.; Binder, Thomas R.; Rediske, Richard R.; O'Keefe, James P.

2013-01-01

We determined the polychlorinated biphenyl (PCB) concentrations of 40 male and 40 female adult sea lampreys Petromyzon marinus captured in the Cheboygan River, a tributary to Lake Huron, during May 2011. In addition, we performed a laboratory experiment using passive integrated transponder tags to determine whether male adult sea lampreys were more active than female adult sea lampreys. Sex had a significant effect on PCB concentration, and PCB concentration at a given level of sea lamprey condition was approximately 25 % greater in males than in females. Adjusting for the difference in condition between the sexes, males averaged a 17 % greater PCB concentration compared with females. Results from the laboratory experiment indicated that males were significantly more active than females. The observed sex difference in PCB concentrations was not due to female sea lampreys releasing eggs at spawning because the sea lamprey is semelparous, and we caught the sea lampreys before spawning. Rather, we attributed the sex difference in PCB concentrations to a greater rate of energy expenditure in males compared with females. We proposed that this greater rate of energy expenditure was likely due to greater activity. Our laboratory experiment results supported this hypothesis. A greater resting metabolic rate may also have contributed to a greater rate of energy expenditure. Our findings should eventually be applicable toward improving control of sea lamprey, a pest responsible for considerable damage to fisheries in lakes where it is not native.

Evaluate Status of Pacific Lamprey in the Clearwater River Drainage, Idaho: Annual Report 2001.

Energy Technology Data Exchange (ETDEWEB)

Cochnauer, Tim; Claire, Christopher

2002-12-01

Recent decline of Pacific lamprey Lampetra tridentata adult migrants to the Snake River drainage has focused attention on the species. Adult Pacific lamprey counted passing Ice Harbor Dam fishway averaged 18,158 during 1962-69 and 361 during 1993-2000. Human resource manipulations in the Snake River and Clearwater River drainages have altered ecosystem habitat in the last 120 years, likely impacting the productive potential of Pacific lamprey habitat. Timber harvest, stream impoundment, road construction, grazing, mining, and community development have dominated habitat alteration in the Clearwater River system and Snake River corridor. Hydroelectric projects in the Snake River corridor impact juvenile/larval Pacific lamprey outmigrants and returning adults. Juvenile and larval lamprey outmigrants potentially pass through turbines, turbine bypass/collection systems, and over spillway structures at the four lower Snake River hydroelectric dams. Clearwater River drainage hydroelectric facilities have impacted Pacific lamprey populations to an unknown degree. The Pacific Power and Light Dam on the Clearwater River in Lewiston, Idaho, restricted chinook salmon Oncorhynchus tshawytscha passage in the 1927-1940 period, altering the migration route of outmigrating Pacific lamprey juveniles/larvae and upstream adult migrants (1927-1972). Dworshak Dam, completed in 1972, eliminated Pacific lamprey spawning and rearing in the North Fork Clearwater River drainage. Construction of the Harpster hydroelectric dam on the South Fork of the Clearwater River resulted in obstructed fish passage 1949-1963. Through Bonneville Power Administration support, the Idaho Department of Fish and Game continued investigation into the status of Pacific lamprey populations in Idaho's Clearwater River drainage in 2001. Trapping, electrofishing, and spawning ground redd surveys were used to determine Pacific lamprey distribution, life history strategies, and habitat requirements in the South

Evidence for a receiver bias underlying female preference for a male mating pheromone in sea lamprey

Science.gov (United States)

Buchinger, Tyler J.; Wang, Huiyong; Li, Weiming; Johnson, Nicholas S.

2013-01-01

Receiver bias models suggest that a male sexual signal became exaggerated to match a pre-existing sensory, perceptual or cognitive disposition of the female. Accordingly, these models predict that females of related taxa possessing the ancestral state of signalling evolved preference for the male trait in a non-sexual context. We postulated that female preference for the male-released bile alcohol mating pheromone, 3 keto petromyzonol sulfate (3kPZS), of the sea lamprey (Petromyzon marinus) evolved as a result of a receiver bias. In particular, we propose that migratory silver lamprey (Ichthyomyzon unicuspis), a basal member of the Petromyzontidae, evolved a preference for 3kPZS released by stream-resident larvae as a means of identifying productive habitat for offspring. Larval silver lamprey released 3kPZS at rates sufficient to be detected by migratory lampreys. Females responded to 3kPZS by exhibiting upstream movement behaviours relevant in a migratory context, but did not exhibit proximate behaviours important to mate search and spawning. Male silver lamprey did not release 3kPZS at rates sufficient to be detected by females in natural high-volume stream environments. We infer that female silver lamprey cue onto 3kPZS excreted by stream-resident larvae as a mechanism to locate habitat conducive to offspring survival and that males do not signal with 3kPZS. We suggest that this female preference for a male signal in a non-sexual context represents a bias leading to the sexual signalling observed in sea lamprey.

Boll weevil eradication: a model for sea lamprey control?

Science.gov (United States)

Smith, James W.; Swink, William D.

2003-01-01

Invasions of boll weevil (Anthonomus grandis) into the United States and sea lamprey (Petromyzon marinus) into the Great Lakes were similar in many ways. Important species (American cotton, Gossypium hirsutum, and lake trout, Salvelinus namaycush) and the industries they supported were negatively affected. Initial control efforts were unsuccessful until pesticides and application technologies were developed. For boll weevils, controls relying on pesticides evolved into an integrated program that included recommended farming practices and poisoned baits. However, the discovery of a boll weevil sex pheromone in 1964 allowed adoption of an ongoing program of eradication. Despite opposition over concept and cost, insecticides, pheromone traps, poisoned baits, and approved farming practices were used to eradicate boll weevils from Virginia, North Carolina, South Carolina, Georgia, Florida, and Alabama by 1999. Using the working back approach along the path of the original invasion, eradication was nearly completed by 2002 in Mississippi and eradication programs were underway in Arkansas, Tennessee, Oklahoma, Louisiana, and parts of Texas. Insecticide use for cotton production decreased 50 to 90%, and cotton yields and farm income increased an average of 78 kg/ha and $190 U.S./ha in areas where boll weevils were eradicated. For sea lampreys, integrated management uses lampricides, barriers to migration, trapping, and release of sterilized males. Although sea lamprey eradication is not currently feasible, recent research on larval and sex pheromones might provide the tools to make it possible. A successful eradication program for sea lampreys starting in Lake Superior and expanding to the lower Great Lakes would ultimately provide huge ecological and economic benefits by eliminating lampricide applications, removing barriers that block teleost fishes, and facilitating the recovery of lake trout. Should the opportunity arise, the concept of sea lamprey eradication should

Hsp90 and hepatobiliary transformation during sea lamprey metamorphosis.

Science.gov (United States)

Chung-Davidson, Yu-Wen; Yeh, Chu-Yin; Bussy, Ugo; Li, Ke; Davidson, Peter J; Nanlohy, Kaben G; Brown, C Titus; Whyard, Steven; Li, Weiming

2015-12-01

Biliary atresia (BA) is a human infant disease with inflammatory fibrous obstructions in the bile ducts and is the most common cause for pediatric liver transplantation. In contrast, the sea lamprey undergoes developmental BA with transient cholestasis and fibrosis during metamorphosis, but emerges as a fecund adult. Therefore, sea lamprey liver metamorphosis may serve as an etiological model for human BA and provide pivotal information for hepatobiliary transformation and possible therapeutics. We hypothesized that liver metamorphosis in sea lamprey is due to transcriptional reprogramming that dictates cellular remodeling during metamorphosis. We determined global gene expressions in liver at several metamorphic landmark stages by integrating mRNA-Seq and gene ontology analyses, and validated the results with real-time quantitative PCR, histological and immunohistochemical staining. These analyses revealed that gene expressions of protein folding chaperones, membrane transporters and extracellular matrices were altered and shifted during liver metamorphosis. HSP90, important in protein folding and invertebrate metamorphosis, was identified as a candidate key factor during liver metamorphosis in sea lamprey. Blocking HSP90 with geldanamycin facilitated liver metamorphosis and decreased the gene expressions of the rate limiting enzyme for cholesterol biosynthesis, HMGCoA reductase (hmgcr), and bile acid biosynthesis, cyp7a1. Injection of hsp90 siRNA for 4 days altered gene expressions of met, hmgcr, cyp27a1, and slc10a1. Bile acid concentrations were increased while bile duct and gall bladder degeneration was facilitated and synchronized after hsp90 siRNA injection. HSP90 appears to play crucial roles in hepatobiliary transformation during sea lamprey metamorphosis. Sea lamprey is a useful animal model to study postembryonic development and mechanisms for hsp90-induced hepatobiliary transformation.

Daytime avoidance of chemosensory alarm cues by adult sea lamprey (Petromyzon marinus)

Science.gov (United States)

Di Rocco, Richard; Belanger, Cowan; Imre, István; Brown, Grant; Johnson, Nicholas S.

2014-01-01

Sea lamprey (Petromyzon marinus) avoid damage-released and predator chemosensory cues at night, but their response to these cues during the day is unknown. Here, we explored (i) whether sea lamprey avoid these cues during the day and (ii) the effect of water temperature on the avoidance of chemosensory alarm cues in two diurnal laboratory experiments. We hypothesized that daytime activity would be temperature-dependent and that only sea lamprey vulnerable to predation (i.e., not hiding) would behaviourally respond to chemosensory alarm cues. Ten groups of ten sea lamprey were exposed to one of a variety of potential chemosensory cues. The experiments were conducted over a range of temperatures to quantify the effect of temperature on avoidance behaviour. Consistent with our hypothesis, a higher proportion of animals were active during daytime as water temperature increased. Moving sea lamprey showed an avoidance response to 2-phenylethylamine (a compound found in mammalian urine) and human saliva once water temperatures had risen to mean (±SD) = 13.7 (±1.4) °C. Resting and hiding sea lamprey did not show an avoidance response to any of the experimental stimuli.

Population genomics of Pacific lamprey: adaptive variation in a highly dispersive species.

Science.gov (United States)

Hess, Jon E; Campbell, Nathan R; Close, David A; Docker, Margaret F; Narum, Shawn R

2013-06-01

Unlike most anadromous fishes that have evolved strict homing behaviour, Pacific lamprey (Entosphenus tridentatus) seem to lack philopatry as evidenced by minimal population structure across the species range. Yet unexplained findings of within-region population genetic heterogeneity coupled with the morphological and behavioural diversity described for the species suggest that adaptive genetic variation underlying fitness traits may be responsible. We employed restriction site-associated DNA sequencing to genotype 4439 quality filtered single nucleotide polymorphism (SNP) loci for 518 individuals collected across a broad geographical area including British Columbia, Washington, Oregon and California. A subset of putatively neutral markers (N = 4068) identified a significant amount of variation among three broad populations: northern British Columbia, Columbia River/southern coast and 'dwarf' adults (F(CT) = 0.02, P ≪ 0.001). Additionally, 162 SNPs were identified as adaptive through outlier tests, and inclusion of these markers revealed a signal of adaptive variation related to geography and life history. The majority of the 162 adaptive SNPs were not independent and formed four groups of linked loci. Analyses with matsam software found that 42 of these outlier SNPs were significantly associated with geography, run timing and dwarf life history, and 27 of these 42 SNPs aligned with known genes or highly conserved genomic regions using the genome browser available for sea lamprey. This study provides both neutral and adaptive context for observed genetic divergence among collections and thus reconciles previous findings of population genetic heterogeneity within a species that displays extensive gene flow. © 2012 John Wiley & Sons Ltd.

Spatial mismatch between sea lamprey behaviour and trap location explains low success at trapping for control

Science.gov (United States)

Rous, Andrew M.; McLean, Adrienne R.; Barber, Jessica; Bravener, Gale; Castro-Santos, Theodore; Holbrook, Christopher M.; Imre, Istvan; Pratt, Thomas C.; McLaughlin, Robert L.

2017-01-01

Crucial to the management of invasive species is understanding space use and the environmental features affecting space use. Improved understanding of space use by invasive sea lamprey (Petromyzon marinus) could help researchers discern why trap success in large rivers is lower than needed for effective control. We tested whether manipulating discharge nightly could increase trap success at a hydroelectric generating station on the St. Marys River. We quantified numbers of acoustically tagged sea lampreys migrating up to, and their space use at, the hydroelectric generating station. In 2011 and 2012, 78% and 68%, respectively, of tagged sea lampreys reached the generating station. Sea lampreys were active along the face, but more likely to occur at the bottom and away from the traps near the surface, especially when discharge was high. Our findings suggest that a low probability of encountering traps was due to spatial (vertical) mismatch between space use by sea lamprey and trap locations and that increasing discharge did not alter space use in ways that increased trap encounter. Understanding space use by invasive species can help managers assess the efficacy of trapping and ways of improving trapping success.

Pacific Lamprey Research and Restoration Project : Annual Report 2000.

Energy Technology Data Exchange (ETDEWEB)

Close, David A.

2002-11-01

This report summarizes results of research activities conducted in 1999-2000. The findings in these chapters represent the efforts of the Confederated Tribes of the Umatilla Indian Reservation (CTUIR) and collaborative efforts among other researchers working on Pacific lampreys (Lampetra tridentata) under this project. The findings in these chapters will help management and recovery of Pacific lampreys in the Columbia River Basin.

Bioenergetics models to estimate numbers of larval lampreys consumed by smallmouth bass in Elk Creek, Oregon

Science.gov (United States)

Schultz, Luke; Heck, Michael; Kowalski, Brandon M; Eagles-Smith, Collin A.; Coates, Kelly C.; Dunham, Jason B.

2017-01-01

Nonnative fishes have been increasingly implicated in the decline of native fishes in the Pacific Northwest. Smallmouth Bass Micropterus dolomieu were introduced into the Umpqua River in southwest Oregon in the early 1960s. The spread of Smallmouth Bass throughout the basin coincided with a decline in counts of upstream-migrating Pacific Lampreys Entosphenus tridentatus. This suggested the potential for ecological interactions between Smallmouth Bass and Pacific Lampreys, as well as freshwater-resident Western Brook Lampreys Lampetra richardsoni. To evaluate the potential effects of Smallmouth Bass on lampreys, we sampled diets of Smallmouth Bass and used bioenergetics models to estimate consumption of larval lampreys in a segment of Elk Creek, a tributary to the lower Umpqua River. We captured 303 unique Smallmouth Bass (mean: 197 mm and 136 g) via angling in July and September. We combined information on Smallmouth Bass diet and energy density with other variables (temperature, body size, growth, prey energy density) in a bioenergetics model to estimate consumption of larval lampreys. Larval lampreys were found in 6.2% of diet samples, and model estimates indicated that the Smallmouth Bass we captured consumed 925 larval lampreys in this 2-month study period. When extrapolated to a population estimate of Smallmouth Bass in this segment, we estimated 1,911 larval lampreys were consumed between July and September. Although the precision of these estimates was low, this magnitude of consumption suggests that Smallmouth Bass may negatively affect larval lamprey populations.

Shifting patterns of nitrogen excretion and amino acid catabolism capacity during the life cycle of the sea lamprey (Petromyzon marinus).

Science.gov (United States)

Wilkie, Michael P; Claude, Jaime F; Cockshutt, Amanda; Holmes, John A; Wang, Yuxiang S; Youson, John H; Walsh, Patrick J

2006-01-01

The jawless fish, the sea lamprey (Petromyzon marinus), spends part of its life as a burrow-dwelling, suspension-feeding larva (ammocoete) before undergoing a metamorphosis into a free swimming, parasitic juvenile that feeds on the blood of fishes. We predicted that animals in this juvenile, parasitic stage have a great capacity for catabolizing amino acids when large quantities of protein-rich blood are ingested. The sixfold to 20-fold greater ammonia excretion rates (J(Amm)) in postmetamorphic (nonfeeding) and parasitic lampreys compared with ammocoetes suggested that basal rates of amino acid catabolism increased following metamorphosis. This was likely due to a greater basal amino acid catabolizing capacity in which there was a sixfold higher hepatic glutamate dehydrogenase (GDH) activity in parasitic lampreys compared with ammocoetes. Immunoblotting also revealed that GDH quantity was 10-fold and threefold greater in parasitic lampreys than in ammocoetes and upstream migrant lampreys, respectively. Higher hepatic alanine and aspartate aminotransferase activities in the parasitic lampreys also suggested an enhanced amino acid catabolizing capacity in this life stage. In contrast to parasitic lampreys, the twofold larger free amino acid pool in the muscle of upstream migrant lampreys confirmed that this period of natural starvation is accompanied by a prominent proteolysis. Carbamoyl phosphate synthetase III was detected at low levels in the liver of parasitic and upstream migrant lampreys, but there was no evidence of extrahepatic (muscle, intestine) urea production via the ornithine urea cycle. However, detection of arginase activity and high concentrations of arginine in the liver at all life stages examined infers that arginine hydrolysis is an important source of urea. We conclude that metamorphosis is accompanied by a metabolic reorganization that increases the capacity of parasitic sea lampreys to catabolize intermittently large amino acid loads arising

A synthesized mating pheromone component increases adult sea lamprey (Petromyzon marinus) trap capture in management scenarios

Science.gov (United States)

Johnson, Nicholas S.; Siefkes, Michael J.; Wagner, C. Michael; Dawson, Heather; Wang, Huiyong; Steeves, Todd; Twohey, Michael; Li, Weiming

2013-01-01

Application of chemical cues to manipulate adult sea lamprey (Petromyzon marinus) behavior is among the options considered for new sea lamprey control techniques in the Laurentian Great Lakes. A male mating pheromone component, 7a,12a,24-trihydroxy-3-one-5a-cholan-24-sulfate (3kPZS), lures ovulated female sea lamprey upstream into baited traps in experimental contexts with no odorant competition. A critical knowledge gap is whether this single pheromone component influences adult sea lamprey behavior in management contexts containing free-ranging sea lampreys. A solution of 3kPZS to reach a final in-stream concentration of 10-12 mol·L-1 was applied to eight Michigan streams at existing sea lamprey traps over 3 years, and catch rates were compared between paired 3kPZS-baited and unbaited traps. 3kPZS-baited traps captured significantly more sexually immature and mature sea lampreys, and overall yearly trapping efficiency within a stream averaged 10% higher during years when 3kPZS was applied. Video analysis of a trap funnel showed that the likelihood of sea lamprey trap entry after trap encounter was higher when the trap was 3kPZS baited. Our approach serves as a model for the development of similar control tools for sea lamprey and other aquatic invaders.

Use of chemosensory cues as repellents for sea lamprey: Potential directions for population management

Science.gov (United States)

Imre, I.; Brown, G.E.; Bergstedt, R.A.; McDonald, R.

2010-01-01

Sea lamprey invaded the Great Lakes in the early 20th century and caused an abrupt decline in the population densities of several native fish species. The integrated management of this invasive species is composed of chemical (lampricide) applications, low-head barrier dams, adult trapping and sterile male release. Recently, there has been an increased emphasis on the development of control methods alternative to lampricide applications. We propose as an alternative-control method the use of chemosensory cues as repellents for sea lamprey population management. Based on the available evidence at this time, we suggest that injury-released chemical alarm cues show promise as repellents for sea lamprey and further research should be directed at determining whether sea lamprey show an avoidance response to these types of chemosensory cues. From a management perspective, these chemosensory cues could be used to restrict sea lamprey access to spawning grounds. Repellents could also be used together with attractants like sex pheromones to manipulate sea lamprey behavior, similar to the "push-pull" strategies utilized with insect pests. ?? 2010 Elsevier B.V.

Determining Lamprey Species Composition, Larval Distribution and Adult Abundance in the Deschutes River Subbasin, Oregon ; 2007 Annual Report.

Energy Technology Data Exchange (ETDEWEB)

Fox, Matt; Graham, Jennifer C. [Department of Natural Resources, Confederated Tribes of the Warm Springs Reservation, Oregon

2009-06-26

We will report results of an ongoing project in the Deschutes River Subbasin to describe Pacific lamprey (Lampetra tridentata) life history. Project objectives were to determine adult lamprey escapement from Sherars Falls located at Rkm 70.4 and determine lamprey focal spawning areas, spawn timing and habitat through radio telemetry. A mark-recapture study and tribal creel was conducted to determine adult escapement. Lamprey were radio tagged and are currently being mobile, aerial and fixed site tracked to describe spawning. Adult lamprey were collected at Sherars Falls using a long-handled dip net from June-September 2007. The fate of lamprey collected at Sherars Falls was determined based on girth measurements. Fish measuring less than 10.5 cm received two markings for the mark-recapture estimation while those measuring 10.5 cm or greater were implanted with radio transmitters. Two-hundred and nine lamprey were marked during first event sampling, 2,501 lamprey inspected for marks and 64 recaptured during second event sampling. We estimate lamprey abundance to be 8,083 (6,352-10,279) with a relative precision of 19.8. Tribal harvest was 2,303 +/- 88. Escapement was estimated at 5,780 adult lamprey. Thirty-six lamprey received radio transmitters. Lamprey were transported upstream 6.3 Rkm for surgery, held to recover from anesthesia and released. Mobile tracking efforts started mid-July 2007 and are on-going. To date 35 of the 36 lamprey have been detected. Upon release, extensive ground-based tracking was conducted until fish became dormant in mid-October. Since, fixed site downloading and tracking have occurred weekly on the mainstem Deschutes River. Majority of lamprey (88%) are holding in the mainstem Deschutes River. Three lamprey moved upstream more than 70 Rkms into westside tributaries from August-December. Three moved approximately 18 Rkms downstream of the release site. Tracking will continue through the spawning season when redd characteristics will be

Survival and metamorphosis of larval sea lamprey (Petromyzon marinus) residing in Lakes Michigan and Huron near river mouths

Science.gov (United States)

Johnson, Nicholas S.; Brenden, Travis O.; Swink, William D.; Lipps, Mathew A.

2016-01-01

Although population demographics of larval lampreys in streams have been studied extensively, demographics in lake environments have not. Here, we estimated survival and rates of metamorphosis for larval sea lamprey (Petromyzon marinus) populations residing in the Great Lakes near river mouths (hereafter termed lentic areas). Tagged larvae were stocked and a Bayesian multi-state tag-recovery model was used to investigate population parameters associated with tag recovery, including survival and metamorphosis probabilities. Compared to previous studies of larvae in streams, larval growth in lentic areas was substantially slower (Brody growth coefficient = 0.00132; estimate based on the recovery of six tagged larvae), survival was slightly greater (annual survival = 63%), and the length at which 50% of the larvae would be expected to metamorphose was substantially shorter (126 mm). Stochastic simulations were used to estimate the production of parasitic stage (juvenile) sea lamprey from a hypothetical population of larvae in a lentic environment. Production of juvenile sea lamprey was substantial because, even though larval growth in these environments was slow relative to stream environments, survival was high and length at metamorphosis was less. However, estimated production of juvenile sea lamprey was less for the lentic environment than for similar simulations for river environments where larvae grew faster. In circumstances where the cost to kill a larva with lampricide was equal and control funds are limited, sea lamprey control effort may be best directed toward larvae in streams with fast-growing larvae, because stream-produced larvae will most likely contribute to juvenile sea lamprey populations.

Low reproductive isolation and highly variable levels of gene flow reveal limited progress towards speciation between European river and brook lampreys.

Science.gov (United States)

Rougemont, Q; Gaigher, A; Lasne, E; Côte, J; Coke, M; Besnard, A-L; Launey, S; Evanno, G

2015-12-01

Ecologically based divergent selection is a factor that could drive reproductive isolation even in the presence of gene flow. Population pairs arrayed along a continuum of divergence provide a good opportunity to address this issue. Here, we used a combination of mating trials, experimental crosses and population genetic analyses to investigate the evolution of reproductive isolation between two closely related species of lampreys with distinct life histories. We used microsatellite markers to genotype over 1000 individuals of the migratory parasitic river lamprey (Lampetra fluviatilis) and freshwater-resident nonparasitic brook lamprey (Lampetra planeri) distributed in 10 sympatric and parapatric population pairs in France. Mating trials, parentage analyses and artificial fertilizations demonstrated a low level of reproductive isolation between species even though size-assortative mating may contribute to isolation. Most parapatric population pairs were strongly differentiated due to the joint effects of geographic distance and barriers to migration. In contrast, we found variable levels of gene flow between sympatric populations ranging from panmixia to moderate differentiation, which indicates a gradient of divergence with some population pairs that may correspond to alternative morphs or ecotypes of a single species and others that remain partially isolated. Ecologically based divergent selection may explain these variable levels of divergence among sympatric population pairs, but incomplete genome swamping following secondary contact could have also played a role. Overall, this study illustrates how highly differentiated phenotypes can be maintained despite high levels of gene flow that limit the progress towards speciation. © 2015 European Society For Evolutionary Biology. Journal of Evolutionary Biology © 2015 European Society For Evolutionary Biology.

Dopamine and the Brainstem Locomotor Networks: From Lamprey to Human

Directory of Open Access Journals (Sweden)

Dimitri Ryczko

2017-05-01

Full Text Available In vertebrates, dopamine neurons are classically known to modulate locomotion via their ascending projections to the basal ganglia that project to brainstem locomotor networks. An increased dopaminergic tone is associated with increase in locomotor activity. In pathological conditions where dopamine cells are lost, such as in Parkinson's disease, locomotor deficits are traditionally associated with the reduced ascending dopaminergic input to the basal ganglia. However, a descending dopaminergic pathway originating from the substantia nigra pars compacta was recently discovered. It innervates the mesencephalic locomotor region (MLR from basal vertebrates to mammals. This pathway was shown to increase locomotor output in lampreys, and could very well play an important role in mammals. Here, we provide a detailed account on the newly found dopaminergic pathway in lamprey, salamander, rat, monkey, and human. In lampreys and salamanders, dopamine release in the MLR is associated with the activation of reticulospinal neurons that carry the locomotor command to the spinal cord. Dopamine release in the MLR potentiates locomotor movements through a D1-receptor mechanism in lampreys. In rats, stimulation of the substantia nigra pars compacta elicited dopamine release in the pedunculopontine nucleus, a known part of the MLR. In a monkey model of Parkinson's disease, a reduced dopaminergic innervation of the brainstem locomotor networks was reported. Dopaminergic fibers are also present in human pedunculopontine nucleus. We discuss the conserved locomotor role of this pathway from lamprey to mammals, and the hypothesis that this pathway could play a role in the locomotor deficits reported in Parkinson's disease.

Electrical guidance efficiency of downstream-migrating juvenile Sea Lamprey decreases with increasing water velocity

Science.gov (United States)

Miehls, Scott M.; Johnson, Nicholas; Haro, Alexander

2017-01-01

We tested the efficacy of a vertically oriented field of pulsed direct current (VEPDC) created by an array of vertical electrodes for guiding downstream-moving juvenile Sea Lampreys Petromyzon marinus to a bypass channel in an artificial flume at water velocities of 10–50 cm/s. Sea Lampreys were more likely to be captured in the bypass channel than in other sections of the flume regardless of electric field status (on or off) or water velocity. Additionally, Sea Lampreys were more likely to be captured in the bypass channel when the VEPDC was active; however, an interaction between the effects of VEPDC and water velocity was observed, as the likelihood of capture decreased with increases in water velocity. The distribution of Sea Lampreys shifted from right to left across the width of the flume toward the bypass channel when the VEPDC was active at water velocities less than 25 cm/s. The VEPDC appeared to have no effect on Sea Lamprey distribution in the flume at water velocities greater than 25 cm/s. We also conducted separate tests to determine the threshold at which Sea Lampreys would become paralyzed. Individuals were paralyzed at a mean power density of 37.0 µW/cm3. Future research should investigate the ability of juvenile Sea Lampreys to detect electric fields and their specific behavioral responses to electric field characteristics so as to optimize the use of this technology as a nonphysical guidance tool across variable water velocities.

Distribution and seasonal differences in Pacific Lamprey and Lampetra spp eDNA across 18 Puget Sound watersheds

Science.gov (United States)

Ostberg, Carl O.; Chase, Dorothy M.; Hayes, Michael C.; Duda, Jeffrey J.

2018-01-01

Lampreys have a worldwide distribution, are functionally important to ecological communities and serve significant roles in many cultures. In Pacific coast drainages of North America, lamprey populations have suffered large declines. However, lamprey population status and trends within many areas of this region are unknown and such information is needed for advancing conservation goals. We developed two quantitative PCR-based, aquatic environmental DNA (eDNA) assays for detection of Pacific Lamprey (Entosphenus tridentatus) and Lampetra spp, using locked nucleic acids (LNAs) in the probe design. We used these assays to characterize the spatial distribution of lamprey in 18 watersheds of Puget Sound, Washington, by collecting water samples in spring and fall. Pacific Lamprey and Lampetraspp were each detected in 14 watersheds and co-occurred in 10 watersheds. Lamprey eDNA detection rates were much higher in spring compared to fall. Specifically, the Pacific Lamprey eDNA detection rate was 3.5 times higher in spring and the Lampetra spp eDNA detection rate was 1.5 times higher in spring even though larval lamprey are present in streams year-round. This significant finding highlights the importance of seasonality on eDNA detection. Higher stream discharge in the fall likely contributed to reduced eDNA detection rates, although seasonal life history events may have also contributed. These eDNA assays differentiate Pacific Lamprey and Lampetra spp across much of their range along the west coast of North America. Sequence analysis indicates the Pacific Lamprey assay also targets other Entosphenus spp and indicates the Lampetra spp assay may have limited or no capability of detecting Lampetra in some locations south of the Columbia River Basin. Nevertheless, these assays will serve as a valuable tool for resource managers and have direct application to lamprey conservation efforts, such as mapping species distributions, occupancy modeling, and monitoring

Discovery of fossil lamprey larva from the Lower Cretaceous reveals its three-phased life cycle.

Science.gov (United States)

Chang, Mee-mann; Wu, Feixiang; Miao, Desui; Zhang, Jiangyong

2014-10-28

Lampreys are one of the two surviving jawless vertebrate groups and one of a few vertebrate groups with the best exemplified metamorphosis during their life cycle, which consists of a long-lasting larval stage, a peculiar metamorphosis, and a relatively short adulthood with a markedly different anatomy. Although the fossil records have revealed that many general features of extant lamprey adults were already formed by the Late Devonian (ca. 360 Ma), little is known about the life cycle of the fossil lampreys because of the lack of fossilized lamprey larvae or transformers. Here we report the first to our knowledge discovery of exceptionally preserved premetamorphic and metamorphosing larvae of the fossil lamprey Mesomyzon mengae from the Lower Cretaceous of Inner Mongolia, China. These fossil ammocoetes look surprisingly modern in having an eel-like body with tiny eyes, oral hood and lower lip, anteriorly positioned branchial region, and a continuous dorsal skin fin fold and in sharing a similar feeding habit, as judged from the detritus left in the gut. In contrast, the larger metamorphosing individuals have slightly enlarged eyes relative to large otic capsules, thickened oral hood or pointed snout, and discernable radials but still anteriorly extended branchial area and lack a suctorial oral disk, which characterize the early stages of the metamorphosis of extant lampreys. Our discovery not only documents the larval conditions of fossil lampreys but also indicates the three-phased life cycle in lampreys emerged essentially in their present mode no later than the Early Cretaceous.

Quantification of Oxidized and Unsaturated Bile Alcohols in Sea Lamprey Tissues by Ultra-High Performance Liquid Chromatography-Tandem Mass Spectrometry

Directory of Open Access Journals (Sweden)

Ke Li

2016-08-01

Full Text Available A sensitive and reliable method was developed and validated for the determination of unsaturated bile alcohols in sea lamprey tissues using liquid-liquid extraction and ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS. The liver, kidney, and intestine samples were extracted with acetonitrile and defatted by n-hexane. Gradient UHPLC separation was performed using an Acquity BEH C18 column with a mobile phase of water and methanol containing 20 mM triethylamine. Multiple reaction monitoring modes of precursor-product ion transitions for each analyte was used. This method displayed good linearity, with correlation coefficients greater than 0.99, and was validated. Precision and accuracy (RSD % were in the range of 0.31%–5.28%, while mean recoveries were between 84.3%–96.3%. With this technique, sea lamprey tissue samples were analyzed for unsaturated bile alcohol analytes. This method is practical and particularly suitable for widespread putative pheromone residue analysis.

Gene expression during ovarian differentiation in parasitic and non-parasitic lampreys: implications for fecundity and life history types.

Science.gov (United States)

Spice, Erin K; Whyard, Steven; Docker, Margaret F

2014-11-01

Lampreys diverged from the jawed vertebrate lineage approximately 500million years ago. Lampreys undergo sex differentiation much later than most other vertebrates, and ovarian differentiation occurs several years before testicular differentiation. The genetic basis of lamprey sex differentiation is of particular interest both because of the phylogenetic importance of lampreys and because of their unusual pattern of sex differentiation. As well, differences between parasitic and non-parasitic lampreys may first become evident at ovarian differentiation. However, nothing is known about the genetic basis of ovarian differentiation in lampreys. This study examined potential differences in gene expression before, during, and after ovarian differentiation in parasitic chestnut lamprey Ichthyomyzon castaneus and non-parasitic northern brook lamprey Ichthyomyzonfossor. Eight target genes (17β-hydroxysteroid dehydrogenase, germ cell-less, estrogen receptor β, insulin-like growth factor 1 receptor, daz-associated protein 1, cytochrome c oxidase subunit III, Wilms' tumour suppressor protein 1, and dehydrocholesterol reductase 7) were examined. Northern brook lamprey displayed higher expression of cytochrome c oxidase subunit III, whereas chestnut lamprey displayed higher expression of insulin-like growth factor 1 receptor; these genes may be involved in apoptosis and oocyte growth, respectively. Presumptive male larvae had higher expression of Wilms' tumour suppressor protein 1, which may be involved in the undifferentiated gonad and/or later testicular development. Differentiated females had higher expression of 17β hydroxysteroid dehydrogenase and daz-associated protein 1, which may be involved in female development. This study is the first to identify genes that may be involved in ovarian differentiation and fecundity in lampreys. Copyright © 2014 Elsevier Inc. All rights reserved.

Development and Functional Organization of the Cranial Nerves in Lampreys.

Science.gov (United States)

Pombal, Manuel A; Megías, Manuel

2018-04-16

Lampreys, together with hagfishes, are the only extant representatives of the oldest branch of vertebrates, the agnathans, which are the sister group of gnathostomes; therefore, studies on these animals are of great evolutionary significance. Lampreys exhibit a particular life cycle with remarkable changes in their behavior, concomitant, in part, with important modifications in the head and its musculature, which might influence the development of the cranial nerves. In this context, some cranial nerves such as the optic nerve and the ocular motor nerves, which develop slowly during an extremely long larval period lasting more than five years, have been more thoroughly investigated; however, much less experimental information is available about others, such as the facial or the hypoglossal nerves. In addition, the possible existence of a "true" accessory nerve in these animals is still a matter of conjecture. Although growing in last decades, investigations on the physiology of the lamprey cranial nerves is scanty. This review focuses on past and recent findings that have contributed to characterize the anatomical organization of the cranial nerves in lampreys, including their components and nuclei, and their relations in the brain; in addition, comments on their development and functional role are also included. Anat Rec, 2018. © 2018 Wiley Periodicals, Inc. © 2018 Wiley Periodicals, Inc.

Odor-conditioned rheotaxis of the sea lamprey: modeling, analysis and validation

International Nuclear Information System (INIS)

Choi, Jongeun; Jeon, Soo; Johnson, Nicholas S; Brant, Cory O; Li, Weiming

2013-01-01

Mechanisms for orienting toward and locating an odor source are sought in both biology and engineering. Chemical ecology studies have demonstrated that adult female sea lamprey show rheotaxis in response to a male pheromone with dichotomous outcomes: sexually mature females locate the source of the pheromone whereas immature females swim by the source and continue moving upstream. Here we introduce a simple switching mechanism modeled after odor-conditioned rheotaxis for the sea lamprey as they search for the source of a pheromone in a one-dimensional riverine environment. In this strategy, the females move upstream only if they detect that the pheromone concentration is higher than a threshold value and drifts down (by turning off control action to save energy) otherwise. In addition, we propose various uncertainty models such as measurement noise, actuator disturbance, and a probabilistic model of a concentration field in turbulent flow. Based on the proposed model with uncertainties, a convergence analysis showed that with this simplistic switching mechanism, the lamprey converges to the source location on average in spite of all such uncertainties. Furthermore, a slightly modified model and its extensive simulation results explain the behaviors of immature female lamprey near the source location. (paper)

Re-examination of sea lamprey control policies for the St. Marys River: Completion of an adaptive management cycle

Science.gov (United States)

Jones, Michael L.; Brenden, Travis O.; Irwin, Brian J.

2015-01-01

The St. Marys River (SMR) historically has been a major producer of sea lampreys (Petromyzon marinus) in the Laurentian Great Lakes. In the early 2000s, a decision analysis (DA) project was conducted to evaluate sea lamprey control policies for the SMR; this project suggested that an integrated policy of trapping, sterile male releases, and Bayluscide treatment was the most cost-effective policy. Further, it concluded that formal assessment of larval sea lamprey abundance and distribution in the SMR would be valuable for future evaluation of control strategies. We updated this earlier analysis, adding information from annual larval assessments conducted since 1999 and evaluating additional control policies. Bayluscide treatments continued to be critical for sea lamprey control, but high recruitment compensation minimized the effectiveness of trapping and sterile male release under current feasible ranges. Because Bayluscide control is costly, development of strategies to enhance trapping success remains a priority. This study illustrates benefits of an adaptive management cycle, wherein models inform decisions, are updated based on learning achieved from those decisions, and ultimately inform future decisions.

Upstream migration of Pacific lampreys in the John Day River, Oregon: Behavior, timing, and habitat use

Science.gov (United States)

Robinson, T. Craig; Bayer, J.M.

2005-01-01

Adult Pacific lamprey migration and habitat preferences for over-winter holding and spawning, and larval rearing in tributaries to the Columbia River are not well understood. The John Day River is one such tributary where larval and adult stages of this species have been documented, and its free-flowing character provided the opportunity to study migration of Pacific lampreys unimpeded by passage constraints. Forty-two adult Pacific lampreys were captured in the John Day River near its mouth during their upstream migration. Pacific lampreys were surgically implanted with radio transmitters and released onsite, and tracked by fixed-site, aerial, and terrestrial telemetry methods for nearly one year. Adults moved upstream exclusively at night, with a mean rate of 11.1 ?? 6.3 km/day. They halted upstream migration by September, and held a single position for approximately six months in the lateral margins of riffles and glides, using boulders for cover. More than half of Pacific lampreys resumed migration in March before ending movement in early May. Pacific lampreys that resumed migration in spring completed a median of 87% of their upstream migration before over-winter holding. Upon completing migration. Pacific lampreys briefly held position before beginning downstream movement at the end of May. Though not directly observed, halting migration and movement downstream were likely the result of spawning and death. Gains in adult Pacific lamprey passage through the Columbia River hydrosystem and tributaries may be made by improvements that would expedite migration during spring and summer and increase the quantity and variety of cover and refuge opportunities. ?? 2005 by the Northwest Scientific Association. All rights reserved.

Chemical Characterization of Lipophilic Constituents in the Skin of Migratory Adult Sea Lamprey from the Great Lakes Region.

Directory of Open Access Journals (Sweden)

Amila A Dissanayake

Full Text Available The sea lamprey (Petromzons marinus is an invasive ectoparasite of large-bodied fishes that adversely affects the fishing industry and ecology of the Laurentian Great Lakes. Lipid content in the whole sea lamprey and muscles, liver and kidney of metamorphosing larval stages has been reported. Similarly, the fatty acid profile of the rope tissues of sexually-mature male sea lampreys has also been reported. The average body weight of a sub-adult migratory sea lamprey is 250 g, which includes 14.4% skin (36 g. Our preliminary extraction data of an adult sea lamprey skin revealed that it contained approximately 8.5% of lipophilic compounds. Lamprey skin is home to a naturally aversive compound (an alarm cue that is being developed into a repellent for use in pest management. As part of an ongoing investigation to identify the chemical structure of the sea lamprey alarm cue, we extracted the skin with water and methanol, respectively. The methanolic extract (1.55% contained exclusively lipophilic compounds and did not include the alarm cue. We chemically characterized all compounds present in the methanolic extract as cholesterol esters (CE, tri- and di-glycerides (TG and DG, cholesterol, free fatty acids (FFA and minor amounts of plasticizers. The free fatty acids fraction was composed of saturated (41.8%, monounsaturated (40.7% and polyunsaturated (17.4% fatty acids, respectively. The plasticizers characterized were phthalate and benzoate and found to be 0.95 mg and 2.54 mg, respectively, per adult sea lamprey skin. This is the first report of the chemical characterization of all the lipophilic constituents in the skin of sub-adult migratory sea lamprey. The CEs isolated and characterized from sea lamprey skin are also for the first time.

Evidence that lake trout served as a buffer against sea lamprey predation on burbot in Lake Erie

Science.gov (United States)

Stapanian, M.A.; Madenjian, C.P.

2007-01-01

The population of burbot Lota lota in Lake Erie recovered during 1986–2003, mainly because of the control of sea lamprey Petromyzon marinus, which began in 1986. Burbot populations continued to grow during 1996–1998, when sea lamprey control was substantially reduced. We calculated mortality parameters for burbot in Lake Erie by estimating age at capture for 2,793 burbot caught in annual gill-net surveys of eastern Lake Erie from 1994 to 2003. Based on catch-curve analysis, annual mortality in Lake Erie during 1994–2003 was estimated as 33%. Annual mortality of the 1992 year-class of burbot was estimated as 30%. The mortality of burbot during the years of reduced sea lamprey control was not different from that during the 3 years preceding reduced control and was significantly lower than that during the entire portion of the time series in which full sea lamprey control was conducted. These results suggest that the reduction in sea lamprey control did not lead to increased burbot mortality. The catch per gill-net lift of large burbot (total length > 600 mm), the size preferred by sea lampreys, was lower than that of adult lake trout Salvelinus namaycush (age 5 and older; total length > 700 mm) before lampricide application was reduced. Although adult lake trout populations declined, the abundance of large burbot did not change during the period of reduced lampricide application. These results support a hypothesis that a healthy population of adult lake trout can serve as a buffer species, acting to reduce predation of burbot by sea lampreys when sea lamprey populations increase. Burbot attained sexual maturity at a relatively early age (3 or 4 years) and a total length (approximately 500 mm) that was smaller than the preferred prey size for sea lampreys. These characteristics and the buffering effect of the lake trout population enabled growth of the burbot population during the brief period when lamprey control was reduced.

Evaluate Status of Pacific Lamprey in the Clearwater River Drainage, Idaho, Annual Report 2002.

Energy Technology Data Exchange (ETDEWEB)

Cochnauer, Tim; Claire, Christopher

2003-10-01

In 2002 Idaho Department of Fish and Game continued investigation into the status of Pacific lamprey populations in Idaho's Clearwater River drainage. Trapping, electrofishing, and spawning ground redd surveys were used to determine Pacific lamprey distribution, life history strategies, and habitat requirements in the South Fork Clearwater River, Lochsa River, Selway River, and Middle Fork Clearwater River subbasins. Five-hundred forty-one ammocoetes were captured electroshocking 70 sites in the South Fork Clearwater River, Lochsa River, Selway River, Middle Fork Clearwater River, Clearwater River, and their tributaries in 2002. Habitat utilization surveys in Red River support previous work indicating Pacific lamprey ammocoete densities are greater in lateral scour pool habitats compared to riffles and rapids. Presence-absence survey findings in 2002 augmented 2000 and 2001 indicating Pacific lamprey macrothalmia and ammocoetes are not numerous or widely distributed. Pacific lamprey distribution was confined to the lower reaches of Red River below rkm 8.0, the South Fork Clearwater River, Lochsa River (Ginger Creek to mouth), Selway River (Race Creek to mouth), Middle Fork Clearwater River, and the Clearwater River (downstream to Potlatch River).

Lampreys, the jawless vertebrates, contain only two ParaHox gene clusters.

Science.gov (United States)

Zhang, Huixian; Ravi, Vydianathan; Tay, Boon-Hui; Tohari, Sumanty; Pillai, Nisha E; Prasad, Aravind; Lin, Qiang; Brenner, Sydney; Venkatesh, Byrappa

2017-08-22

ParaHox genes ( Gsx , Pdx , and Cdx ) are an ancient family of developmental genes closely related to the Hox genes. They play critical roles in the patterning of brain and gut. The basal chordate, amphioxus, contains a single ParaHox cluster comprising one member of each family, whereas nonteleost jawed vertebrates contain four ParaHox genomic loci with six or seven ParaHox genes. Teleosts, which have experienced an additional whole-genome duplication, contain six ParaHox genomic loci with six ParaHox genes. Jawless vertebrates, represented by lampreys and hagfish, are the most ancient group of vertebrates and are crucial for understanding the origin and evolution of vertebrate gene families. We have previously shown that lampreys contain six Hox gene loci. Here we report that lampreys contain only two ParaHox gene clusters (designated as α- and β-clusters) bearing five ParaHox genes ( Gsxα , Pdxα , Cdxα , Gsxβ , and Cdxβ ). The order and orientation of the three genes in the α-cluster are identical to that of the single cluster in amphioxus. However, the orientation of Gsxβ in the β-cluster is inverted. Interestingly, Gsxβ is expressed in the eye, unlike its homologs in jawed vertebrates, which are expressed mainly in the brain. The lamprey Pdxα is expressed in the pancreas similar to jawed vertebrate Pdx genes, indicating that the pancreatic expression of Pdx was acquired before the divergence of jawless and jawed vertebrate lineages. It is likely that the lamprey Pdxα plays a crucial role in pancreas specification and insulin production similar to the Pdx of jawed vertebrates.

Factors influencing capture of invasive sea lamprey in traps baited with a synthesized sex pheromone component

Science.gov (United States)

Johnson, Nicholas; Siefkes, Michael J.; Wagner, C. Michael; Bravener, Gale; Steeves, Todd; Twohey, Michael; Li, Weiming

2015-01-01

The sea lamprey, Petromyzon marinus, is emerging as a model organism for understanding how pheromones can be used for manipulating vertebrate behavior in an integrated pest management program. In a previous study, a synthetic sex pheromone component 7α,12α, 24-trihydroxy-5α-cholan-3-one 24-sulfate (3kPZS) was applied to sea lamprey traps in eight streams at a final in-stream concentration of 10−12 M. Application of 3kPZS increased sea lamprey catch, but where and when 3kPZS had the greatest impact was not determined. Here, by applying 3kPZS to additional streams, we determined that overall increases in yearly exploitation rate (proportion of sea lampreys that were marked, released, and subsequently recaptured) were highest (20–40 %) in wide streams (~40 m) with low adult sea lamprey abundance (pheromone), sea lamprey may have been more responsive to a partial sex pheromone blend emitted from traps. Furthermore, we found that the largest and most consistent responses to 3kPZS were during nights early in the trapping season, when water temperatures were increasing. This may have occurred because, during periods of increasing water temperatures, sea lamprey become more active and males at large may not have begun to release sex pheromone. In general, our results are consistent with those for pheromones of invertebrates, which are most effective when pest density is low and when pheromone competition is low.

Status report of the Pacific lamprey (Lampetra tridentata) in the Columbia River Basin

International Nuclear Information System (INIS)

Close, D.A.; Fitzpatrick, M.; Li, H.; James, G.

1995-07-01

The widespread decline of Pacific lamprey (Lampetra tridentata) in the Pacific Northwest, especially in the Columbia River system has led to concerns and questions from a number of regional agencies, Native American tribes, and the public. To address these concerns, new research efforts must focus on specific problems associated with this understudied species. The preservation and restoration of this species is critical for a number of reasons, including its importance to the tribes and its importance as an indicator of ecosystem health. Historically lamprey have been labeled a pest species due to the problems associated with the exotic sea lamprey, (Petromyzon marinus), invading the Great Lakes

Status Report of the Pacific Lamprey (Lampetra Trzdentata) in the Columbia River Basin.

Energy Technology Data Exchange (ETDEWEB)

Close, David A.; Parker, Blaine; James, gary

1995-07-01

The widespread decline of Pacific lamprey (Lampetra tridentata) in the Pacific Northwest, especially in the Columbia River system has led to concerns and questions from a number of regional agencies, Native American tribes, and the public. To address these concerns, new research efforts must focus on specific problems associated with this understudied species. The preservation and restoration of this species is critical for a number of reasons, including its importance to the tribes and its importance as an indicator of ecosystem health. Historically lamprey have been labeled a pest species due to the problems associated with the exotic sea lamprey, (Petromyzon marinus), invading the Great Lakes.

Large wood and in-stream habitat for juvenile coho salmon and larval lampreys in a Pacific Northwest stream

Science.gov (United States)

Gonzalez, Rosalinda; Dunham, Jason B.; Lightcap, Scott W.; McEnroe, Jeffery R.

2017-01-01

The influences of large wood on Pacific salmon are well-studied, but studies of nonsalmonid species such as lampreys are uncommon. To address this need, we evaluated the potential effects of large wood on larval lampreys (Pacific Lamprey, Entosphenus tridentatus; and potentially Western Brook Lamprey Lampetra richardsoni), as well as juvenile Coho Salmon Oncorhynchus kisutch, in a small coastal Oregon stream. Our objectives were to 1) identify in-stream habitat characteristics associated with the presence of larval lampreys and abundance of juvenile Coho Salmon; and 2) evaluate how these characteristics were associated with in-stream wood. To address habitat use, we quantified presence of larval lampreys in 92 pools and abundance of juvenile Coho Salmon in 44 pools during summer low flows. We focused on a study reach where large wood was introduced into the stream between 2008 and 2009. Results indicated that presence of larval lampreys was significantly associated with availability of fine sediment and deeper substrate. The abundance of juvenile Coho Salmon (fish/pool) was strongly associated with pool surface area and to a weaker extent with the proportion of cobble and boulder substrates in pools. Pools with wood, regardless of whether they were formed by wood, had significantly greater coverage of fine sediment, deeper substrate, and greater pool surface area. Taken together, these results suggest that in-stream wood can provide habitat associated with presence of larval lampreys and greater abundance of juvenile Coho Salmon.

Tradeoff between assessment and control of aquatic invasive species: A case study of sea lamprey management in the St. Marys River

Science.gov (United States)

Robinson, Jason M.; Wilberg, Michael J.; Adams, Jean V.; Jones, Michael L.

2016-01-01

Allocating resources between the gathering of information to guide management actions and implementing those actions presents an inherent tradeoff. This tradeoff is evident for control of the Sea Lamprey Petromyzon marinus in the St. Marys River, connecting Lakes Huron and Superior and a major source of parasitic Sea Lampreys to Lake Huron and northern Lake Michigan. Larval Sea Lampreys in the St. Marys River are controlled through the application of Bayluscide, which is applied to areas of high larval density. Bayluscide applications are guided with an annual deepwater electrofishing survey to estimate larval Sea Lamprey density at relatively fine spatial scales. We took a resampling approach to describe the effect of sampling intensity on the success of the larval Sea Lamprey management program and explicitly incorporated the economic tradeoff between assessment and control efforts to maximize numbers of larvae killed in the St. Marys River. When no tradeoff between assessment and control was incorporated, increasing assessment always led to more larvae killed for the same treatment budget. When the tradeoff was incorporated, the sampling intensity that maximized the number of larvae killed depended on the overall budget available. Increased sampling intensities maximized effectiveness under medium to large budgets (US \\$0.4 to \\$2.0 million), and intermediate sampling intensities maximized effectiveness under low budgets. Sea Lamprey control actions based on assessment information outperformed those that were implemented with no assessment under all budget scenarios.

Thyroid hormone and retinoid X receptor function and expression during sea lamprey (Petromyzon marinus) metamorphosis.

Science.gov (United States)

Manzon, Lori A; Youson, John H; Holzer, Guillaume; Staiano, Leopoldo; Laudet, Vincent; Manzon, Richard G

2014-08-01

expression data suggests that THs, either directly or via a metabolite, may function to positively modulate changes at the tissue or organ levels during lamprey metamorphosis. Collectively the results presented herein support the hypothesis that THs have a dual functional role in the lamprey life cycle whereby high levels promote larval feeding, growth and lipogenesis and low levels promote metamorphosis. Copyright © 2014 Elsevier Inc. All rights reserved.

The effect of rapid and sustained decompression on barotrauma in juvenile brook lamprey and Pacific lamprey: implications for passage at hydroelectric facilities

Energy Technology Data Exchange (ETDEWEB)

Colotelo, Alison HA; Pflugrath, Brett D.; Brown, Richard S.; Brauner, Colin J.; Mueller, Robert P.; Carlson, Thomas J.; Deng, Zhiqun; Ahmann, Martin L.; Trumbo, Bradly A.

2012-10-01

Fish passing downstream through hydroelectric facilities may pass through hydroturbines where they experience a rapid decrease in barometric pressure as they pass by turbine blades, which can lead to barotraumas including swim bladder rupture, exopthalmia, emboli, and hemorrhaging. In juvenile Chinook salmon, the main mechanism for injury is thought to be expansion of existing gases (particularly those present in the swim bladder) and the rupture of the swim bladder ultimately leading to exopthalmia, emboli and hemorrhaging. In fish that lack a swim bladder, such as lamprey, the rate and severity of barotraumas due to rapid decompression may be reduced however; this has yet to be extensively studied. Another mechanism for barotrauma can be gases coming out of solution and the rate of this occurrence may vary among species. In this study, juvenile brook and Pacific lamprey acclimated to 146.2 kPa (equivalent to a depth of 4.6 m) were subjected to rapid (<1 sec; brook lamprey only) or sustained decompression (17 minutes) to a very low pressure (13.8 kPa) using a protocol previously applied to juvenile Chinook salmon. No mortality or evidence of barotraumas, as indicated by the presence of hemorrhages, emboli or exopthalmia, were observed during rapid or sustained decompression, nor following recovery for up to 120 h following sustained decompression. In contrast, mortality or injury would be expected for 97.5% of juvenile Chinook salmon exposed to a similar rapid decompression to these very low pressures. Additionally, juvenile Chinook salmon experiencing sustained decompression died within 7 minutes, accompanied by emboli in the fins and gills and hemorrhaging in the tissues. Thus, juvenile lamprey may not be susceptible to barotraumas associated with hydroturbine passage to the same degree as juvenile salmonids, and management of these species should be tailored to their specific morphological and physiological characteristics.

Anesthesia of juvenile Pacific Lampreys with MS-222, BENZOAK, AQUI-S 20E, and Aquacalm

Science.gov (United States)

Christiansen, Helena E.; Gee, Lisa P.; Mesa, Matthew G.

2013-01-01

Effective anesthetics are a critical component of safe and humane fish handling procedures. We tested three concentrations each of four anesthetics—Finquel (tricaine methanesulfonate, herein referred to as MS-222), BENZOAK (20% benzocaine), AQUI-S 20E (10% eugenol), and Aquacalm (metomidate hydrochloride)—for efficacy and safety in metamorphosed, outmigrating juvenile Pacific Lampreys Entosphenus tridentatus. The anesthetics MS-222 (100 mg/L) and BENZOAK (60 mg/L) were the most effective for anesthetizing juvenile Pacific Lampreys to a handleable state with minimal irritation to the fish. Fish anesthetized with BENZOAK also had lower rates of fungal infection than those exposed to MS-222, AQUI-S 20E, or no anesthetic. Exposure to AQUI-S 20E irritated juvenile Pacific Lampreys, causing them to leap or climb out of the anesthetic solution, and Aquacalm anesthetized fish to a handleable state too slowly and incompletely for effective use with routine handling procedures. Our results indicate that MS-222 and BENZOAK are effective anesthetics for juvenile Pacific Lampreys, but field studies are needed to determine whether exposure to MS-222 increases risk of fungal infection in juvenile Pacific Lampreys released to the wild.

Genetic engineering of chimeric antigen receptors using lamprey derived variable lymphocyte receptors

Directory of Open Access Journals (Sweden)

Robert Moot

2016-01-01

Full Text Available Chimeric antigen receptors (CARs are used to redirect effector cell specificity to selected cell surface antigens. Using CARs, antitumor activity can be initiated in patients with no prior tumor specific immunity. Although CARs have shown promising clinical results, the technology remains limited by the availability of specific cognate cell target antigens. To increase the repertoire of targetable tumor cell antigens we utilized the immune system of the sea lamprey to generate directed variable lymphocyte receptors (VLRs. VLRs serve as membrane bound and soluble immune effectors analogous but not homologous to immunoglobulins. They have a fundamentally different structure than immunoglobulin (Ig-based antibodies while still demonstrating high degrees of specificity and affinity. To test the functionality of VLRs as the antigen recognition domain of CARs, two VLR-CARs were created. One contained a VLR specific for a murine B cell leukemia and the other contained a VLR specific for the human T cell surface antigen, CD5. The CAR design consisted of the VLR sequence, myc-epitope tag, CD28 transmembrane domain, and intracellular CD3ζ signaling domain. We demonstrate proof of concept, including gene transfer, biosynthesis, cell surface localization, and effector cell activation for multiple VLR-CAR designs. Therefore, VLRs provide an alternative means of CAR-based cancer recognition.

Variations in the presence of chloride cells in the gills of lampreys (Petromyzontiformes) and their evolutionary implications.

Science.gov (United States)

Bartels, H; Docker, M F; Krappe, M; White, M M; Wrede, C; Potter, I C

2015-04-01

Although confined to fresh water, non-parasitic species of lampreys and the landlocked parasitic sea lamprey, all of which were derived relatively recently from an adromous ancestors, still develop chloride cells, whose function in their ancestors was for osmoregulation in marine waters during the adult parasitic phase. In contrast, such cells are not developed by the non-parasitic least brook lamprey Lampetra aepyptera, which has been separated from its ancestor for >2 million years, nor by the freshwater parasitic species of the genus Ichthyomyzon. The length of time that a non-parasitic species or landlocked parasitic form or species has spent in fresh water is thus considered the overriding factor determining whether chloride cells are developed by those lampreys.

On the Mechanism(s of Membrane Permeability Transition in Liver Mitochondria of Lamprey, Lampetra fluviatilis L.: Insights from Cadmium

Directory of Open Access Journals (Sweden)

Elena A. Belyaeva

2014-01-01

Full Text Available Previously we have shown that opening of the mitochondrial permeability transition pore in its low conductance state is the case in hepatocytes of the Baltic lamprey (Lampetra fluviatilis L. during reversible metabolic depression taking place in the period of its prespawning migration when the exogenous feeding is switched off. The depression is observed in the last year of the lamprey life cycle and is conditioned by reversible mitochondrial dysfunction (mitochondrial uncoupling in winter and coupling in spring. To further elucidate the mechanism(s of induction of the mitochondrial permeability transition pore in the lamprey liver, we used Cd2+ and Ca2+ plus Pi as the pore inducers. We found that Ca2+ plus Pi induced the high-amplitude swelling of the isolated “winter” mitochondria both in isotonic sucrose and ammonium nitrate medium while both low and high Cd2+ did not produce the mitochondrial swelling in these media. Low Cd2+ enhanced the inhibition of basal respiration rate of the “winter” mitochondria energized by NAD-dependent substrates whereas the same concentrations of the heavy metal evoked its partial stimulation on FAD-dependent substrates. The above changes produced by Cd2+ or Ca2+ plus Pi in the “winter” mitochondria were only weakly (if so sensitive to cyclosporine A (a potent pharmacological desensitizer of the nonselective pore added alone and they were not sensitive to dithiothreitol (a dithiol reducing agent. Under monitoring of the transmembrane potential of the “spring” lamprey liver mitochondria, we revealed that Cd2+ produced its decrease on both types of the respiratory substrates used that was strongly hampered by cyclosporine A, and the membrane potential was partially restored by dithiothreitol. The effects of different membrane permeability modulators on the lamprey liver mitochondria function and the seasonal changes in their action are discussed.

Origins of gonadotropin-releasing hormone (GnRH) in vertebrates: identification of a novel GnRH in a basal vertebrate, the sea lamprey.

Science.gov (United States)

Kavanaugh, Scott I; Nozaki, Masumi; Sower, Stacia A

2008-08-01

We cloned a cDNA encoding a novel (GnRH), named lamprey GnRH-II, from the sea lamprey, a basal vertebrate. The deduced amino acid sequence of the newly identified lamprey GnRH-II is QHWSHGWFPG. The architecture of the precursor is similar to that reported for other GnRH precursors consisting of a signal peptide, decapeptide, a downstream processing site, and a GnRH-associated peptide; however, the gene for lamprey GnRH-II does not have introns in comparison with the gene organization for all other vertebrate GnRHs. Lamprey GnRH-II precursor transcript was widely expressed in a variety of tissues. In situ hybridization of the brain showed expression and localization of the transcript in the hypothalamus, medulla, and olfactory regions, whereas immunohistochemistry using a specific antiserum showed only GnRH-II cell bodies and processes in the preoptic nucleus/hypothalamus areas. Lamprey GnRH-II was shown to stimulate the hypothalamic-pituitary axis using in vivo and in vitro studies. Lamprey GnRH-II was also shown to activate the inositol phosphate signaling system in COS-7 cells transiently transfected with the lamprey GnRH receptor. These studies provide evidence for a novel lamprey GnRH that has a role as a third hypothalamic GnRH. In summary, the newly discovered lamprey GnRH-II offers a new paradigm of the origin of the vertebrate GnRH family. We hypothesize that due to a genome/gene duplication event, an ancestral gene gave rise to two lineages of GnRHs: the gnathostome GnRH and lamprey GnRH-II.

Temporal genetic population structure and interannual variation in migration behavior of Pacific Lamprey Entosphenus tridentatus

Science.gov (United States)

Clemens, Benjamin J.; Wyss, Lance A.; McCoun, Rebecca; Courter, Ian; Schwabe, Lawrence; Peery, Christopher; Schreck, Carl B.; Spice, Erin K.; Docker, Margaret F.

2017-01-01

Studies using neutral loci suggest that Pacific lamprey, Entosphenus tridentatus, lack strong spatial genetic population structure. However, it is unknown whether temporal genetic population structure exists. We tested whether adult Pacific lamprey: (1) show temporal genetic population structure; and (2) migrate different distances between years. We non-lethally sampled lamprey for DNA in 2009 and 2010 and used eight microsatellite loci to test for genetic population structure. We used telemetry to record the migration behaviors of these fish. Lamprey were assignable to three moderately differentiated genetic clusters (FST = 0.16–0.24 for all pairwise comparisons): one cluster was composed of individuals from 2009, and the other two contained individuals from 2010. The FST value between years was 0.13 and between genetic clusters within 2010 was 0.20. A total of 372 (72.5%) fish were detected multiple times during their migrations. Most fish (69.9%) remained in the mainstem Willamette River; the remaining 30.1% migrated into tributaries. Eighty-two lamprey exhibited multiple back-and-forth movements among tributaries and the mainstem, which may indicate searching behaviors. All migration distances were significantly greater in 2010, when the amplitude of river discharge was greater. Our data suggest genetic structuring between and within years that may reflect different cohorts.

Rapid evolution meets invasive species control: The potential for pesticide resistance in sea lamprey

Science.gov (United States)

Dunlop, Erin S.; McLaughlin, Robert L.; Adams, Jean V.; Jones, Michael L.; Birceanu, Oana; Christie, Mark R.; Criger, Lori A.; Hinderer, Julia L.M.; Hollingworth, Robert M.; Johnson, Nicholas; Lantz, Stephen R.; Li, Weiming; Miller, James R.; Morrison, Bruce J.; Mota-Sanchez, David; Muir, Andrew M.; Sepulveda, Maria S.; Steeves, Todd B.; Walter, Lisa; Westman, Erin; Wirgin, Isaac; Wilkie, Michael P.

2018-01-01

Rapid evolution of pest, pathogen and wildlife populations can have undesirable effects; for example, when insects evolve resistance to pesticides or fishes evolve smaller body size in response to harvest. A destructive invasive species in the Laurentian Great Lakes, the sea lamprey (Petromyzon marinus) has been controlled with the pesticide 3-trifluoromethyl-4-nitrophenol (TFM) since the 1950s. We evaluated the likelihood of sea lamprey evolving resistance to TFM by (1) reviewing sea lamprey life history and control; (2) identifying physiological and behavioural resistance strategies; (3) estimating the strength of selection from TFM; (4) assessing the timeline for evolution; and (5) analyzing historical toxicity data for evidence of resistance. The number of sea lamprey generations exposed to TFM was within the range observed for fish populations where rapid evolution has occurred. Mortality from TFM was estimated as 82-90%, suggesting significant selective pressure. However, 57 years of toxicity data revealed no increase in lethal concentrations of TFM. Vigilance and the development of alternative controls are required to prevent this aquatic invasive species from evolving strategies to evade control.

Management strategy evaluation of pheromone-baited trapping techniques to improve management of invasive sea lamprey

Science.gov (United States)

Dawson, Heather; Jones, Michael L.; Irwin, Brian J.; Johnson, Nicholas; Wagner, Michael C.; Szymanski, Melissa

2016-01-01

We applied a management strategy evaluation (MSE) model to examine the potential cost-effectiveness of using pheromone-baited trapping along with conventional lampricide treatment to manage invasive sea lamprey. Four pheromone-baited trapping strategies were modeled: (1) stream activation wherein pheromone was applied to existing traps to achieve 10−12 mol/L in-stream concentration, (2) stream activation plus two additional traps downstream with pheromone applied at 2.5 mg/hr (reverse-intercept approach), (3) trap activation wherein pheromone was applied at 10 mg/hr to existing traps, and (4) trap activation and reverse-intercept approach. Each new strategy was applied, with remaining funds applied to conventional lampricide control. Simulating deployment of these hybrid strategies on fourteen Lake Michigan streams resulted in increases of 17 and 11% (strategies 1 and 2) and decreases of 4 and 7% (strategies 3 and 4) of the lakewide mean abundance of adult sea lamprey relative to status quo. MSE revealed performance targets for trap efficacy to guide additional research because results indicate that combining lampricides and high efficacy trapping technologies can reduce sea lamprey abundance on average without increasing control costs.

A new clarification method to visualize biliary degeneration during liver metamorphosis in sea lamprey (Petromyzon marinus)

Science.gov (United States)

Chung-Davidson, Yu-Wen; Davidson, Peter J.; Scott, Anne M.; Walaszczyk, Erin J.; Brant, Cory O.; Buchinger, Tyler; Johnson, Nicholas S.; Li, Weiming

2014-01-01

Biliary atresia is a rare disease of infancy, with an estimated 1 in 15,000 frequency in the southeast United States, but more common in East Asian countries, with a reported frequency of 1 in 5,000 in Taiwan. Although much is known about the management of biliary atresia, its pathogenesis is still elusive. The sea lamprey (Petromyzon marinus) provides a unique opportunity to examine the mechanism and progression of biliary degeneration. Sea lamprey develop through three distinct life stages: larval, parasitic, and adult. During the transition from larvae to parasitic juvenile, sea lamprey undergo metamorphosis with dramatic reorganization and remodeling in external morphology and internal organs. In the liver, the entire biliary system is lost, including the gall bladder and the biliary tree. A newly-developed method called “CLARITY” was modified to clarify the entire liver and the junction with the intestine in metamorphic sea lamprey. The process of biliary degeneration was visualized and discerned during sea lamprey metamorphosis by using laser scanning confocal microscopy. This method provides a powerful tool to study biliary atresia in a unique animal model.

An annotated bibliography for lamprey habitat in the White Salmon River, Washington

Science.gov (United States)

Allen, M. Brady

2012-01-01

The October 2011 decommissioning of Condit Dam on the White Salmon River at river kilometer (rkm) 5.3 removed a significant fish passage barrier from the White Salmon River basin for the first time in nearly a century. This affords an opportunity to regain a potentially important drainage basin for Pacific lamprey (Entosphenus tridentatus) production. In anticipation of Pacific lamprey recolonization or reintroduction, aquatic resource managers, such as the Yakama Nation (YN), are planning to perform surveys in the White Salmon River and its tributaries. The likely survey objectives will be to investigate the presence of lamprey, habitat conditions, and habitat availability. In preparation for this work, a compilation and review of the relevant aquatic habitat and biological information on the White Salmon River was conducted. References specific to the White Salmon River were collected and an annotated bibliography was produced including reports containing:

Anadromous sea lampreys recolonize a Maine coastal river tributary after dam removal

Science.gov (United States)

Hogg, Robert; Coghlan, Stephen M.; Zydlewski, Joseph D.

2013-01-01

Sedgeunkedunk Stream, a third-order tributary to the Penobscot River, Maine, historically supported several anadromous fishes, including the Atlantic Salmon Salmo salar, AlewifeAlosa pseudoharengus, and Sea Lamprey Petromyzon marinus. However, two small dams constructed in the 1800s reduced or eliminated spawning runs entirely. In 2009, efforts to restore marine–freshwater connectivity in the system culminated with removal of the lowermost dam, thus providing access to an additional 4.6 km of lotic habitat. Because Sea Lampreys utilized accessible habitat prior to dam removal, they were chosen as a focal species with which to quantify recolonization. During spawning runs of 2008–2011 (before and after dam removal), individuals were marked with PIT tags and their activity was tracked with daily recapture surveys. Open-population mark–recapture models indicated a fourfold increase in the annual abundance of spawning-phase Sea Lampreys, with estimates rising from 59±4 () before dam removal (2008) to 223±18 and 242±16 after dam removal (2010 and 2011, respectively). Accompanying the marked increase in annual abundance was a greater than fourfold increase in nesting sites: the number of nests increased from 31 in 2008 to 128 and 131 in 2010 and 2011, respectively. During the initial recolonization event (i.e., in 2010), Sea Lampreys took 6 d to move past the former dam site and 9 d to expand into the furthest upstream reaches. Conversely, during the 2011 spawning run, Sea Lampreys took only 3 d to penetrate into the upstream reaches, thus suggesting a potential positive feedback in which larval recruitment into the system may have attracted adult spawners via conspecific pheromone cues. Although more research is needed to verify the migratory pheromone hypothesis, our study clearly demonstrates that small-stream dam removal in coastal river systems has the potential to enhance recovery of declining anadromous fish populations.

Anatomical recovery of the GABAergic system after a complete spinal cord injury in lampreys.

Science.gov (United States)

Romaus-Sanjurjo, D; Valle-Maroto, S M; Barreiro-Iglesias, A; Fernández-López, B; Rodicio, M C

2018-03-15

Lampreys recover locomotion spontaneously several weeks after a complete spinal cord injury. Dysfunction of the GABAergic system following SCI has been reported in mammalian models. So, it is of great interest to understand how the GABAergic system of lampreys adapts to the post-injury situation and how this relates to spontaneous recovery. The spinal cord of lampreys contains 3 populations of GABAergic neurons and most of the GABAergic innervation of the spinal cord comes from these local cells. GABAB receptors are expressed in the spinal cord of lampreys and they play important roles in the control of locomotion. The aims of the present study were to quantify: 1) the changes in the number of GABAergic neurons and innervation of the spinal cord and 2) the changes in the expression of the gabab receptor subunits b1 and b2 in the spinal cord of the sea lamprey after SCI. We performed complete spinal cord transections at the level of the fifth gill of mature larval lampreys and GABA immunohistochemistry or gabab in situ hybridization experiments. Animals were analysed up to 10 weeks post-lesion (wpl), when behavioural analyses showed that they recovered normal appearing locomotion (stage 6 in the Ayer's scale of locomotor recovery). We observed a significant decrease in the number of GABA-ir cells and fibres 1 h after lesion both rostral and caudal to the lesion site. GABA-ir cell numbers and innervation were recovered to control levels 1 to 2 wpl. At 1, 4 and 10 wpl the expression of gabab1 and gabab2 transcripts was significantly decreased in the spinal cord compared to control un-lesioned animals. This is the first study reporting the quantitative long-term changes in the number of GABAergic cells and fibres and in the expression of gabab receptors in the spinal cord of any vertebrate following a traumatic SCI. Our results show that in lampreys there is a full recovery of the GABAergic neurons and a decrease in the expression of gabab receptors when functional

Anatomy of the lamprey ear: morphological evidence for occurrence of horizontal semicircular ducts in the labyrinth of Petromyzon marinus

Science.gov (United States)

Maklad, Adel; Reed, Caitlyn; Johnson, Nicholas S.; Fritzsch, Bernd

2014-01-01

In jawed (gnathostome) vertebrates, the inner ears have three semicircular canals arranged orthogonally in the three Cartesian planes: one horizontal (lateral) and two vertical canals. They function as detectors for angular acceleration in their respective planes. Living jawless craniates, cyclostomes (hagfish and lamprey) and their fossil records seemingly lack a lateral horizontal canal. The jawless vertebrate hagfish inner ear is described as a torus or doughnut, having one vertical canal, and the jawless vertebrate lamprey having two. These observations on the anatomy of the cyclostome (jawless vertebrate) inner ear have been unchallenged for over a century, and the question of how these jawless vertebrates perceive angular acceleration in the yaw (horizontal) planes has remained open. To provide an answer to this open question we reevaluated the anatomy of the inner ear in the lamprey, using stereoscopic dissection and scanning electron microscopy. The present study reveals a novel observation: the lamprey has two horizontal semicircular ducts in each labyrinth. Furthermore, the horizontal ducts in the lamprey, in contrast to those of jawed vertebrates, are located on the medial surface in the labyrinth rather than on the lateral surface. Our data on the lamprey horizontal duct suggest that the appearance of the horizontal canal characteristic of gnathostomes (lateral) and lampreys (medial) are mutually exclusive and indicate a parallel evolution of both systems, one in cyclostomes and one in gnathostome ancestors.

Population ecology of the sea lamprey (Petromyzon marinus) as an invasive species in the Laurentian Great Lakes and an imperiled species in Europe

Science.gov (United States)

Hansen, Michael J.; Madenjian, Charles P.; Slade, Jeffrey W.; Steeves, Todd B.; Almeida, Pedro R.; Quintella, Bernardo R.

2016-01-01

The sea lamprey Petromyzon marinus (Linnaeus) is both an invasive non-native species in the Laurentian Great Lakes of North America and an imperiled species in much of its native range in North America and Europe. To compare and contrast how understanding of population ecology is useful for control programs in the Great Lakes and restoration programs in Europe, we review current understanding of the population ecology of the sea lamprey in its native and introduced range. Some attributes of sea lamprey population ecology are particularly useful for both control programs in the Great Lakes and restoration programs in the native range. First, traps within fish ladders are beneficial for removing sea lampreys in Great Lakes streams and passing sea lampreys in the native range. Second, attractants and repellants are suitable for luring sea lampreys into traps for control in the Great Lakes and guiding sea lamprey passage for conservation in the native range. Third, assessment methods used for targeting sea lamprey control in the Great Lakes are useful for targeting habitat protection in the native range. Last, assessment methods used to quantify numbers of all life stages of sea lampreys would be appropriate for measuring success of control in the Great Lakes and success of conservation in the native range.

Cloning, phylogeny, and regional expression of a Y5 receptor mRNA in the brain of the sea lamprey (Petromyzon marinus).

Science.gov (United States)

Pérez-Fernández, Juan; Megías, Manuel; Pombal, Manuel A

2014-04-01

The NPY receptors known as Y receptors are classified into three subfamilies, Y1, Y2, and Y5, and are involved in different physiological functions. The Y5 receptor is the only member of the Y5 subfamily, and it is present in all vertebrate groups, except for teleosts. Both molecular and pharmacological studies show that Y5 receptor is highly conserved during vertebrate evolution. Furthermore, this receptor is widely expressed in the mammalian brain, including the hypothalamus, where it is thought to take part in feeding and homeostasis regulation. Lampreys belong to the agnathan lineage, and they are thought to have branched out between the two whole-genome duplications that occurred in vertebrates. Therefore, they are in a key position for studies on the evolution of gene families in vertebrates. Here we report the cloning, phylogeny, and brain expression pattern of the sea lamprey Y5 receptor. In phylogenetic studies, the lamprey Y5 receptor clusters in a basal position, together with Y5 receptors of other vertebrates. The mRNA of this receptor is broadly expressed in the lamprey brain, being especially abundant in hypothalamic areas. Its expression pattern is roughly similar to that reported for other vertebrates and parallels the expression pattern of the Y1 receptor subtype previously described by our group, as it occurs in mammals. Altogether, these results confirm that a Y5 receptor is present in lampreys, thus being highly conserved during the evolution of vertebrates, and suggest that it is involved in many brain functions, the only known exception being teleosts. Copyright © 2013 Wiley Periodicals, Inc.

White sucker Catostomus commersonii respond to conspecific and sea lamprey Petromyzon marinus alarm cues but not potential predator cues

Science.gov (United States)

Jordbro, Ethan J.; Di Rocco, Richard T.; Imre, Istvan; Johnson, Nicholas; Brown, Grant E.

2016-01-01

Recent studies proposed the use of chemosensory alarm cues to control the distribution of invasive sea lamprey Petromyzon marinus populations in the Laurentian Great Lakes and necessitate the evaluation of sea lamprey chemosensory alarm cues on valuable sympatric species such as white sucker. In two laboratory experiments, 10 replicate groups (10 animals each) of migratory white suckers were exposed to deionized water (control), conspecific whole-body extract, heterospecific whole-body extract (sea lamprey) and two potential predator cues (2-phenylethylamine HCl (PEA HCl) and human saliva) during the day, and exposed to the first four of the above cues at night. White suckers avoided the conspecific and the sea lamprey whole-body extract both during the day and at night to the same extent. Human saliva did not induce avoidance during the day. PEA HCl did not induce avoidance at a higher concentration during the day, or at night at the minimum concentration that was previously shown to induce maximum avoidance by sea lamprey under laboratory conditions. Our findings suggest that human saliva and PEA HCl may be potential species-specific predator cues for sea lamprey.

Elimination of super(14)C-bisazir residues in adult sea lamprey (Petromyzon marinus)

Science.gov (United States)

Allen, J.L.; Dawson, V.K.

1987-01-01

Bisazir (P.P-bis(1-aziridinyl)-N-methylphosphinothioic amide), a chemosterilant, was administered to sea lampreys (Petromyzon marinus ) by intraperitoneal injection of 100 mg/kg or by immersion for 2 h in a 100- mg/L aqueous solution of the chemical. Whole body analysis of the injected lampreys showed that total residue concentrations decreased to 4.65 in males and 10.07 in females during the first day after injection, and to 1.46 in males and 3.74 in females after 10 days of withdrawal. Lampreys exposed by bath immersion contained residues of about 25 mu g/g of tissue immediately after exposure. The concentration ( mu g/g) decreased to 1.02 in males and 2.11 in females after 1 day of withdrawal and to 0.51 in males and 0.85 in females after 10 days.

Seasonal patterns in growth, blood consumption, and effects on hosts by parasitic-phase sea lampreys in the Great Lakes: an individual-based model approach

Science.gov (United States)

Madenjian, Charles P.; Cochran, Philip A.; Bergstedt, Roger A.

2003-01-01

An individual-based model (IBM) was developed for sea lamprey (Petromyzon marinus) populations in the Laurentian Great Lakes. The IBM was then calibrated to observed growth, by season, for sea lampreys in northern Lake Huron under two different water temperature regimes: a regime experienced by Seneca-strain lake trout (Salvelinus namaycush) and a regime experienced by Marquettestrain lake trout. Modeling results indicated that seasonal blood consumption under the Seneca regime was very similar to that under the Marquette regime. Simulated mortality of lake trout directly due to blood removal by sea lampreys occurred at nearly twice the rate during August and September under the Marquette regime than under the Seneca regime. However, cumulative sea lamprey-induced mortality on lake trout over the entire duration of the sea lamprey's parasitic phase was only 7% higher for the Marquette regime compared with the Seneca regime. Thus, these modeling results indicated that the strain composition of the host (lake trout) population was not important in determining total number of lake trout deaths or total blood consumption attributable to the sea lamprey population, given the sea lamprey growth pattern. Regardless of water temperature regime, both blood consumption rate by sea lampreys and rate of sea lamprey-induced mortality on lake trout peaked in late October. Elevated blood consumption in late October appeared to be unrelated to changes in water temperature. The IBM approach should prove useful in optimizing control of sea lampreys in the Laurentian Great Lakes.

A thermogenic secondary sexual character in male sea lamprey

Science.gov (United States)

Chung-Davidson, Yu-Wen; Priess, M. Cody; Yeh, Chu-Yin; Brant, Cory O.; Johnson, Nicholas S.; Li, Ke; Nanlohy, Kaben G.; Bryan, Mara B.; Brown, C. Titus; Choi, Jongeun; Li, Weiming

2013-01-01

Secondary sexual characters in animals are exaggerated ornaments or weapons for intrasexual competition. Unexpectedly, we found that a male secondary sexual character in sea lamprey (Petromyzon marinus ) is a thermogenic adipose tissue that instantly increases its heat production during sexual encounters. This secondary sexual character, developed in front of the anterior dorsal fin of mature males, is a swollen dorsal ridge known as the ‘rope’ tissue. It contains nerve bundles, multivacuolar adipocytes and interstitial cells packed with small lipid droplets and mitochondria with dense and highly organized cristae. The fatty acid composition of the rope tissue is rich in unsaturated fatty acids. The cytochrome c oxidase activity is high but the ATP concentration is very low in the mitochondria of the rope tissue compared with those of the gill and muscle tissues. The rope tissue temperature immediately rose up to 0.3°C when the male encountered a conspecific. Mature males generated more heat in the rope and muscle tissues when presented with a mature female than when presented with a male (paired t-test, P-3 more heat than the muscle in 10 min. Transcriptome analyses revealed that genes involved in fat cell differentiation are upregulated whereas those involved in oxidative-phosphorylation-coupled ATP synthesis are downregulated in the rope tissue compared with the gill and muscle tissues. Sexually mature male sea lamprey possess the only known thermogenic secondary sexual character that shows differential heat generation toward individual conspecifics.

Growth and survival of sea lampreys from metamorphosis to spawning in Lake Huron

Science.gov (United States)

Swink, William D.; Johnson, Nicholas S.

2014-01-01

Larval Sea Lampreys Petromyzon marinus live burrowed in stream bottoms and then metamorphose into their parasitic stage. Among larvae that metamorphose in a given year (i.e., parasitic cohort), autumn out-migrants (October–December) to the Laurentian Great Lakes can feed on fish for up to 6 months longer than spring outmigrants (March–May), which overwinter in streams without feeding. We evaluated whether the season of outmigration affected growth or survival of newlymetamorphosed Sea Lampreys in LakeHuron. Newlymetamorphosed individuals (n=2,718) from three parasitic cohorts were netted during their out-migration from BlackMallard Creek, Michigan, to LakeHuron during autumn 1997 through spring 2000; each out-migrant was injected with a sequentially numbered coded wire tag and was released back into the creek. After up to 18 months of feeding in the Great Lakes, 224 (8.2%) Sea Lampreys were recaptured (in 1999–2001) as upstream-migrating adults in tributaries to Lakes Huron and Michigan. Recovery rates of autumn and spring out-migrants as adults were 9.4% and 7.8%, respectively, and these rates did not significantly differ. Overwinter feeding (i.e., as parasites) by autumn out-migrants did not produce adult mean sizes greater than those of spring out-migrants. Because we detected no growth or survival differences between autumn and spring out-migrants, the capture of newly metamorphosed Sea Lampreys at any point during their out-migration should provide equal reductions in damage to Great Lakes fisheries. The absence of a difference in growth or survival between autumn and spring out-migrants is an aspect of Sea Lamprey life history that yields resiliency to this invasive parasite and complicates efforts for its control in the Great Lakes.

Affinity Crystallography: A New Approach to Extracting High-Affinity Enzyme Inhibitors from Natural Extracts.

Science.gov (United States)

Aguda, Adeleke H; Lavallee, Vincent; Cheng, Ping; Bott, Tina M; Meimetis, Labros G; Law, Simon; Nguyen, Nham T; Williams, David E; Kaleta, Jadwiga; Villanueva, Ivan; Davies, Julian; Andersen, Raymond J; Brayer, Gary D; Brömme, Dieter

2016-08-26

Natural products are an important source of novel drug scaffolds. The highly variable and unpredictable timelines associated with isolating novel compounds and elucidating their structures have led to the demise of exploring natural product extract libraries in drug discovery programs. Here we introduce affinity crystallography as a new methodology that significantly shortens the time of the hit to active structure cycle in bioactive natural product discovery research. This affinity crystallography approach is illustrated by using semipure fractions of an actinomycetes culture extract to isolate and identify a cathepsin K inhibitor and to compare the outcome with the traditional assay-guided purification/structural analysis approach. The traditional approach resulted in the identification of the known inhibitor antipain (1) and its new but lower potency dehydration product 2, while the affinity crystallography approach led to the identification of a new high-affinity inhibitor named lichostatinal (3). The structure and potency of lichostatinal (3) was verified by total synthesis and kinetic characterization. To the best of our knowledge, this is the first example of isolating and characterizing a potent enzyme inhibitor from a partially purified crude natural product extract using a protein crystallographic approach.

Deep ancestry of programmed genome rearrangement in lampreys.

Science.gov (United States)

Timoshevskiy, Vladimir A; Lampman, Ralph T; Hess, Jon E; Porter, Laurie L; Smith, Jeramiah J

2017-09-01

In most multicellular organisms, the structure and content of the genome is rigorously maintained over the course of development. However some species have evolved genome biologies that permit, or require, developmentally regulated changes in the physical structure and content of the genome (programmed genome rearrangement: PGR). Relatively few vertebrates are known to undergo PGR, although all agnathans surveyed to date (several hagfish and one lamprey: Petromyzon marinus) show evidence of large scale PGR. To further resolve the ancestry of PGR within vertebrates, we developed probes that allow simultaneous tracking of nearly all sequences eliminated by PGR in P. marinus and a second lamprey species (Entosphenus tridentatus). These comparative analyses reveal conserved subcellular structures (lagging chromatin and micronuclei) associated with PGR and provide the first comparative embryological evidence in support of the idea that PGR represents an ancient and evolutionarily stable strategy for regulating inherent developmental/genetic conflicts between germline and soma. Copyright © 2017 Elsevier Inc. All rights reserved.

Phylogeographical analysis reveals multiple conservation units in brook lampreys Lampetra planeri of Portuguese streams.

Science.gov (United States)

Pereira, A M; Robalo, J I; Freyhof, J; Maia, C; Fonseca, J P; Valente, A; Almada, V C

2010-08-01

The populations of brook lamprey Lampetra planeri of Portuguese Rivers were analysed phylogeographically using a fragment of 644 bp of the mitochondrial control region of 158 individuals from six populations. Samples representing L. planeri and migratory lampreys Lampetra fluviatilis of rivers draining to the North Sea and the Baltic Sea were also included to assess the relationships of Portuguese samples. The data support a clear differentiation of all the populations studied. Several populations, which are isolated among themselves and also from the migratory lampreys, proved to be entirely composed of private haplotypes, a finding that supports some time of independent evolutionary history for these populations. This, combined with the geographic confinement to small water bodies, justifies the recognition of at least four conservation units in the Portuguese rivers Sado, São Pedro, Nabão and Inha.

Effects of sea lamprey substrate modification and carcass nutrients on macroinvertebrate assemblages in a small Atlantic coastal stream

Science.gov (United States)

Weaver, Daniel M.; Coghlan, Stephen M.; Zydlewski, Joseph D.

2018-01-01

Aquatic macroinvertebrates respond to patch dynamics arising from interactions of physical and chemical disturbances across space and time. Anadromous fish, such as sea lamprey, Petromyzon marinus, migrate from the ocean and alter physical and chemical properties of recipient spawning streams. Sea lamprey disturb stream benthos physically through nest construction and spawning, and enrich food webs through nutrient deposition from decomposing carcasses. Sea lamprey spawning nests support greater macroinvertebrate abundance than adjacent reference areas, but concurrent effects of stream bed modification and nutrient supplementation have not been examined sequentially. We added carcasses and cleared substrate experimentally to mimic the physical disturbance and nutrient enrichment associated with lamprey spawning, and characterized effects on macroinvertebrate assemblage structure. We found that areas receiving cleared substrate and carcass nutrients were colonized largely by Simuliidae compared to upstream and downstream control areas that were colonized largely by Hydropsychidae, Philopotamidae, and Chironomidae. Environmental factors such as stream flow likely shape assemblages by physically constraining macroinvertebrate establishment and feeding. Our results indicate potential changes in macroinvertebrate assemblages from the physical and chemical changes to streams brought by spawning populations of sea lamprey.

Determining Adult Pacific Lamprey Abundance and Spawning Habitat in the Lower Deschutes River Sub-Basin, Oregon, 2008 Annual Report.

Energy Technology Data Exchange (ETDEWEB)

Fox, Matt; Graham, Jennifer C. [Confederated Tribes of the Warm Springs Reservation, Oregon

2009-04-30

An adult Pacific lamprey (Lampetra tridentata) escapement estimate was generated in the lower Deschutes River during run year 2008. This included a mark-recapture study to determine adult abundance and a tribal subsistence creel. Fish measuring less than 10.5 cm received two marks for the mark-recapture estimate while those measuring greater than 10.5 cm were surgically implanted with radio transmitters to monitor migration upstream of Sherars Falls (rkm 70.4). Radio telemetry was used to determine habitat, focal spawning areas and spawn timing. All fish were collected at the Sherars Falls fish ladder from July-October 2008 using a long handled dip-net. Escapement was generated using a two event mark-recapture experiment. Adult lamprey populations were estimated at 3,471 (95% CI = 2,384-5,041; M = 101; C = 885 R = 25) using Chapman's modification of the Peterson estimate. The relative precision around the estimate was 31.42. Tribal harvest was approximately 806 adult lamprey (95% CI = +/- 74) with a total escapement of 2,669. Fourteen lamprey received radio tags and were released at Lower Blue Hole recreation site (rkm 77.3). Movement was recorded by mobile, fixed site and aerial telemetry methods. Upstream movements of lamprey were documented from July through December 2008 with most lamprey over-wintering in the mainstem Deschutes River.

Occurrence of Ergasilus megaceros Wilson, 1916, in the sea lamprey and other fishes from North America

Science.gov (United States)

Muzzall, Patrick M.; Hudson, Patrick L.

2004-01-01

Ergasilus megaceros (Copepoda: Ergasilidae) was recovered from the nasal fossae (lamellae) of the olfactory sac in 1 (1.8%) of 56 sea lampreys, Petromyzon marinus Linne, 1758, collected in May 2002 from the Cheboygan River, Michigan. Although the sea lamprey is a new host record for E. megaceros, this fish species may not be a preferred host because of its low prevalence. Ergasilus megaceros is the second ergasilid species reported from the sea lamprey in North America. This is the third report of an ergasilid species infecting the nasal fossae of fishes in North America, with E. rhinos being the only other species reported from this site.

Prosomeric map of the lamprey forebrain based on calretinin immunocytochemistry, Nissl stain, and ancillary markers.

Science.gov (United States)

Pombal, M A; Puelles, L

1999-11-22

The structural organization of the lamprey extratelencephalic forebrain is re-examined from the perspective of the prosomeric segmental paradigm. The question asked was whether the prosomeric forebrain model used for gnathostomes is of material advantage for interpreting subdivisions in the lamprey forebrain. To this aim, the main longitudinal and transverse landmarks recognized by the prosomeric model in other vertebrates were identified in Nissl-stained lamprey material. Lines of cytoarchitectural discontinuity and contours of migrated neuronal groups were mapped in a two-dimensional sagittal representation and were also classified according to their radial position. Immunocytochemical mapping of calretinin expression in adjacent sections served to define particular structural units better, in particular, the dorsal thalamus. These data were complemented by numerous other chemoarchitectonic observations obtained with ancillary markers, which identified additional specific formations, subdivisions, or boundaries. Emphasis was placed on studying whether such chemically defined neuronal groups showed boundaries aligned with the postulated inter- or intraprosomeric boundaries. The course of diverse axonal tracts was studied also with regard to their prosomeric topography. This analysis showed that the full prosomeric model applies straightforwardly to the lamprey forebrain. This finding implies that a common segmental and longitudinal organization of the neural tube may be primitive for all vertebrates. Interesting novel aspects appear in the interpretation of the lamprey pretectum, the dorsal and ventral thalami, and the hypothalamus. The topologic continuity of the prosomeric forebrain regions with evaginated or non-evaginated portions of the telencephalon was also examined. Copyright 1999 Wiley-Liss, Inc.

The sterile-male-release technique in Great Lakes sea lamprey management

Science.gov (United States)

Twohey, Michael B.; Heinrich, John W.; Seelye, James G.; Fredricks, Kim T.; Bergstedt, Roger A.; Kaye, Cheryl A.; Scholefield, Ron J.; McDonald, Rodney B.; Christie, Gavin C.

2003-01-01

The implementation of a sterile-male-release technique from 1991 through 1999 and evaluation of its effectiveness in the Great Lakes sea lamprey (Petromyzon marinus) management program is reviewed. Male sea lampreys were injected with the chemosterilant bisazir (P,P-bis(1-aziridinyl)-N-methylphosphinothioic amide) using a robotic device. Quality assurance testing indicated the device delivered a consistent and effective dose of bisazir. Viability of embryos in an untreated control group was 64% compared to 1% in a treatment group. A task force developed nine hypotheses to guide implementation and evaluation of the technique. An annual average of 26,000 male sea lampreys was harvested from as many as 17 Great Lakes tributaries for use in the technique. An annual average of 16,100 sterilized males was released into 33 tributaries of Lake Superior to achieve a theoretical 59% reduction in larval production during 1991 to 1996. The average number of sterile males released in the St. Marys River increased from 4,000 during 1991 to 1996 to 20,100 during 1997 to 1999. The theoretical reduc-stertion in reproduction when combined with trapping was 57% during 1991 to 1996 and 86% during 1997 to 1999. Evaluation studies demonstrated that sterilized males were competitive and reduced production of larvae in streams. Field studies and simulation models suggest reductions in reproduction will result in fewer recruits, but there is risk of periodic high recruitment events independent of sterile-male release. Strategies to reduce reproduction will be most reliable when low densities of reproducing females are achieved. Expansion of the technique is limited by access to additional males for sterilization. Sterile-male release and other alternative controls are important in delivering integrated pest management and in reducing reliance on pesticides.

Does DNA methylation regulate metamorphosis? The case of the sea lamprey (Petromyzon marinus) as an example.

Science.gov (United States)

Covelo-Soto, Lara; Saura, María; Morán, Paloma

2015-07-01

Lampreys represent one of the most ancient vertebrate lineages enclosing a special interest for genetic and epigenetic studies. The sea lamprey (Petromyzon marinus) is an anadromous species that experiences metamorphosis all the way up to the adult stage. Although representing a gradual process, metamorphosis in this species involves dramatic conversions with regard to physiological together with structural body changes preparing individuals for a marine and parasitic life; in consequence, multiple gene expression modifications are expected. The implications of thyroid hormones and HOX gene expression changes have previously been reported in this species and also in other vertebrate species. Nonetheless, information lacks on how these genes are regulated in lampreys. We here report about the existence of methylation pattern differences between the adult and the larvae sea lamprey life cycle stages making use of the Methylation-Sensitive Amplified Polymorphism (MSAP) technique. Differentially methylated fragment sequencing allowed to establish homologous identities with HOX genes involved in morphogenesis, along with genes related to the water balance and to the osmotic homoeostasis, all associated to a marine environment adaptation. These results provide evidences revealing that DNA methylation plays a role in the epigenetic regulation of the P. marinus post-natal development representing a starting point for future studies. To the best of our knowledge, this is the first study which detects DNA methylation changes associated with metamorphosis in lampreys. Copyright © 2015 Elsevier Inc. All rights reserved.

Estimating reach-specific fish movement probabilities in rivers with a Bayesian state-space model: application to sea lamprey passage and capture at dams

Science.gov (United States)

Holbrook, Christopher M.; Johnson, Nicholas S.; Steibel, Juan P.; Twohey, Michael B.; Binder, Thomas R.; Krueger, Charles C.; Jones, Michael L.

2014-01-01

Improved methods are needed to evaluate barriers and traps for control and assessment of invasive sea lamprey (Petromyzon marinus) in the Great Lakes. A Bayesian state-space model provided reach-specific probabilities of movement, including trap capture and dam passage, for 148 acoustic tagged invasive sea lamprey in the lower Cheboygan River, Michigan, a tributary to Lake Huron. Reach-specific movement probabilities were combined to obtain estimates of spatial distribution and abundance needed to evaluate a barrier and trap complex for sea lamprey control and assessment. Of an estimated 21 828 – 29 300 adult sea lampreys in the river, 0%–2%, or 0–514 untagged lampreys, could have passed upstream of the dam, and 46%–61% were caught in the trap. Although no tagged lampreys passed above the dam (0/148), our sample size was not sufficient to consider the lock and dam a complete barrier to sea lamprey. Results also showed that existing traps are in good locations because 83%–96% of the population was vulnerable to existing traps. However, only 52%–69% of lampreys vulnerable to traps were caught, suggesting that traps can be improved. The approach used in this study was a novel use of Bayesian state-space models that may have broader applications, including evaluation of barriers for other invasive species (e.g., Asian carp (Hypophthalmichthys spp.)) and fish passage structures for other diadromous fishes.

Evaluating harvest-based control of invasive fish with telemetry: Performance of sea lamprey traps in the Great Lakes

Science.gov (United States)

Holbrook, Christopher; Bergstedt, Roger A.; Barber, Jessica M.; Bravener, Gale A; Jones, Michael L.; Krueger, Charles C.

2016-01-01

Physical removal (e.g., harvest via traps or nets) of mature individuals may be a cost-effective or socially acceptable alternative to chemical control strategies for invasive species, but requires knowledge of the spatial distribution of a population over time. We used acoustic telemetry to determine the current and possible future role of traps to control and assess invasive sea lampreys, Petromyzon marinus, in the St. Marys River, the connecting channel between Lake Superior and Lake Huron. Exploitation rates (i.e., fractions of an adult sea lamprey population removed by traps) at two upstream locations were compared among three years and two points of entry to the system. Telemetry receivers throughout the drainage allowed trap performance (exploitation rate) to be partitioned into two components: proportion of migrating sea lampreys that visited trap sites (availability) and proportion of available sea lampreys that were caught by traps (local trap efficiency). Estimated exploitation rates were well below those needed to provide population control in the absence of lampricides and were limited by availability and local trap efficiency. Local trap efficiency estimates for acoustic-tagged sea lampreys were lower than analogous estimates regularly obtained using traditional mark–recapture methods, suggesting that abundance had been previously underestimated. Results suggested major changes would be required to substantially increase catch, including improvements to existing traps, installation of new traps, or other modifications to attract and retain more sea lampreys. This case study also shows how bias associated with telemetry tags can be estimated and incorporated in models to improve inferences about parameters that are directly relevant to fishery management.

Strategies for swimming: explorations of the behaviour of a neuro-musculo-mechanical model of the lamprey

Directory of Open Access Journals (Sweden)

Thelma L. Williams

2015-02-01

Full Text Available Experiments were performed on a neuro-musculo-mechanical model of a lamprey, to explore the strategies for controlling swimming speed. The muscle component of the model was based on previous experiments on isolated lamprey muscle. The patterns of muscle activation were those found in EMG studies on swimming lampreys. The fluid mechanics were modelled with G.I. Taylor's simplification. Tail beat frequencies of 2–6 sec−1 were combined with muscle activation strengths of 0.1% to 20% of maximum tetanic isometric strength. The resulting forward swimming speed and changing body shape were recorded. From the changing body shape the speed of the backward-travelling wave of curvature was calculated, as well as the ratio between the speeds of the waves of activation and curvature. For any given activation strength there was a tail beat frequency that gave maximal forward speed. Furthermore, for all the combinations of activation strength and tail beat frequency that gave such maximum swimming speeds, the ratio of the speed of the wave of curvature to the wave of muscle activation was approximately 0.75. This is similar to the ratio found in swimming lampreys.

15alpha-Hydroxyprogesterone in male sea lampreys, Petromyzon marinus L

Czech Academy of Sciences Publication Activity Database

Bryan, M. B.; Scott, A. P.; Černý, Ivan; Young, B. A.; Li, W.

2004-01-01

Roč. 69, - (2004), s. 473-481 ISSN 0039-128X Institutional research plan: CEZ:AV0Z4055905 Keywords : steroid * 15alpha-hydroxyprogesterone * sea lamprey Subject RIV: CC - Organic Chemistry Impact factor: 2.337, year: 2004

Regulation of a putative corticosteroid, 17, 21-dihydroxypregn-4-ene, 3, 20-one, in sea lamprey, Petromyzon marinus.

Science.gov (United States)

Roberts, Brent W.; Didier, Wes; Satbir, Rai; Johnson, Nicholas S.; Libants, Scot V.; Sang-Seon, Yun; Close, David

2013-01-01

In higher vertebrates, in response to stress, the hypothalamus produces corticotropin-releasing hormone (CRH), which stimulates cells in the anterior pituitary to produce adrenocorticotropic hormone (ACTH), which in turn stimulates production of either cortisol (F) or corticosterone (B) by the adrenal tissues. In lampreys, however, neither of these steroids is present. Instead, it has been proposed that the stress steroid is actually 17,21-dihydroxypregn-4-ene-3,20-dione (11-deoxycortisol; S). However, there have been no studies yet to determine its mechanism of regulation or site of production. Here we demonstrate that (1) intraperitoneal injections of lamprey-CRH increase plasma S in a dose dependent manner, (2) intraperitoneal injections of four lamprey-specific ACTH peptides at 100 lg/kg, did not induce changes in plasma S concentrations in either males or females; (3) two lamprey-specific gonadotropin-releasing hormones (GnRH I and III) and arginine-vasotocin (AVT), all at single doses, stimulated S production as well as, or to an even greater extent than CRH; (4) sea lamprey mesonephric kidneys, in vitro, converted tritiated 17a-hydroxyprogesterone (17a-P) into a steroid that had the same chromatographic properties (on HPLC and TLC) as S; (5) kidney tissues released significantly more immunoassayable S into the incubation medium than gill, liver or gonad tissues. One interpretation of these results is that the corticosteroid production of the sea lamprey, one of the oldest extant vertebrates, is regulated through multiple pathways rather than the classical HPI-axis. However, the responsiveness of this steroid to the GnRH peptides means that a reproductive rather than a stress role for this steroid cannot yet be ruled out.

Test of a non-physical barrier consisting of light, sound, and bubble screen to block upstream movement of sea lamprey in an experimental raceway

Science.gov (United States)

Miehls, Scott M.; Johnson, Nicholas S.; Hrodey, Pete J.

2017-01-01

Control of the invasive Sea Lamprey Petromyzon marinus is critical for management of commercial and recreational fisheries in the Laurentian Great Lakes. Use of physical barriers to block Sea Lampreys from spawning habitat is a major component of the control program. However, the resulting interruption of natural streamflow and blockage of nontarget species present substantial challenges. Development of an effective nonphysical barrier would aid the control of Sea Lampreys by eliminating their access to spawning locations while maintaining natural streamflow. We tested the effect of a nonphysical barrier consisting of strobe lights, low-frequency sound, and a bubble screen on the movement of Sea Lampreys in an experimental raceway designed as a two-choice maze with a single main channel fed by two identical inflow channels (one control and one blocked). Sea Lampreys were more likely to move upstream during trials when the strobe light and low-frequency sound were active compared with control trials and trials using the bubble screen alone. For those Sea Lampreys that did move upstream to the confluence of inflow channels, no combination of stimuli or any individual stimulus significantly influenced the likelihood that Sea Lampreys would enter the blocked inflow channel, enter the control channel, or return downstream.

A sea lamprey (Petromyzon marinus) sex pheromone mixture increases trap catch relative to a single synthesized component in specific environments

Science.gov (United States)

Johnson, Nicholas S.; Tix, John A.; Hlina, Benjamin L.; Wagner, C. Michael; Siefkes, Michael J.; Wang, Huiyong; Li, Weiming

2015-01-01

Spermiating male sea lamprey (Petromyzon marinus) release a sex pheromone, of which a component, 7α, 12α, 24-trihydoxy-3-one-5α-cholan-24-sulfate (3kPZS), has been identified and shown to induce long distance preference responses in ovulated females. However, other pheromone components exist, and when 3kPZS alone was used to control invasive sea lamprey populations in the Laurentian Great Lakes, trap catch increase was significant, but gains were generally marginal. We hypothesized that free-ranging sea lamprey populations discriminate between a partial and complete pheromone while migrating to spawning grounds and searching for mates at spawning grounds. As a means to test our hypothesis, and to test two possible uses of sex pheromones for sea lamprey control, we asked whether the full sex pheromone mixture released by males (spermiating male washings; SMW) is more effective than 3kPZS in capturing animals in traditional traps (1) en route to spawning grounds and (2) at spawning grounds. At locations where traps target sea lampreys en route to spawning grounds, SMW-baited traps captured significantly more sea lampreys than paired 3kPZS-baited traps (~10 % increase). At spawning grounds, no difference in trap catch was observed between 3kPZS and SMW-baited traps. The lack of an observed difference at spawning grounds may be attributed to increased pheromone competition and possible involvement of other sensory modalities to locate mates. Because fishes often rely on multiple and sometimes redundant sensory modalities for critical life history events, the addition of sex pheromones to traditionally used traps is not likely to work in all circumstances. In the case of the sea lamprey, sex pheromone application may increase catch when applied to specifically designed traps deployed in streams with low adult density and limited spawning habitat.

An anti-steroidogenic inhibitory primer pheromone in male sea lamprey (Petromyzon marinus)

Science.gov (United States)

Chung-Davidson, Yu-Wen; Wang, Huiyong; Bryan, Mara B.; Wu, Hong; Johnson, Nicholas S.; Li, Weiming

2013-01-01

Reproductive functions can be modulated by both stimulatory and inhibitory primer pheromones released by conspecifics. Many stimulatory primer pheromones have been documented, but relatively few inhibitory primer pheromones have been reported in vertebrates. The sea lamprey male sex pheromone system presents an advantageous model to explore the stimulatory and inhibitory primer pheromone functions in vertebrates since several pheromone components have been identified. We hypothesized that a candidate sex pheromone component, 7α, 12α-dihydroxy-5α-cholan-3-one-24-oic acid (3 keto-allocholic acid or 3kACA), exerts priming effects through the hypothalamic-pituitary-gonadal (HPG) axis. To test this hypothesis, we measured the peptide concentrations and gene expressions of lamprey gonadotropin releasing hormones (lGnRH) and the HPG output in immature male sea lamprey exposed to waterborne 3kACA. Exposure to waterborne 3kACA altered neuronal activation markers such as jun and jun N-terminal kinase (JNK), and lGnRH mRNA levels in the brain. Waterborne 3kACA also increased lGnRH-III, but not lGnRH-I or -II, in the forebrain. In the plasma, 3kACA exposure decreased all three lGnRH peptide concentrations after 1 h exposure. After 2 h exposure, 3kACA increased lGnRHI and -III, but decreased lGnRH-II peptide concentrations in the plasma. Plasma lGnRH peptide concentrations showed differential phasic patterns. Group housing condition appeared to increase the averaged plasma lGnRH levels in male sea lamprey compared to isolated males. Interestingly, 15α-hydroxyprogesterone (15α-P) concentrations decreased after prolonged 3kACA exposure (at least 24 h). To our knowledge, this is the only known synthetic vertebrate pheromone component that inhibits steroidogenesis in males.

Quantifying high-affinity binding of hydrophobic ligands by isothermal titration calorimetry.

Science.gov (United States)

Krainer, Georg; Broecker, Jana; Vargas, Carolyn; Fanghänel, Jörg; Keller, Sandro

2012-12-18

A fast and reliable quantification of the binding thermodynamics of hydrophobic high-affinity ligands employing a new calorimetric competition experiment is described. Although isothermal titration calorimetry is the method of choice for a quantitative characterization of intermolecular interactions in solution, a reliable determination of a dissociation constant (K(D)) is typically limited to the range 100 μM > K(D) > 1 nM. Interactions displaying higher or lower K(D) values can be assessed indirectly, provided that a suitable competing ligand is available whose K(D) falls within the directly accessible affinity window. This established displacement assay, however, requires the high-affinity ligand to be soluble at high concentrations in aqueous buffer and, consequently, poses serious problems in the study of protein binding involving small-molecule ligands dissolved in organic solvents--a familiar case in many drug-discovery projects relying on compound libraries. The calorimetric competition assay introduced here overcomes this limitation, thus allowing for a detailed thermodynamic description of high-affinity receptor-ligand interactions involving poorly water-soluble compounds. Based on a single titration of receptor into a dilute mixture of the two competing ligands, this competition assay provides accurate and precise values for the dissociation constants and binding enthalpies of both high- and moderate-affinity ligands. We discuss the theoretical background underlying the approach, demonstrate its practical application to metal ion chelation and high-affinity protein-inhibitor interactions, and explore its potential and limitations with the aid of simulations and statistical analyses.

Migratory behavior of adult sea lamprey and cumulative passage performance through four fishways

Science.gov (United States)

Castro-Santos, Theodore R.; Shi, Xiaotao; Haro, Alexander

2017-01-01

This article describes a study of PIT-tagged sea lamprey (Petromyzon marinus) ascending four fishways comprising three designs at two dams on the Connecticut River, USA. Migration between dams was rapid (median migration rate = 23 km·day−1). Movement through the fishways was much slower, however (median = 0.02–0.33 km·day−1). Overall delay at dams was substantial (median = 13.6–14.6 days); many fish failed to pass (percent passage ranged from 29% to 55%, depending on fishway), and repeated passage attempts compounded delay for both passers and failers. Cox regression revealed that fishway entry rates were influenced by flow, temperature, and diel cycle, with most lampreys entering at night and at elevated flows, but with no apparent effect of sex or length. Overall delay was influenced by slow movement through the fishways, but repeated failures were the primary factor determining delay. These data suggest that although some lamprey were able to pass fishways, they did so with difficulty, and delays incurred as they attempted to pass may act to limit their distribution within their native range.

High affinity hemoglobin and Parkinson's disease.

Science.gov (United States)

Graham, Jeffrey; Hobson, Douglas; Ponnampalam, Arjuna

2014-12-01

Parkinson's disease (PD) is a neurodegenerative disorder characterized by the loss of dopaminergic neurons in the substantia nigra (SN) region of the midbrain. Oxidative damage in this region has been shown to play an important role in the pathogenesis of this disease. Human neurons have been discovered to contain hemoglobin, with an increased concentration seen in the neurons of the SN. High affinity hemoglobin is a clinical entity resulting from mutations that create a functional increase in the binding of hemoglobin to oxygen and an inability to efficiently unload it to tissues. This can result in a number of metabolic compensatory changes, including an elevation in circulating hemoglobin and an increase in the molecule 2,3-diphosphoglycerate (2,3-DPG). Population based studies have revealed that patients with PD have elevated hemoglobin as well as 2,3-DPG levels. Based on these observations, we hypothesize that the oxidative damage seen in PD is related to an underlying high affinity hemoglobin subtype. Copyright © 2014 Elsevier Ltd. All rights reserved.

Metabolism of a sea lamprey pesticide by fish liver enzymes part A: identification and synthesis of TFM metabolites.

Science.gov (United States)

Bussy, Ugo; Chung-Davidson, Yu-Wen; Buchinger, Tyler; Li, Ke; Smith, Scott A; Jones, A Daniel; Li, Weiming

2018-02-01

The sea lamprey (Petromyzon marinus) is a destructive invasive species in the Great Lakes that contributed to the collapse of native fish populations in the mid-1900s. 3-Trifluoromethyl-4-nitrophenol (TFM) is a selective pesticide that has been applied to sea lamprey infested tributaries of the Great Lakes to kill larvae since the 1960s and has reduced the populations by as much as 90%. However, the metabolism of TFM by sea lamprey and non-target species is not fully illuminated. Elucidation of TFM metabolism is critical for understanding its mode of action and possible environmental impact. Here, we describe the screening, identification, synthesis and structural characterization of TFM metabolites in livers from sea lamprey and three non-target species that differ in their ability to survive TFM exposure. We identified glucuronidation, sulfation, N-acetylation, glutathione conjugation, and aromatic nitro group reduction as potential detoxification mechanisms. Seven metabolites were synthesized for use as markers of TFM metabolism in fish. Quantitative 1 H NMR was used to assay synthesized metabolite stock solutions that were then used as standard material to develop a quantitative LC-MS/MS method for TFM metabolites.

The lamprey: a jawless vertebrate model system for examining origin of the neural crest and other vertebrate traits.

Science.gov (United States)

Green, Stephen A; Bronner, Marianne E

2014-01-01

Lampreys are a group of jawless fishes that serve as an important point of comparison for studies of vertebrate evolution. Lampreys and hagfishes are agnathan fishes, the cyclostomes, which sit at a crucial phylogenetic position as the only living sister group of the jawed vertebrates. Comparisons between cyclostomes and jawed vertebrates can help identify shared derived (i.e. synapomorphic) traits that might have been inherited from ancestral early vertebrates, if unlikely to have arisen convergently by chance. One example of a uniquely vertebrate trait is the neural crest, an embryonic tissue that produces many cell types crucial to vertebrate features, such as the craniofacial skeleton, pigmentation of the skin, and much of the peripheral nervous system (Gans and Northcutt, 1983). Invertebrate chordates arguably lack unambiguous neural crest homologs, yet have cells with some similarities, making comparisons with lampreys and jawed vertebrates essential for inferring characteristics of development in early vertebrates, and how they may have evolved from nonvertebrate chordates. Here we review recent research on cyclostome neural crest development, including research on lamprey gene regulatory networks and differentiated neural crest fates. Copyright © 2014 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

Engineering of bispecific affinity proteins with high affinity for ERBB2 and adaptable binding to albumin.

Directory of Open Access Journals (Sweden)

Johan Nilvebrant

Full Text Available The epidermal growth factor receptor 2, ERBB2, is a well-validated target for cancer diagnostics and therapy. Recent studies suggest that the over-expression of this receptor in various cancers might also be exploited for antibody-based payload delivery, e.g. antibody drug conjugates. In such strategies, the full-length antibody format is probably not required for therapeutic effect and smaller tumor-specific affinity proteins might be an alternative. However, small proteins and peptides generally suffer from fast excretion through the kidneys, and thereby require frequent administration in order to maintain a therapeutic concentration. In an attempt aimed at combining ERBB2-targeting with antibody-like pharmacokinetic properties in a small protein format, we have engineered bispecific ERBB2-binding proteins that are based on a small albumin-binding domain. Phage display selection against ERBB2 was used for identification of a lead candidate, followed by affinity maturation using second-generation libraries. Cell surface display and flow-cytometric sorting allowed stringent selection of top candidates from pools pre-enriched by phage display. Several affinity-matured molecules were shown to bind human ERBB2 with sub-nanomolar affinity while retaining the interaction with human serum albumin. Moreover, parallel selections against ERBB2 in the presence of human serum albumin identified several amino acid substitutions that dramatically modulate the albumin affinity, which could provide a convenient means to control the pharmacokinetics. The new affinity proteins competed for ERBB2-binding with the monoclonal antibody trastuzumab and recognized the native receptor on a human cancer cell line. Hence, high affinity tumor targeting and tunable albumin binding were combined in one small adaptable protein.

High-throughput fragment screening by affinity LC-MS.

Science.gov (United States)

Duong-Thi, Minh-Dao; Bergström, Maria; Fex, Tomas; Isaksson, Roland; Ohlson, Sten

2013-02-01

Fragment screening, an emerging approach for hit finding in drug discovery, has recently been proven effective by its first approved drug, vemurafenib, for cancer treatment. Techniques such as nuclear magnetic resonance, surface plasmon resonance, and isothemal titration calorimetry, with their own pros and cons, have been employed for screening fragment libraries. As an alternative approach, screening based on high-performance liquid chromatography separation has been developed. In this work, we present weak affinity LC/MS as a method to screen fragments under high-throughput conditions. Affinity-based capillary columns with immobilized thrombin were used to screen a collection of 590 compounds from a fragment library. The collection was divided into 11 mixtures (each containing 35 to 65 fragments) and screened by MS detection. The primary screening was performed in 3500 fragments per day). Thirty hits were defined, which subsequently entered a secondary screening using an active site-blocked thrombin column for confirmation of specificity. One hit showed selective binding to thrombin with an estimated dissociation constant (K (D)) in the 0.1 mM range. This study shows that affinity LC/MS is characterized by high throughput, ease of operation, and low consumption of target and fragments, and therefore it promises to be a valuable method for fragment screening.

Flexibility in the patterning and control of axial locomotor networks in lamprey.

Science.gov (United States)

Buchanan, James T

2011-12-01

In lower vertebrates, locomotor burst generators for axial muscles generally produce unitary bursts that alternate between the two sides of the body. In lamprey, a lower vertebrate, locomotor activity in the axial ventral roots of the isolated spinal cord can exhibit flexibility in the timings of bursts to dorsally-located myotomal muscle fibers versus ventrally-located myotomal muscle fibers. These episodes of decreased synchrony can occur spontaneously, especially in the rostral spinal cord where the propagating body waves of swimming originate. Application of serotonin, an endogenous spinal neurotransmitter known to presynaptically inhibit excitatory synapses in lamprey, can promote decreased synchrony of dorsal-ventral bursting. These observations suggest the possible existence of dorsal and ventral locomotor networks with modifiable coupling strength between them. Intracellular recordings of motoneurons during locomotor activity provide some support for this model. Pairs of motoneurons innervating myotomal muscle fibers of similar ipsilateral dorsoventral location tend to have higher correlations of fast synaptic activity during fictive locomotion than do pairs of motoneurons innervating myotomes of different ipsilateral dorsoventral locations, suggesting their control by different populations of premotor interneurons. Further, these different motoneuron pools receive different patterns of excitatory and inhibitory inputs from individual reticulospinal neurons, conveyed in part by different sets of premotor interneurons. Perhaps, then, the locomotor network of the lamprey is not simply a unitary burst generator on each side of the spinal cord that activates all ipsilateral body muscles simultaneously. Instead, the burst generator on each side may comprise at least two coupled burst generators, one controlling motoneurons innervating dorsal body muscles and one controlling motoneurons innervating ventral body muscles. The coupling strength between these two

Mixtures of Two Bile Alcohol Sulfates Function as a Proximity Pheromone in Sea Lamprey.

Directory of Open Access Journals (Sweden)

Cory O Brant

Full Text Available Unique mixtures of pheromone components are commonly identified in insects, and have been shown to increase attractiveness towards conspecifics when reconstructed at the natural ratio released by the signaler. In previous field studies of pheromones that attract female sea lamprey (Petromyzon marinus, L., putative components of the male-released mating pheromone included the newly described bile alcohol 3,12-diketo-4,6-petromyzonene-24-sulfate (DkPES and the well characterized 3-keto petromyzonol sulfate (3kPZS. Here, we show chemical evidence that unequivocally confirms the elucidated structure of DkPES, electrophysiological evidence that each component is independently detected by the olfactory epithelium, and behavioral evidence that mature female sea lamprey prefer artificial nests activated with a mixture that reconstructs the male-released component ratio of 30:1 (3kPZS:DkPES, molar:molar. In addition, we characterize search behavior (sinuosity of swim paths of females approaching ratio treatment sources. These results suggest unique pheromone ratios may underlie reproductive isolating mechanisms in vertebrates, as well as provide utility in pheromone-integrated control of invasive sea lamprey in the Great Lakes.

A pheromone outweighs temperature in influencing migration of sea lamprey

Science.gov (United States)

Brant, Cory O.; Li, Ke; Johnson, Nicholas S.; Li, Weiming

2015-01-01

Organisms continuously acquire and process information from surrounding cues. While some cues complement one another in delivering more reliable information, others may provide conflicting information. How organisms extract and use reliable information from a multitude of cues is largely unknown. We examined movement decisions of sea lampreys (Petromyzon marinus L.) exposed to a conspecific and an environmental cue during pre-spawning migration. Specifically, we predicted that the mature male-released sex pheromone 3-keto petromyzonol sulfate (3kPZS) will outweigh the locomotor inhibiting effects of cold stream temperature (less than 15°C). Using large-scale stream bioassays, we found that 3kPZS elicits an increase (more than 40%) in upstream movement of pre-spawning lampreys when the water temperatures were below 15°C. Both warming temperatures and conspecific cues increase upstream movement when the water temperature rose above 15°C. These patterns define an interaction between abiotic and conspecific cues in modulating animal decision-making, providing an example of the hierarchy of contradictory information.

Proadifen-sensitive high affinity binding of 3H-alaproclate to liver membranes

International Nuclear Information System (INIS)

Ross, S.B.

1987-01-01

3 H-alaproclate, a selective 5 h ydroxytryptamine uptake inhibitor, was found to bind to microsomal membranes from the rat liver with high affinity (K D -=3 nM) and large capacity (B max about 2 nmol/g liver). This binding was stereoselective since S-( - )-alaproclate was 30 times more potent than the R-( + )-enantiomer to displace the 3 H-labelled racemate. Proadifen (SKF 525A), an inhibitor of cytochrome P-450, displaced the 3 H-alaproclate binding with the same, high affinity (K i =3 nM) as alaproclate itself. Repeated treatment with phenobarbital sodium (5x75 mg/kg intraperitoneally) increased the number of alaproclate binding sites 7-8 times without changing the affinity. However, most of the phenobarbital induced 3 H-alaproclate binding was not displaceable by proadifen, showing the presence of at least two different high affinity binding sites. The possible involvement of cytochrome P-450 in the alaproclate binding is discussed. (author)

Structure of Greyhound hemoglobin: origin of high oxygen affinity.

Science.gov (United States)

Bhatt, Veer S; Zaldívar-López, Sara; Harris, David R; Couto, C Guillermo; Wang, Peng G; Palmer, Andre F

2011-05-01

This study presents the crystal structure of Greyhound hemoglobin (GrHb) determined to 1.9 Å resolution. GrHb was found to crystallize with an α₁β₁ dimer in the asymmetric unit and belongs to the R2 state. Oxygen-affinity measurements combined with the fact that GrHb crystallizes in the R2 state despite the high-salt conditions used for crystallization strongly indicate that GrHb can serve as a model high-oxygen-affinity hemoglobin (Hb) for higher mammals, especially humans. Structural analysis of GrHb and its comparison with the R2-state of human Hb revealed several regions that can potentially contribute to the high oxygen affinity of GrHb and serve to rationalize the additional stability of the R2-state of GrHb. A previously well studied hydrophobic cluster of bar-headed goose Hb near α119 was also incorporated in the comparison between GrHb and human Hb. Finally, a structural comparison with generic dog Hb and maned wolf Hb was conducted, revealing that in contrast to GrHb these structures belong to the R state of Hb and raising the intriguing possibility of an additional allosteric factor co-purifying with GrHb that can modulate its quaternary structure.

Reconstitution of high-affinity opioid agonist binding in brain membranes

Energy Technology Data Exchange (ETDEWEB)

Remmers, A.E.; Medzihradsky, F. (Univ. of Michigan Medical School, Ann Arbor (United States))

1991-03-15

In synaptosomal membranes from rat brain cortex, the {mu} selective agonist ({sup 3}H)dihydromorphine in the absence of sodium, and the nonselective antagonist ({sup 3}H)naltrexone in the presence of sodium, bound to two populations of opioid receptor sites with K{sub d} values of 0.69 and 8.7 nM for dihydromorphine, and 0.34 and 5.5 nM for naltrexone. The addition of 5 {mu}M guanosine 5{prime}-({gamma}-thio)triphosphate (GTP({gamma}S)) strongly reduced high-affinity agonist but not antagonist binding. Exposure of the membranes to high pH reduced the number of GTP({gamma}-{sup 35}S) binding sites by 90% and low K{sub m}, opioid-sensitive GTPase activity by 95%. In these membranes, high-affinity agonist binding was abolished and modulation of residual binding by GTP({gamma}S) was diminished. Alkali treatment of the glioma cell membranes prior to fusion inhibited most of the low K{sub m} GTPase activity and prevented the reconstitution of agonist binding. The results show that high-affinity opioid agonist binding reflects the ligand-occupied receptor - guanine nucleotide binding protein complex.

Haemoglobin Pierre-Benite--a high affinity variant associated with relative polycythaemia.

Science.gov (United States)

Beard, M E; Potter, H C; Spearing, R L; Brennan, S O

2001-12-01

This is the second reported example of Hb Pierre--Benite (beta90 Glu-->Asp). This mutation is associated with increased oxygen affinity and polycythaemia. No instability was found and there was no charge shift detected by cellulose acetate electrophoresis at pH 8.3. The mutation was however, clearly indicated by electrospray ionization mass spectrometry (ESI MS), which showed an abnormal beta chain with a 14 Da decrease in mass. Blood volume studies documented a relative rather than a true polycythaemia and this finding has been reported in at least two other high affinity haemoglobin variants--Hb Heathrow and Hb Rahere. This finding led to delay in diagnosis because high oxygen affinity variants are conventionally considered to cause a true polycythaemia.

High affinity binding of [3H]cocaine to rat liver microsomes

International Nuclear Information System (INIS)

El-Maghrabi, E.A.; Calligaro, D.O.; Eldefrawi, M.E.

1988-01-01

] 3 H]cocaine bound reversible, with high affinity and stereospecificity to rat liver microsomes. Little binding was detected in the lysosomal, mitochondrial and nuclear fractions. The binding kinetics were slow and the kinetically calculated K/sub D/ was 2 nM. Induction of mixed function oxidases by phenobarbital did not produce significant change in [ 3 H]cocaine binding. On the other hand, chronic administration of cocaine reduced [ 3 H]cocaine binding drastically. Neither treatment affected the affinity of the liver binding protein for cocaine. Microsomes from mouse and human livers had less cocaine-binding protein and lower affinity for cocaine than those from rat liver. Binding of [ 3 H]cocaine to rat liver microsomes was insensitive to monovalent cations and > 10 fold less sensitive to biogenic amines than the cocaine receptor in rat striatum. However, the liver protein had higher affinity for cocaine and metabolites except for norcocaine. Amine uptake inhibitors displaced [ 3 H]cocaine binding to liver with a different rank order of potency than their displacement of [ 3 H]cocaine binding to striatum. This high affinity [ 3 H]cocaine binding protein in liver is not likely to be monooxygenase, but may have a role in cocaine-induced hepatotoxicity

Characterization of a novel bile alcohol sulfate released by sexually mature male sea lamprey (Petromyzon marinus.

Directory of Open Access Journals (Sweden)

Ke Li

Full Text Available A sulphate-conjugated bile alcohol, 3,12-diketo-4,6-petromyzonene-24-sulfate (DKPES, was identified using bioassay-guided fractionation from water conditioned with sexually mature male sea lamprey (Petromyzon marinus. The structure and relative stereochemistry of DKPES was established using spectroscopic data. The electro-olfactogram (EOG response threshold of DKPES was 10(-7 Molar (M and that of 3-keto petromyzonol sulfate (3 KPZS; a known component of the male sea lamprey sex pheromone was 10(-10 M. Behavioural studies indicated that DKPES can be detected at low concentrations by attracting sexually mature females to nests when combined with 3 KPZS. Nests baited with a mixture of DKPES and 3 KPZS (ratio 1∶29.8 attracted equal numbers of sexually mature females compared to an adjacent nest baited with 3 KPZS alone. When DKPES and 3 KPZS mixtures were applied at ratios of 2∶29.8 and 10∶29.8, the proportion of sexually mature females that entered baited nests increased to 73% and 70%, respectively. None of the sexually mature females released were attracted to nests baited with DKPES alone. These results indicated that DKPES is a component of the sex pheromone released by sexually mature male sea lamprey, and is the second biologically active compound identified from this pheromone. DKPES represents the first example that a minor component of a vertebrate pheromone can be combined with a major component to elicit critical sexual behaviors. DKPES holds considerable promise for increasing the effectiveness of pheromone-baited trapping as a means of sea lamprey control in the Laurentian Great Lakes.

Influence of body size, metabolic rate and life history stage on the uptake and excretion of the lampricide 3-trifluoromethyl-4-nitrophenol (TFM) by invasive sea lampreys (Petromyzon marinus).

Science.gov (United States)

Tessier, Laura R; Long, Tristan A F; Wilkie, Michael P

2018-01-01

Invasive sea lamprey (Petromyzon marinus) are controlled in the Great Lakes using the lampricide 3-trifluoromethyl-4-nitrophenol (TFM), which is applied to streams infested with larval lamprey. However, lamprey that survive treatments (residuals) remain a challenge because they may subsequently undergo metamorphosis into parasitic juvenile animals that migrate downstream to the Great Lakes, where they feed on important sport and commercial fishes. The goal of this study was to determine if body size and life stage could potentially influence sea lamprey tolerance to TFM by influencing patterns of TFM uptake and elimination. Because mass specific rates of oxygen consumption (M˙O 2 ) are lower in larger compared to smaller lamprey, we predicted that TFM uptake would be negatively correlated to body size, suggesting that large larvae would be more tolerant to TFM exposure. Accordingly, TFM uptake and M˙O 2 were measured in larvae ranging in size from 0.2-4.2g using radio-labelled TFM ( 14 C-TFM) and static respirometry. Both were inversely proportional to wet mass (M), and could be described usingthe allometric power relationship: Y=aM b , in which M˙O 2 =1.86M 0.53 and TFM Uptake=7.24M 0.34 . We also predicted that body size would extend to rates of TFM elimination, which was measured following the administration of 14 C-TFM (via intraperitoneal injection). However, there were no differences in the half-lives of elimination of TFM (T 1/2 -TFM). There were also no differences in M˙O 2 or TFM uptake amongst size-matched larval, metamorphosing (stages 6-7), or post-metamorphic (juvenile) sea lamprey. However, the T 1/2 -TFM was significantly lower in larval than post-metamorphic lamprey (juvenile), indicating the larval lamprey cleared TFM more efficiently than juvenile lamprey. We conclude that larger larval sea lamprey are more likely to survive TFM treatments suggesting that body size might be an important variable to consider when treating streams with TFM to

Cloning and analysis of an HMG gene from the lamprey Lampetra fluviatilis

DEFF Research Database (Denmark)

Sharman, A C; Hay-Schmidt, Anders; Holland, P W

1997-01-01

Evolution has shaped the organisation of vertebrate genomes, including the human genome. To shed further light on genome history, we have cloned and analysed an HMG gene from lamprey, representing one of the earliest vertebrate lineages. Genes of the HMG1/2 family encode chromosomal proteins...

Retinal amino acid neurochemistry of the southern hemisphere lamprey, Geotria australis.

Directory of Open Access Journals (Sweden)

Lisa Nivison-Smith

Full Text Available Lampreys are one of the two surviving groups of the agnathan (jawless stages in vertebrate evolution and are thus ideal candidates for elucidating the evolution of visual systems. This study investigated the retinal amino acid neurochemistry of the southern hemisphere lamprey Geotria australis during the downstream migration of the young, recently-metamorphosed juveniles to the sea and during the upstream migration of the fully-grown and sexually-maturing adults to their spawning areas. Glutamate and taurine were distributed throughout the retina, whilst GABA and glycine were confined to neurons of the inner retina matching patterns seen in most other vertebrates. Glutamine and aspartate immunoreactivity was closely matched to Müller cell morphology. Between the migratory phases, few differences were observed in the distribution of major neurotransmitters i.e. glutamate, GABA and glycine, but changes in amino acids associated with retinal metabolism i.e. glutamine and aspartate, were evident. Taurine immunoreactivity was mostly conserved between migrant stages, consistent with its role in primary cell functions such as osmoregulation. Further investigation of glutamate signalling using the probe agmatine (AGB to map cation channel permeability revealed entry of AGB into photoreceptors and horizontal cells followed by accumulation in inner retinal neurons. Similarities in AGB profiles between upstream and downstream migrant of G. australis confirmed the conservation of glutamate neurotransmission. Finally, calcium binding proteins, calbindin and calretinin were localized to the inner retina whilst recoverin was localized to photoreceptors. Overall, conservation of major amino acid neurotransmitters and calcium-associated proteins in the lamprey retina confirms these elements as essential features of the vertebrate visual system. On the other hand, metabolic elements of the retina such as neurotransmitter precursor amino acids and Müller cells

Effects of coded-wire-tagging on stream-dwelling Sea Lamprey larvae

Science.gov (United States)

Johnson, Nicholas; Swink, William D.; Dawson, Heather A.; Jones, Michael L.

2016-01-01

The effects of coded wire tagging Sea Lamprey Petromyzon marinus larvae from a known-aged stream-dwelling population were assessed. Tagged larvae were significantly shorter on average than untagged larvae from 3 to 18 months after tagging. However, 30 months after tagging, the length distribution of tagged and untagged larvae did not differ and tagged Sea Lampreys were in better condition (i.e., higher condition factor) and more likely to have undergone metamorphosis than the untagged population. The reason why tagged larvae were more likely to metamorphose is not clear, but the increased likelihood of metamorphosis could have been a compensatory response to the period of slower growth after tagging. Slower growth after tagging was consistent across larval size-classes, so handling and displacement from quality habitat during the early part of the growing season was likely the cause rather than the tag burden. The tag effects observed in this study, if caused by displacement and handling, may be minimized in future studies if tagging is conducted during autumn after growth has concluded for the year.

Analysis of DNA damage in sea lamprey (Petromyzon marinus) spermatozoa by UV, hydrogen peroxide, and the toxicant bisazir

International Nuclear Information System (INIS)

Ciereszko, Andrzej; Wolfe, Tobie D.; Dabrowski, Konrad

2005-01-01

In this study we sought to demonstrate that Comet assay can be applied to sea lamprey sperm DNA fragmentation and used to describe the relationship between sperm DNA damage and sperm fertilizing ability. We show that the assay can be used reliably and accurately, and unlike in the case of mammals, there is no need for additional steps related to improvement of efficacy of lysis and DNA decondensation. This agrees with the presence of histone proteins in lamprey sperm. An increase in DNA fragmentation was noted during short-term storage of milt on ice (0-4 days). We demonstrated genotoxic effects of UV radiation and oxidative stress (exposure to hydrogen peroxide) and found that oxidative damage to sperm DNA was likely repaired after fertilization in the embryo. Repairing capacity of the oocyte toward sperm DNA lesions caused by UV was restricted. Toxic effect of p,p-bis-(1-aziridinyl)-N-methylphosphinothioic acid (p,p-bis(1-aziridinyl)-N-methylphosphinothioic amide), a sea lamprey chemosterilant, could not be linked to DNA fragmentation in the in vitro tests. Its genotoxicity in vivo may possibly be associated with other mechanisms of DNA degradation (oxidation or DNA-protein and DNA-DNA cross-linking). In conclusion, this study demonstrates that Comet assay can be successfully applied to monitor effects of environmental disturbances and imposed injuries in sea lamprey spermatozoa and possibly other species of ancient fish with acrosomal sperm

Analysis of DNA damage in sea lamprey (Petromyzon marinus) spermatozoa by UV, hydrogen peroxide, and the toxicant bisazir

Energy Technology Data Exchange (ETDEWEB)

Ciereszko, Andrzej [School of Natural Resources, Ohio State University, 210 Kottman Hall, 2021 Coffey Rd., Columbus, OH 434210 (United States); Semen Biology Group, Institute of Animal Reproduction and Food Research, Polish Academy of Sciences, 10-747 Olsztyn (Poland); Wolfe, Tobie D. [School of Natural Resources, Ohio State University, 210 Kottman Hall, 2021 Coffey Rd., Columbus, OH 434210 (United States); Dabrowski, Konrad [School of Natural Resources, Ohio State University, 210 Kottman Hall, 2021 Coffey Rd., Columbus, OH 434210 (United States)]. E-mail: dabrowski.1@osu.edu

2005-06-15

In this study we sought to demonstrate that Comet assay can be applied to sea lamprey sperm DNA fragmentation and used to describe the relationship between sperm DNA damage and sperm fertilizing ability. We show that the assay can be used reliably and accurately, and unlike in the case of mammals, there is no need for additional steps related to improvement of efficacy of lysis and DNA decondensation. This agrees with the presence of histone proteins in lamprey sperm. An increase in DNA fragmentation was noted during short-term storage of milt on ice (0-4 days). We demonstrated genotoxic effects of UV radiation and oxidative stress (exposure to hydrogen peroxide) and found that oxidative damage to sperm DNA was likely repaired after fertilization in the embryo. Repairing capacity of the oocyte toward sperm DNA lesions caused by UV was restricted. Toxic effect of p,p-bis-(1-aziridinyl)-N-methylphosphinothioic acid (p,p-bis(1-aziridinyl)-N-methylphosphinothioic amide), a sea lamprey chemosterilant, could not be linked to DNA fragmentation in the in vitro tests. Its genotoxicity in vivo may possibly be associated with other mechanisms of DNA degradation (oxidation or DNA-protein and DNA-DNA cross-linking). In conclusion, this study demonstrates that Comet assay can be successfully applied to monitor effects of environmental disturbances and imposed injuries in sea lamprey spermatozoa and possibly other species of ancient fish with acrosomal sperm.

Genetic diversity, endemism and phylogeny of lampreys within the genus Lampetra sensu stricto (Petromyzontiformes: Petromyzontidae) in western North America.

Science.gov (United States)

Boguski, D A; Reid, S B; Goodman, D H; Docker, M F

2012-11-01

Phylogenetic structure of four Lampetra species from the Pacific drainage of North America (western brook lamprey Lampetra richardsoni, Pacific brook lamprey Lampetra pacifica, river lamprey Lampetra ayresii and Kern brook lamprey Lampetra hubbsi) and unidentified Lampetra specimens (referred to as Lampetra sp.) from 36 locations was estimated using the mitochondrial cytochrome b gene. Maximum parsimony and Bayesian inferences did not correspond with any taxonomic scheme proposed to date. Rather, although L. richardsoni (from Alaska to California) and L. ayresii (from British Columbia to California) together constituted a well-supported clade distinct from several genetically divergent Lampetra populations in Oregon and California, these two species were not reciprocally monophyletic. The genetically divergent populations included L. pacifica (from the Columbia River basin) and L. hubbsi (from the Kern River basin) and four Lampetra sp. populations in Oregon (Siuslaw River and Fourmile Creek) and California (Kelsey and Mark West Creeks). These four Lampetra sp. populations showed genetic divergence between 2.3 and 5.7% from any known species (and up to 8.0% from each other), and may represent morphologically cryptic and thus previously undescribed species. A fifth population (from Paynes Creek, California) may represent a range extension of L. hubbsi into the Upper Sacramento River. © 2012 The Authors. Journal of Fish Biology © 2012 The Fisheries Society of the British Isles.

Evolution of vertebrate adaptive immunity: immune cells and tissues, and AID/APOBEC cytidine deaminases.

Science.gov (United States)

Hirano, Masayuki

2015-08-01

All surviving jawed vertebrate representatives achieve diversity in immunoglobulin-based B and T cell receptors for antigen recognition through recombinatorial rearrangement of V(D)J segments. However, the extant jawless vertebrates, lampreys and hagfish, instead generate three types of variable lymphocyte receptors (VLRs) through a template-mediated combinatorial assembly of different leucine-rich repeat (LRR) sequences. The clonally diverse VLRB receptors are expressed by B-like lymphocytes, while the VLRA and VLRC receptors are expressed by lymphocyte lineages that resemble αβ and γδ T lymphocytes, respectively. These findings suggest that three basic types of lymphocytes, one B-like and two T-like, are an essential feature of vertebrate adaptive immunity. Around 500 million years ago, a common ancestor of jawed and jawless vertebrates evolved a genetic program for the development of prototypic lymphoid cells as a foundation for an adaptive immune system. This acquisition preceded the convergent evolution of alternative types of clonally diverse receptors for antigens in all vertebrates, as reviewed in this article. © 2015 WILEY Periodicals, Inc.

Neural crest contributions to the lamprey head

Science.gov (United States)

McCauley, David W.; Bronner-Fraser, Marianne

2003-01-01

The neural crest is a vertebrate-specific cell population that contributes to the facial skeleton and other derivatives. We have performed focal DiI injection into the cranial neural tube of the developing lamprey in order to follow the migratory pathways of discrete groups of cells from origin to destination and to compare neural crest migratory pathways in a basal vertebrate to those of gnathostomes. The results show that the general pathways of cranial neural crest migration are conserved throughout the vertebrates, with cells migrating in streams analogous to the mandibular and hyoid streams. Caudal branchial neural crest cells migrate ventrally as a sheet of cells from the hindbrain and super-pharyngeal region of the neural tube and form a cylinder surrounding a core of mesoderm in each pharyngeal arch, similar to that seen in zebrafish and axolotl. In addition to these similarities, we also uncovered important differences. Migration into the presumptive caudal branchial arches of the lamprey involves both rostral and caudal movements of neural crest cells that have not been described in gnathostomes, suggesting that barriers that constrain rostrocaudal movement of cranial neural crest cells may have arisen after the agnathan/gnathostome split. Accordingly, neural crest cells from a single axial level contributed to multiple arches and there was extensive mixing between populations. There was no apparent filling of neural crest derivatives in a ventral-to-dorsal order, as has been observed in higher vertebrates, nor did we find evidence of a neural crest contribution to cranial sensory ganglia. These results suggest that migratory constraints and additional neural crest derivatives arose later in gnathostome evolution.

Neither philopatric nor panmictic: microsatellite and mtDNA evidence suggests lack of natal homing but limits to dispersal in Pacific lamprey.

Science.gov (United States)

Spice, Erin K; Goodman, Damon H; Reid, Stewart B; Docker, Margaret F

2012-06-01

Most species with lengthy migrations display some degree of natal homing; some (e.g. migratory birds and anadromous salmonids) show spectacular feats of homing. However, studies of the sea lamprey (Petromyzon marinus) indicate that this anadromous species locates spawning habitat based on pheromonal cues from larvae rather than through philopatry. Previous genetic studies in the anadromous Pacific lamprey (Entosphenus tridentatus) have both supported and rejected the hypothesis of natal homing. To resolve this, we used nine microsatellite loci to examine the population structure in 965 Pacific lamprey from 20 locations from central British Columbia to southern California and supplemented this analysis with mitochondrial DNA restriction fragment length polymorphism analysis on a subset of 530 lamprey. Microsatellite analysis revealed (i) relatively low but often statistically significant genetic differentiation among locations (97% pairwise F(ST) values were <0.04 but 73.7% were significant); and (ii) weak but significant isolation by distance (r(2) = 0.0565, P = 0.0450) but no geographic clustering of samples. The few moderate F(ST) values involved comparisons with sites that were geographically distant or far upstream. The mtDNA analysis--although providing less resolution among sites (only 4.7%F(ST) values were significant)--was broadly consistent with the microsatellite results: (i) the southernmost site and some sites tributary to the Salish Sea were genetically distinct; and (ii) southern sites showed higher haplotype and private haplotype richness. These results are inconsistent with philopatry, suggesting that anadromous lampreys are unusual among species with long migrations, but suggest that limited dispersal at sea precludes panmixia in this species. © 2012 Blackwell Publishing Ltd.

Proadifen-sensitive high affinity binding of /sup 3/H-alaproclate to liver membranes

Energy Technology Data Exchange (ETDEWEB)

Ross, S.B.

1987-01-01

/sup 3/H-alaproclate, a selective 5/sub h/ydroxytryptamine uptake inhibitor, was found to bind to microsomal membranes from the rat liver with high affinity (K/sub D/-=3 nM) and large capacity (B/sub max/ about 2 nmol/g liver). This binding was stereoselective since S-( - )-alaproclate was 30 times more potent than the R-( + )-enantiomer to displace the /sup 3/H-labelled racemate. Proadifen (SKF 525A), an inhibitor of cytochrome P-450, displaced the /sup 3/H-alaproclate binding with the same, high affinity (K/sub i/=3 nM) as alaproclate itself. Repeated treatment with phenobarbital sodium (5x75 mg/kg intraperitoneally) increased the number of alaproclate binding sites 7-8 times without changing the affinity. However, most of the phenobarbital induced /sup 3/H-alaproclate binding was not displaceable by proadifen, showing the presence of at least two different high affinity binding sites. The possible involvement of cytochrome P-450 in the alaproclate binding is discussed.

The monoclonal S9.6 antibody exhibits highly variable binding affinities towards different R-loop sequences.

Directory of Open Access Journals (Sweden)

Fabian König

Full Text Available The monoclonal antibody S9.6 is a widely-used tool to purify, analyse and quantify R-loop structures in cells. A previous study using the surface plasmon resonance technology and a single-chain variable fragment (scFv of S9.6 showed high affinity (0.6 nM for DNA-RNA and also a high affinity (2.7 nM for RNA-RNA hybrids. We used the microscale thermophoresis method allowing surface independent interaction studies and electromobility shift assays to evaluate additional RNA-DNA hybrid sequences and to quantify the binding affinities of the S9.6 antibody with respect to distinct sequences and their GC-content. Our results confirm high affinity binding to previously analysed sequences, but reveals that binding affinities are highly sequence specific. Our study presents R-loop sequences that independent of GC-content and in different sequence variations exhibit either no binding, binding affinities in the micromolar range and as well high affinity binding in the nanomolar range. Our study questions the usefulness of the S9.6 antibody in the quantitative analysis of R-loop sequences in vivo.

Survival and metamorphosis of low-density populations of larval sea lampreys (Petromyzon marinus) in streams following lampricide treatment

Science.gov (United States)

Johnson, Nicholas S.; Swink, William D.; Brenden, Travis O.; Slade, Jeffrey W.; Steeves, Todd B.; Fodale, Michael F.; Jones, Michael L.

2014-01-01

Sea lamprey Petromyzon marinus control in the Great Lakes primarily involves application of lampricides to streams where larval production occurs to kill larvae prior to their metamorphosing and entering the lakes as parasites (juveniles). Because lampricides are not 100% effective, larvae that survive treatment maymetamorphose before streams are again treated. Larvae that survive treatment have not beenwidely studied, so their dynamics are notwell understood.Wetagged and released larvae in six Great Lake tributaries following lampricide treatment and estimated vital demographic rates using multistate tag-recovery models. Model-averaged larval survivals ranged from 56.8 to 57.6%. Model-averaged adult recovery rates, which were the product of juvenile survivals and adult capture probabilities, ranged from 6.8 to 9.3%. Using stochastic simulations, we estimated production of juvenile sea lampreys from a hypothetical population of treatment survivors under different growth conditions based on parameter estimates from this research. For fast-growing populations, juvenile production peaked 2 years after treatment. For slow-growing populations, juvenile production was approximately one-third that of fast-growing populations,with production not peaking until 4 years after treatment. Our results suggest that dynamics (i.e., survival, metamorphosis) of residual larval populations are very similar to those of untreated larval populations. Consequently, residual populations do not necessarily warrant special consideration for the purpose of sea lamprey control and can be ranked for treatment along with other populations. Consecutive lampricide treatments, which are under evaluation by the sea lamprey control program, would bemost effective for reducing juvenile production in large, fast-growing populations.

Evolution of the vertebrate claudin gene family: insights from a basal vertebrate, the sea lamprey.

Science.gov (United States)

Mukendi, Christian; Dean, Nicholas; Lala, Rushil; Smith, Jeramiah; Bronner, Marianne E; Nikitina, Natalya V

2016-01-01

Claudins are major constituents of tight junctions, contributing both to their intercellular sealing and selective permeability properties. While claudins and claudin-like molecules are present in some invertebrates, the association of claudins with tight junctions has been conclusively documented only in vertebrates. Here we report the sequencing, phylogenetic analysis and comprehensive spatiotemporal expression analysis of the entire claudin gene family in the basal extant vertebrate, the sea lamprey. Our results demonstrate that clear orthologues to about half of all mammalian claudins are present in the lamprey, suggesting that at least one round of whole genome duplication contributed to the diversification of this gene family. Expression analysis revealed that claudins are expressed in discrete and specific domains, many of which represent vertebrate-specific innovations, such as in cranial ectodermal placodes and the neural crest; whereas others represent structures characteristic of chordates, e.g. pronephros, notochord, somites, endostyle and pharyngeal arches. By comparing the embryonic expression of claudins in the lamprey to that of other vertebrates, we found that ancestral expression patterns were often preserved in higher vertebrates. Morpholino mediated loss of Cldn3b demonstrated a functional role for this protein in placode and pharyngeal arch morphogenesis. Taken together, our data provide novel insights into the origins and evolution of the claudin gene family and the significance of claudin proteins in the evolution of vertebrates.

A simple, cost-effective emitter for controlled release of fish pheromones: development, testing, and application to management of the invasive sea lamprey

Science.gov (United States)

Wagner, Michael C.; Hanson, James E.; Meckley, Trevor D.; Johnson, Nicholas; Bals, Jason D.

2018-01-01

Semiochemicals that elicit species-specific attraction or repulsion have proven useful in the management of terrestrial pests and hold considerable promise for control of nuisance aquatic species, particularly invasive fishes. Because aquatic ecosystems are typically large and open, use of a semiochemical to control a spatially dispersed invader will require the development of a cost-effective emitter that is easy to produce, environmentally benign, inexpensive, and controls the release of the semiochemical without altering its structure. We examined the release properties of five polymers, and chose polyethylene glycol (PEG) as the best alternative. In a series of laboratory and field experiments, we examined the response of the invasive sea lamprey to PEG, and to a partial sex pheromone emitted from PEG that has proven effective as a trap bait to capture migrating sea lamprey prior to spawning. Our findings confirm that the sea lamprey does not behaviorally respond to PEG, and that the attractant response to the pheromone component was conserved when emitted from PEG. Further, we deployed the pheromone-PEG emitters as trap bait during typical control operations in three Great Lakes tributaries, observing similar improvements in trap performance when compared to a previous study using mechanically pumped liquid pheromone. Finally, the polymer emitters tended to dissolve unevenly in high flow conditions. We demonstrate that housing the emitter stabilizes the dissolution rate at high water velocity. We conclude the performance characteristics of PEG emitters to achieve controlled-release of a semiochemical are sufficient to recommend its use in conservation and management activities related to native and invasive aquatic organisms.

The role of CH/π interactions in the high affinity binding of streptavidin and biotin.

Science.gov (United States)

Ozawa, Motoyasu; Ozawa, Tomonaga; Nishio, Motohiro; Ueda, Kazuyoshi

2017-08-01

The streptavidin-biotin complex has an extraordinarily high affinity (Ka: 10 15 mol -1 ) and contains one of the strongest non-covalent interactions known. This strong interaction is widely used in biological tools, including for affinity tags, detection, and immobilization of proteins. Although hydrogen bond networks and hydrophobic interactions have been proposed to explain this high affinity, the reasons for it remain poorly understood. Inspired by the deceased affinity of biotin observed for point mutations of streptavidin at tryptophan residues, we hypothesized that a CH/π interaction may also contribute to the strong interaction between streptavidin and biotin. CH/π interactions were explored and analyzed at the biotin-binding site and at the interface of the subunits by the fragment molecular orbital method (FMO) and extended applications: PIEDA and FMO4. The results show that CH/π interactions are involved in the high affinity for biotin at the binding site of streptavidin. We further suggest that the involvement of CH/π interactions at the subunit interfaces and an extended CH/π network play more critical roles in determining the high affinity, rather than involvement at the binding site. Copyright © 2017 Elsevier Inc. All rights reserved.

Identification of high- and low-affinity NGF receptors during development of the chicken central nervous system

International Nuclear Information System (INIS)

Escandon, E.; Chao, M.V.

1990-01-01

In order to study regulation of the nerve growth factor (NGF) receptor during embryogenesis in chick brain, we have used affinity crosslinking of tissues with 125 I-NGF. NGF interacts with high- and low-affinity receptors; high-affinity receptors are required for the majority of NGF's actions. Most measurements of receptor levels do not distinguish between high- and low-affinity forms of the receptor. We have used the lipophilic crosslinking agent HSAB to identify the high-affinity, functional receptor during development of the chicken central nervous system. A peak of expression during Embryonic Days 5-10 was detected in all regions of the chicken central nervous system, but, shortly after birth, only the cerebellar region displays significant levels of NGF receptor protein. The time course of expression confirms the dramatic regulation of the NGF receptor gene during defined embryonic periods. The detection of high-affinity NGF receptors in brain and neural retina provides strong evidence that NGF is involved in essential ontogenetic events in the development of the chicken central nervous system

High affinity calmodulin target sequence in the signalling molecule PI 3-kinase

DEFF Research Database (Denmark)

Fischer, R; Julsgart, J; Berchtold, M W

1998-01-01

M-binding peptide derived from the p110gamma isoform interacts with CaM in a calcium-dependent way. Using gel shift analysis and fluorescence spectrophotometry we discovered that the peptide forms a high affinity complex with CaM. Titration experiments using dansylated CaM gave an affinity constant of 5 n...

Reconnaissance of contaminants in larval Pacific lamprey (Entosphenus tridentatus) tissues and habitats in the Columbia River Basin, Oregon and Washington, USA

Science.gov (United States)

Nilsen, Elena B.; Hapke, Whitney B.; McIlraith, Brian; Markovchick, Dennis J.

2015-01-01

Pacific lampreys (Entosphenus tridentatus) have resided in the Columbia River Basin for millennia and have great ecological and cultural importance. The role of habitat contamination in the recent decline of the species has rarely been studied and was the main objective of this effort. A wide range of contaminants (115 analytes) was measured in sediments and tissues at 27 sites across a large geographic area of diverse land use. This is the largest dataset of contaminants in habitats and tissues of Pacific lamprey in North America and the first study to compare contaminant bioburden during the larval life stage and the anadromous, adult portion of the life cycle. Bioaccumulation of pesticides, flame retardants, and mercury was observed at many sites. Based on available data, contaminants are accumulating in larval Pacific lamprey at levels that are likely detrimental to organism health and may be contributing to the decline of the species.

Genetic models reveal historical patterns of sea lamprey population fluctuations within Lake Champlain

Directory of Open Access Journals (Sweden)

Cassidy C. D’Aloia

2015-10-01

Full Text Available The origin of sea lamprey (Petromyzon marinus in Lake Champlain has been heavily debated over the past decade. Given the lack of historical documentation, two competing hypotheses have emerged in the literature. First, it has been argued that the relatively recent population size increase and concomitant rise in wounding rates on prey populations are indicative of an invasive population that entered the lake through the Champlain Canal. Second, recent genetic evidence suggests a post-glacial colonization at the end of the Pleistocene, approximately 11,000 years ago. One limitation to resolving the origin of sea lamprey in Lake Champlain is a lack of historical and current measures of population size. In this study, the issue of population size was explicitly addressed using nuclear (nDNA and mitochondrial DNA (mtDNA markers to estimate historical demography with genetic models. Haplotype network analysis, mismatch analysis, and summary statistics based on mtDNA noncoding sequences for NCI (479 bp and NCII (173 bp all indicate a recent population expansion. Coalescent models based on mtDNA and nDNA identified two potential demographic events: a population decline followed by a very recent population expansion. The decline in effective population size may correlate with land-use and fishing pressure changes post-European settlement, while the recent expansion may be associated with the implementation of the salmonid stocking program in the 1970s. These results are most consistent with the hypothesis that sea lamprey are native to Lake Champlain; however, the credibility intervals around parameter estimates demonstrate that there is uncertainty regarding the magnitude and timing of past demographic events.

Molecular basis of a high affinity murine interleukin-5 receptor.

OpenAIRE

Devos, R; Plaetinck, G; Van der Heyden, J; Cornelis, S; Vandekerckhove, J; Fiers, W; Tavernier, J

1991-01-01

The mouse interleukin-5 receptor (mIL-5R) consists of two components one of which, the mIL-5R alpha-chain, binds mIL-5 with low affinity. Recently we demonstrated that monoclonal antibodies (Mabs) recognizing the second mIL-5R beta-chain, immunoprecipitate a p130-140 protein doublet which corresponds to the mIL-3R and the mIL-3R-like protein, the latter chain for which so far no ligand has been identified. In this study we show that a high affinity mIL-5R can be reconstituted on COS1 cells by...

Single-experiment displacement assay for quantifying high-affinity binding by isothermal titration calorimetry.

Science.gov (United States)

Krainer, Georg; Keller, Sandro

2015-04-01

Isothermal titration calorimetry (ITC) is the gold standard for dissecting the thermodynamics of a biomolecular binding process within a single experiment. However, reliable determination of the dissociation constant (KD) from a single titration is typically limited to the range 100 μM>KD>1 nM. Interactions characterized by a lower KD can be assessed indirectly by so-called competition or displacement assays, provided that a suitable competitive ligand is available whose KD falls within the directly accessible window. However, this protocol is limited by the fact that it necessitates at least two titrations to characterize one high-affinity inhibitor, resulting in considerable consumption of both sample material and time. Here, we introduce a fast and efficient ITC displacement assay that allows for the simultaneous characterization of both a high-affinity ligand and a moderate-affinity ligand competing for the same binding site on a receptor within a single experiment. The protocol is based on a titration of the high-affinity ligand into a solution containing the moderate-affinity ligand bound to the receptor present in excess. The resulting biphasic binding isotherm enables accurate and precise determination of KD values and binding enthalpies (ΔH) of both ligands. We discuss the theoretical background underlying the approach, demonstrate its practical application to metal ion chelation, explore its potential and limitations with the aid of simulations and statistical analyses, and elaborate on potential applications to protein-inhibitor interactions. Copyright © 2014 Elsevier Inc. All rights reserved.

Quantification of a male sea lamprey pheromone in tributaries of Laurentian Great Lakes by liquid chromatography-tandem mass spectrometry

Science.gov (United States)

Xi, X.; Johnson, N.S.; Brant, C.O.; Yun, S.-S.; Chambers, K.L.; Jones, A.D.; Li, W.

2011-01-01

We developed an assay for measuring 7α,12α,24-trihydroxy-5a-cholan-3-one-24-sulfate (3kPZS), a mating pheromone released by male sea lampreys (Petromyzon marinus), at low picomolar concentrations in natural waters to assess the presence of invasive populations. 3kPZS was extracted from streamwater at a rate of recovery up to 90% using a single cation-exchange and reversed-phase mixed-mode cartridge, along with [2H5]3kPZS as an internal standard, and quantified using ultrahigh performance liquid chromatography-tandem mass spectrometry. The limit of detection was below 0.1 ng L–1 (210 fM), which was the lowest concentration tested. Intra- and interday coefficients of variation were between 0.3–11.6% and 4.8–9.8%, respectively, at 1 ng 3kPZS L–1 and 5 ng 3kPZS L–1. This assay was validated by repeat measurements of water samples from a stream spiked with synthesized 3kPZS to reach 4.74 ng L–1 or 0.24 ng L–1. We further verified the utility of this assay to detect spawning populations of lampreys; in the seven tributaries to the Laurentian Great Lakes sampled, 3kPZS concentrations were found to range between 0.15 and 2.85 ng L–1 during the spawning season in known sea lamprey infested segments and were not detectable in uninfested segments. The 3kPZS assay may be useful for the integrated management of sea lamprey, an invasive species in the Great Lakes where pheromone-based control and assessment techniques are desired.

Selective induction of high-ouabain-affinity isoform of Na+-K+-ATPase by thyroid hormone

International Nuclear Information System (INIS)

Haber, R.S.; Loeb, J.N.

1988-01-01

The administration of thyroid hormone is known to result in an induction of the Na + -K + -adenosinetriphosphatase (Na + -K + -ATPase) in rat skeletal muscle and other thyroid hormone-responsive tissues. Since the Na + -K + -ATPase in a variety of mammalian tissues has recently been reported to exist in at least two forms distinguishable by differing affinities for the inhibitory cardiac glycoside ouabain. The authors have studied the effects of 3,3',5-triiodo-L-thyronine (T 3 ) treatment on these two forms of the enzyme in rat diaphragm. The inhibition of Na + -K + -ATPase activity in a crude membrane fraction by varying concentrations of ouabain conformed to a biphasic pattern consistent with the presence of two distinct isoforms with inhibition constants (K I s) for ouabain of ∼10 -7 and 10 -4 M, respectively. Measurement of the specific binding of [ 3 H]ouabain to these membranes confirmed the presence of a class of high-affinity ouabain binding sites with a dissociation constant (K d ) of slightly less than 10 -7 M, whose maximal binding capacity was increased by T 3 treatment by 185%. Binding studies in unfractionated homogenates of diaphragm similarly demonstrated the presence of high-affinity sites whose maximal binding capacity was increased by T 3 treatment. Quantitation of both the high- and low-ouabain-affinity forms of the Na + -K + -ATPase by ouabain-dependent phosphorylation from [ 32 P]orthophosphate confirmed that T 3 treatment markedly increased the number of high-affinity sites while having little effect on the number of low-affinity sites. These observations provide, to our knowledge, the first demonstration that these two forms of the Na + -K + -ATPase are subject to selective hormonal induction

High Performance Affinity Chromatography of Antithrombin III Based on Monodisperse Poly (glycidyl methacrylate) Beads

Institute of Scientific and Technical Information of China (English)

无

2001-01-01

A new approach for the separation of antithrombin III with high performance affinity chromatography (HPAC) was described. A novel monodisperse,non-porous,cross-linked poly (glycidyl methacrylate) beads (PGMA) were used as the affinity support. With the water-soluble carbodiimide,heparin was linked covalently to amino-PGMA-beads,which was prepared by amination of PGMA. The adsorbent obtained exhibits high binding activity to antithrombin III (ATIII),good resolution and excellent mechanical properties and can be used under high flow rate.

Robotic high-throughput purification of affinity-tagged recombinant proteins.

Science.gov (United States)

Wiesler, Simone C; Weinzierl, Robert O J

2015-01-01

Affinity purification of recombinant proteins has become the method of choice to obtain good quantities and qualities of proteins for a variety of downstream biochemical applications. While manual or FPLC-assisted purification techniques are generally time-consuming and labor-intensive, the advent of high-throughput technologies and liquid handling robotics has simplified and accelerated this process significantly. Additionally, without the human factor as a potential source of error, automated purification protocols allow for the generation of large numbers of proteins simultaneously and under directly comparable conditions. The delivered material is ideal for activity comparisons of different variants of the same protein. Here, we present our strategy for the simultaneous purification of up to 24 affinity-tagged proteins for activity measurements in biochemical assays. The protocol described is suitable for the scale typically required in individual research laboratories.

Fatty acid and drug binding to a low-affinity component of human serum albumin, purified by affinity chromatography

DEFF Research Database (Denmark)

Vorum, H; Pedersen, A O; Honoré, B

1992-01-01

Binding equilibria for decanoate to a defatted, commercially available human serum albumin preparation were investigated by dialysis exchange rate determinations. The binding isotherm could not be fitted by the general binding equation. It was necessary to assume that the preparation was a mixture...... of two albumin components about 40% of the albumin having high affinity and about 60% having low affinity. By affinity chromatography we succeeded in purifying the low-affinity component from the mixture. The high-affinity component, however, could not be isolated. We further analyzed the fatty acid...... and drug binding abilities of the low-affinity component. The fatty acids decanoate, laurate, myristate and palmitate were bound with higher affinity to the mixture than to the low-affinity component. Diazepam was bound with nearly the same affinity to the low-affinity component as to the albumin mixture...

Characteristics of high affinity and low affinity adenosine binding sites in human cerebral cortex

International Nuclear Information System (INIS)

John, D.; Fox, I.V.

1986-01-01

The binding characteristics of human brain cortical membrane fractions were evaluated to test the hypothesis that there are A 1 and A 2 adenosine binding sites. The ligands used were 2-chloro(8- 3 H) adenosine and N 6 -(adenine-2, 8- 3 H) cyclohexayladenosine. Binding of chloroadenosine to human brain cortical membranes was time dependent, reversible and concentration dependent. The kinetic constant determinations from binding studies of the adenosine receptor are presented. Utilizing tritium-cyclohexyladenosine as ligand the authors observed evidence for a high affinity binding site in human brain cortical membranes with a kd of 5 nM

Characterization of high-affinity (/sup 3/H)ouabain binding in the rat central nervous system

Energy Technology Data Exchange (ETDEWEB)

Hauger, R.; Luu, H.M.; Meyer, D.K.; Goodwin, F.K.; Paul, S.M.

1985-06-01

The characteristics of (/sup 3/H)ouabain binding were examined in various areas of rat brain. In the striatum, Scatchard analysis revealed a single class of high-affinity binding sites with an apparent binding affinity (KD) of 10.4 +/- 0.9 nM and an estimated binding capacity (Bmax) of 7.6 +/- 1.9 pmol/mg protein. Similar monophasic Scatchard plots were found in the brainstem, cerebellum, hypothalamus, and frontal cerebral cortex. (/sup 3/H)Ouabain binding to rat brain was sodium- and ATP-dependent and strongly inhibited by potassium. Proscillariden A was the most potent cardiac glycoside tested in inhibiting specific (/sup 3/H)ouabain binding to brain membranes, and the rank order of inhibitory potencies for a series of cardiac glycosides was similar to that previously reported for inhibition of heart Na,K-ATPase. To assess whether the high-affinity binding sites for (/sup 3/H)ouabain were localized to neuronal or nonneuronal membranes, the effect of discrete kainic acid lesions on striatal (/sup 3/H)ouabain binding was examined. Kainic acid lesions of the striatum reduced (/sup 3/H)ouabain binding to striatal homogenates by 79.6 +/- 1.6%. This suggests that the high-affinity (/sup 3/H)ouabain binding sites measured in our experiments are localized to neuronal elements. Thus, the high-affinity binding of (/sup 3/H)ouabain to brain membranes may selectively label a neuronal form or conformation of Na,K-ATPase.

The influence of acclimation temperature on the lipid composition of the larval lamprey, Petromyzon marinus, depends on tissue and lipid class.

Science.gov (United States)

Kao, Yung-Hsi; Sheridan, Mark A; Holmes, John A; Youson, John H

2010-11-01

This study was designed to examine the effect of thermal acclimation on the lipid composition of fat depot organs the liver and kidneys of larval sea lamprey, Petromyzon marinus. We found that 21 °C-acclimated larvae possessed lower total lipid amounts in the liver (39% lower) and kidneys (30% lower) than 13 °C-acclimated larvae. Relatively lower lipid contents in the liver and kidneys of 21 °C-acclimated lamprey primarily resulted from a reduction in stored lipid reserve, triacylglycerol, but not the structural lipid, phospholipid. Compared to 21 °C-acclimated larvae, 13 °C-acclimated larvae were found to possess fewer saturated fatty acids (SFAs) and more unsaturated fatty acids (USFAs) in renal triacylglycerol and phospholipid classes, while there were no significant differences in the SFAs and USFAs of hepatic triacylglycerol, phospholipid, cholesteryl ester, fatty acid, and monoacylglycerol classes. Fewer SFAs, found in the kidney triacylglycerol of 13 °C-acclimated lamprey, were due to lower 12:0 and 14:0 fatty acids, but those in the renal phospholipid class were characterized by fewer 14:0, 15:0, and 16:0 fatty acids. More USFAs in renal triacylglycerol, as indicated by a higher unsaturation index, primarily resulted from higher polyunsaturated fatty acids (18:2ω6, 18:3ω3, and 18:4ω3); whereas, in the renal phospholipid class, this was a result of higher monoenes (18:1, 20:1, and 22:1ω9) and ω3 polyunsaturated fatty acids (18:4ω3). These data suggest that the influence of thermal acclimation on the lipid composition of lamprey fat depot organs depends on tissue and lipid class.

Selective high-affinity polydentate ligands and methods of making such

Energy Technology Data Exchange (ETDEWEB)

Denardo, Sally J.; Denardo, Gerald L.; Balhorn, Rodney L.

2018-02-06

This invention provides novel polydentate selective high affinity ligands (SHALs) that can be used in a variety of applications in a manner analogous to the use of antibodies. SHALs typically comprise a multiplicity of ligands that each bind different region son the target molecule. The ligands are joined directly or through a linker thereby forming a polydentate moiety that typically binds the target molecule with high selectivity and avidity.

High-aluminum-affinity silica is a nanoparticle that seeds secondary aluminosilicate formation.

Directory of Open Access Journals (Sweden)

Ravin Jugdaohsingh

Full Text Available Despite the importance and abundance of aluminosilicates throughout our natural surroundings, their formation at neutral pH is, surprisingly, a matter of considerable debate. From our experiments in dilute aluminum and silica containing solutions (pH ~ 7 we previously identified a silica polymer with an extraordinarily high affinity for aluminium ions (high-aluminum-affinity silica polymer, HSP. Here, further characterization shows that HSP is a colloid of approximately 2.4 nm in diameter with a mean specific surface area of about 1,000 m(2 g(-1 and it competes effectively with transferrin for Al(III binding. Aluminum binding to HSP strongly inhibited its decomposition whilst the reaction rate constant for the formation of the β-silicomolybdic acid complex indicated a diameter between 3.6 and 4.1 nm for these aluminum-containing nanoparticles. Similarly, high resolution microscopic analysis of the air dried aluminum-containing silica colloid solution revealed 3.9 ± 1.3 nm sized crystalline Al-rich silica nanoparticles (ASP with an estimated Al:Si ratio of between 2 and 3 which is close to the range of secondary aluminosilicates such as imogolite. Thus the high-aluminum-affinity silica polymer is a nanoparticle that seeds early aluminosilicate formation through highly competitive binding of Al(III ions. In niche environments, especially in vivo, this may serve as an alternative mechanism to polyhydroxy Al(III species binding monomeric silica to form early phase, non-toxic aluminosilicates.

High-Aluminum-Affinity Silica Is a Nanoparticle That Seeds Secondary Aluminosilicate Formation

Science.gov (United States)

Jugdaohsingh, Ravin; Brown, Andy; Dietzel, Martin; Powell, Jonathan J.

2013-01-01

Despite the importance and abundance of aluminosilicates throughout our natural surroundings, their formation at neutral pH is, surprisingly, a matter of considerable debate. From our experiments in dilute aluminum and silica containing solutions (pH ~ 7) we previously identified a silica polymer with an extraordinarily high affinity for aluminium ions (high-aluminum-affinity silica polymer, HSP). Here, further characterization shows that HSP is a colloid of approximately 2.4 nm in diameter with a mean specific surface area of about 1,000 m2 g-1 and it competes effectively with transferrin for Al(III) binding. Aluminum binding to HSP strongly inhibited its decomposition whilst the reaction rate constant for the formation of the β-silicomolybdic acid complex indicated a diameter between 3.6 and 4.1 nm for these aluminum-containing nanoparticles. Similarly, high resolution microscopic analysis of the air dried aluminum-containing silica colloid solution revealed 3.9 ± 1.3 nm sized crystalline Al-rich silica nanoparticles (ASP) with an estimated Al:Si ratio of between 2 and 3 which is close to the range of secondary aluminosilicates such as imogolite. Thus the high-aluminum-affinity silica polymer is a nanoparticle that seeds early aluminosilicate formation through highly competitive binding of Al(III) ions. In niche environments, especially in vivo, this may serve as an alternative mechanism to polyhydroxy Al(III) species binding monomeric silica to form early phase, non-toxic aluminosilicates. PMID:24349573

Differences between high-affinity forskolin binding sites in dopamine-riche and other regions of rat brain

International Nuclear Information System (INIS)

Poat, J.A.; Cripps, H.E.; Iversen, L.L.

1988-01-01

Forskolin labelled with [ 3 H] bound to high- and low-affinity sites in the rat brain. The high-affinity site was discretely located, with highest densities in the striatum, nucleus accumbens, olfactory tubercule, substantia nigra, hippocampus, and the molecular layers of the cerebellum. This site did not correlate well with the distribution of adenylate cyclase. The high-affinity striatal binding site may be associated with a stimulatory guanine nucleotide-binding protein. Thus, the number of sites was increased by the addition of Mg 2+ and guanylyl imidodiphosphate. Cholera toxin stereotaxically injected into rat striatum increased the number of binding sites, and no further increase was noted following the subsequent addition of guanyl nucleotide. High-affinity forskolin binding sites in non-dopamine-rich brain areas (hippocampus and cerebullum) were modulated in a qualitatively different manner by guanyl nucleotides. In these areas the number of binding sites was significantly reduced by the addition of guanyl nucleotide. These results suggest that forskolin may have a potential role in identifying different functional/structural guanine nucleotide-binding proteins

The fourth dimension in immunological space: how the struggle for nutrients selects high-affinity lymphocytes.

Science.gov (United States)

Wensveen, Felix M; van Gisbergen, Klaas P J M; Eldering, Eric

2012-09-01

Lymphocyte activation via the antigen receptor is associated with radical shifts in metabolism and changes in requirements for nutrients and cytokines. Concomitantly, drastic changes occur in the expression of pro-and anti-apoptotic proteins that alter the sensitivity of lymphocytes to limiting concentrations of key survival factors. Antigen affinity is a primary determinant for the capacity of activated lymphocytes to access these vital resources. The shift in metabolic needs and the variable access to key survival factors is used by the immune system to eliminate activated low-affinity cells and to generate an optimal high-affinity response. In this review, we focus on the control of apoptosis regulators in activated lymphocytes by nutrients, cytokines, and costimulation. We propose that the struggle among individual clones that leads to the formation of high-affinity effector cell populations is in effect an 'invisible' fourth signal required for effective immune responses. © 2012 John Wiley & Sons A/S.

Neurochemical characterization of sea lamprey taste buds and afferent gustatory fibers: presence of serotonin, calretinin, and CGRP immunoreactivity in taste bud bi-ciliated cells of the earliest vertebrates.

Science.gov (United States)

Barreiro-Iglesias, Antón; Villar-Cerviño, Verona; Villar-Cheda, Begoña; Anadón, Ramón; Rodicio, María Celina

2008-12-01

Neuroactive substances such as serotonin and other monoamines have been suggested to be involved in the transmission of gustatory signals from taste bud cells to afferent fibers. Lampreys are the earliest vertebrates that possess taste buds, although these differ in structure from taste buds in jawed vertebrates, and their neurochemistry remains unknown. We used immunofluorescence methods with antibodies raised against serotonin, tyrosine hydroxylase (TH), gamma-aminobutyric acid (GABA), glutamate, calcitonin gene-related peptide (CGRP), neuropeptide Y (NPY), calretinin, and acetylated alpha-tubulin to characterize the neurochemistry and innervation of taste buds in the sea lamprey, Petromyzon marinus L. For localization of proliferative cells in taste buds we used bromodeoxyuridine labeling and proliferating cell nuclear antigen immunohistochemistry. Results with both markers indicate that proliferating cells are restricted to a few basal cells and that almost all cells in taste buds are nonproliferating. A large number of serotonin-, calretinin-, and CGRP-immunoreactive bi-ciliated cells were revealed in lamprey taste buds. This suggests that serotonin participates in the transmission of gustatory signals and indicates that this substance appeared early on in vertebrate evolution. The basal surface of the bi-ciliated taste bud cells was contacted by tubulin-immunoreactive fibers. Some of the fibers surrounding the taste bud were calretinin immunoreactive. Lamprey taste bud cells or afferent fibers did not exhibit TH, GABA, glutamate, or NPY immunoreactivity, which suggests that expression of these substances evolved in taste buds of some gnathostomes lines after the separation of gnathostomes and lampreys. (c) 2008 Wiley-Liss, Inc.

Detection of Waterborne Viruses Using High Affinity Molecularly Imprinted Polymers.

Science.gov (United States)

Altintas, Zeynep; Gittens, Micah; Guerreiro, Antonio; Thompson, Katy-Anne; Walker, Jimmy; Piletsky, Sergey; Tothill, Ibtisam E

2015-07-07

Molecularly imprinted polymers (MIPs) are artificial receptor ligands which can recognize and specifically bind to a target molecule. They are more resistant to chemical and biological damage and inactivation than antibodies. Therefore, target specific-MIP nanoparticles are aimed to develop and implemented to biosensors for the detection of biological toxic agents such as viruses, bacteria, and fungi toxins that cause many diseases and death due to the environmental contamination. For the first time, a molecularly imprinted polymer (MIP) targeting the bacteriophage MS2 as the template was investigated using a novel solid-phase synthesis method to obtain the artificial affinity ligand for the detection and removal of waterborne viruses through optical-based sensors. A high affinity between the artificial ligand and the target was found, and a regenerative MIP-based virus detection assay was successfully developed using a new surface plasmon resonance (SPR)-biosensor which provides an alternative technology for the specific detection and removal of waterborne viruses that lead to high disease and death rates all over the world.

Production and Identification of High Affinity Monoclonal Antibodies Against Pesticide Carbofuran

Institute of Scientific and Technical Information of China (English)

无

2007-01-01

To produce high-affinity monoclonal antibodies against pesticide carbofuran, and the develop immunochemical assays for people's health and environmental protection, the hapten 4-[[(2,3-dihydro-2,2-dimethyl-7-benzofuranyloxy) carbonyl]-amino]-butanoic acid (BFNB) of carbofuran was synthesized and Balb/c mice were immunized by the hapten-carrier (BFNB-bovine serum albumin, BFNB-BSA) conjugates. The splenocytes of immunized mice were fused with Sp2/0 cells and the cultural supernatants of hybridoma cells were screened by the indirect enzyme-linked immunoabsorbent assay (ELISA), based on BFNB-ovoalbumin conjugates (BFNB-OVA). Purified monoclonal antibody (McAb) was obtained from fluids of ascites, deposited by octanoic acid and ammonium sulfate. The affinity and the specificity of McAb were characterized by ELISA or indirect competitive ELISA. A hybridoma cell line (5D3) secreting anti-carbofuran McAb had been established. The titer of culture medium and ascites was up to 1:2.048 × 103 and 1:1.024 × 106, respectively, and the subtype of the McAb was IgG1. The affinity constant of the McAb was about 2.54 × 109 L mol-1, with an IC50 value of 1.18 ng mL-1 and a detection limit of 0.01 ng mL-1. Cross-reactivity studies showed that the McAb was quiet specific for carbofuran, as among the four analogous compounds, they were all hardly recognized (4.59 × 10-4% for 2,3-dihydro-2,2-dimethyl-7-benzofuranol and less than 3.0 × 10-4% for others). The prepared McAb had a very high affinity and specificity,and it could be used to develop ELISA for rapid determination of carbofuran.

Lamins of the sea lamprey (Petromyzon marinus) and the evolution of the vertebrate lamin protein family.

Science.gov (United States)

Schilf, Paul; Peter, Annette; Hurek, Thomas; Stick, Reimer

2014-07-01

Lamin proteins are found in all metazoans. Most non-vertebrate genomes including those of the closest relatives of vertebrates, the cephalochordates and tunicates, encode only a single lamin. In teleosts and tetrapods the number of lamin genes has quadrupled. They can be divided into four sub-types, lmnb1, lmnb2, LIII, and lmna, each characterized by particular features and functional differentiations. Little is known when during vertebrate evolution these features have emerged. Lampreys belong to the Agnatha, the sister group of the Gnathostomata. They split off first within the vertebrate lineage. Analysis of the sea lamprey (Petromyzon marinus) lamin complement presented here, identified three functional lamin genes, one encoding a lamin LIII, indicating that the characteristic gene structure of this subtype had been established prior to the agnathan/gnathostome split. Two other genes encode lamins for which orthology to gnathostome lamins cannot be designated. Search for lamin gene sequences in all vertebrate taxa for which sufficient sequence data are available reveals the evolutionary time frame in which specific features of the vertebrate lamins were established. Structural features characteristic for A-type lamins are not found in the lamprey genome. In contrast, lmna genes are present in all gnathostome lineages suggesting that this gene evolved with the emergence of the gnathostomes. The analysis of lamin gene neighborhoods reveals noticeable similarities between the different vertebrate lamin genes supporting the hypothesis that they emerged due to two rounds of whole genome duplication and makes clear that an orthologous relationship between a particular vertebrate paralog and lamins outside the vertebrate lineage cannot be established. Copyright © 2014 Elsevier GmbH. All rights reserved.

The Dopamine D2 Receptor Gene in Lamprey, Its Expression in the Striatum and Cellular Effects of D2 Receptor Activation

Science.gov (United States)

Robertson, Brita; Huerta-Ocampo, Icnelia; Ericsson, Jesper; Stephenson-Jones, Marcus; Pérez-Fernández, Juan; Bolam, J. Paul; Diaz-Heijtz, Rochellys; Grillner, Sten

2012-01-01

All basal ganglia subnuclei have recently been identified in lampreys, the phylogenetically oldest group of vertebrates. Furthermore, the interconnectivity of these nuclei is similar to mammals and tyrosine hydroxylase-positive (dopaminergic) fibers have been detected within the input layer, the striatum. Striatal processing is critically dependent on the interplay with the dopamine system, and we explore here whether D2 receptors are expressed in the lamprey striatum and their potential role. We have identified a cDNA encoding the dopamine D2 receptor from the lamprey brain and the deduced protein sequence showed close phylogenetic relationship with other vertebrate D2 receptors, and an almost 100% identity within the transmembrane domains containing the amino acids essential for dopamine binding. There was a strong and distinct expression of D2 receptor mRNA in a subpopulation of striatal neurons, and in the same region tyrosine hydroxylase-immunoreactive synaptic terminals were identified at the ultrastructural level. The synaptic incidence of tyrosine hydroxylase-immunoreactive boutons was highest in a region ventrolateral to the compact layer of striatal neurons, a region where most striatal dendrites arborise. Application of a D2 receptor agonist modulates striatal neurons by causing a reduced spike discharge and a diminished post-inhibitory rebound. We conclude that the D2 receptor gene had already evolved in the earliest group of vertebrates, cyclostomes, when they diverged from the main vertebrate line of evolution (560 mya), and that it is expressed in striatum where it exerts similar cellular effects to that in other vertebrates. These results together with our previous published data (Stephenson-Jones et al. 2011, 2012) further emphasize the high degree of conservation of the basal ganglia, also with regard to the indirect loop, and its role as a basic mechanism for action selection in all vertebrates. PMID:22563388

Computational modeling and molecular imprinting for the development of acrylic polymers with high affinity for bile salts.

Science.gov (United States)

Yañez, Fernando; Chianella, Iva; Piletsky, Sergey A; Concheiro, Angel; Alvarez-Lorenzo, Carmen

2010-02-05

This work has focused on the rational development of polymers capable of acting as traps of bile salts. Computational modeling was combined with molecular imprinting technology to obtain networks with high affinity for cholate salts in aqueous medium. The screening of a virtual library of 18 monomers, which are commonly used for imprinted networks, identified N-(3-aminopropyl)-methacrylate hydrochloride (APMA.HCl), N,N-diethylamino ethyl methacrylate (DEAEM) and ethyleneglycol methacrylate phosphate (EGMP) as suitable functional monomers with medium-to-high affinity for cholic acid. The polymers were prepared with a fix cholic acid:functional monomer mole ratio of 1:4, but with various cross-linking densities. Compared to polymers prepared without functional monomer, both imprinted and non-imprinted microparticles showed a high capability to remove sodium cholate from aqueous medium. High affinity APMA-based particles even resembled the performance of commercially available cholesterol-lowering granules. The imprinting effect was evident in most of the networks prepared, showing that computational modeling and molecular imprinting can act synergistically to improve the performance of certain polymers. Nevertheless, both the imprinted and non-imprinted networks prepared with the best monomer (APMA.HCl) identified by the modeling demonstrated such high affinity for the template that the imprinting effect was less important. The fitting of adsorption isotherms to the Freundlich model indicated that, in general, imprinting increases the population of high affinity binding sites, except when the affinity of the functional monomer for the target molecule is already very high. The cross-linking density was confirmed as a key parameter that determines the accessibility of the binding points to sodium cholate. Materials prepared with 9% mol APMA and 91% mol cross-linker showed enough affinity to achieve binding levels of up to 0.4 mmol g(-1) (i.e., 170 mg g(-1)) under flow

Multiple functions of a multi-component mating pheromone in sea lamprey Petromyzon marinus

Science.gov (United States)

Johnson, N.S.; Yun, S.-S.; Buchinger, T.J.; Li, W.

2012-01-01

The role of the C24 sulphate in the mating pheromone component, 7α,12α,24-trihydroxy-5α-cholan-3-one 24-sulphate (3kPZS), to specifically induce upstream movement in ovulated female sea lampreys Petromyzon marinus was investigated. 7α,12α-dihydroxy-5α-cholan-3-one 24-oic acid (3kACA), a structurally similar bile acid released by spermiated males, but lacking the C24 sulphate ester, was tested in bioassays at concentrations between 10−11 and 10−14 molar (M). 3kACA did not induce upstream movement in females or additional reproductive behaviours. In contrast, spermiated male washings induced upstream movement, prolonged retention on a nest and induced an array of nesting behaviours. Differential extraction and elution by solid-phase extraction resins showed that components other than 3kPZS + 3kACA are necessary to retain females on nests and induce nest cleaning behaviours. All pheromone components, including components in addition to 3kPZS + 3kACA that retain females and induce nest cleaning behaviours were released from the anterior region of the males, as had been reported for 3kPZS. It is concluded that the sea lamprey male mating pheromone has multiple functions and is composed of multiple components.

A rhodium(III) complex for high-affinity DNA base-pair mismatch recognition

Science.gov (United States)

Junicke, Henrik; Hart, Jonathan R.; Kisko, Jennifer; Glebov, Oleg; Kirsch, Ilan R.; Barton, Jacqueline K.

2003-01-01

A rhodium(III) complex, rac-[Rh(bpy)2phzi]3+ (bpy, 2,2′-bipyridine; phzi, benzo[a]phenazine-5,6-quinone diimine) has been designed as a sterically demanding intercalator targeted to destabilized mismatched sites in double-helical DNA. The complex is readily synthesized by condensation of the phenazine quinone with the corresponding diammine complex. Upon photoactivation, the complex promotes direct strand scission at single-base mismatch sites within the DNA duplex. As with the parent mismatch-specific reagent, [Rh(bpy)2(chrysi)]3+ [chrysene-5,6-quinone diimine (chrysi)], mismatch selectivity depends on the helix destabilization associated with mispairing. Unlike the parent chrysi complex, the phzi analogue binds and cleaves with high affinity and efficiency. The specific binding constants for CA, CC, and CT mismatches within a 31-mer oligonucleotide duplex are 0.3, 1, and 6 × 107 M−1, respectively; site-specific photocleavage is evident at nanomolar concentrations. Moreover, the specificity, defined as the ratio in binding affinities for mispaired vs. well paired sites, is maintained. The increase in affinity is attributed to greater stability in the mismatched site associated with stacking by the heterocyclic aromatic ligand. The high-affinity complex is also applied in the differential cleavage of DNA obtained from cell lines deficient in mismatch repair vs. those proficient in mismatch repair. Agreement is found between photocleavage by the mismatch-specific probes and deficiency in mismatch repair. This mismatch-specific targeting, therefore, offers a potential strategy for new chemotherapeutic design. PMID:12610209

High blood oxygen affinity in the air-breathing swamp eel Monopterus albus.

Science.gov (United States)

Damsgaard, Christian; Findorf, Inge; Helbo, Signe; Kocagoz, Yigit; Buchanan, Rasmus; Huong, Do Thi Thanh; Weber, Roy E; Fago, Angela; Bayley, Mark; Wang, Tobias

2014-12-01

The Asian swamp eel (Monopterus albus, Zuiew 1793) is a facultative air-breathing fish with reduced gills. Previous studies have shown that gas exchange seems to occur across the epithelium of the buccopharyngeal cavity, the esophagus and the integument, resulting in substantial diffusion limitations that must be compensated by adaptations in others steps of the O₂ transport system to secure adequate O₂ delivery to the respiring tissues. We therefore investigated O₂ binding properties of whole blood, stripped hemoglobin (Hb), two major isoHb components and the myoglobin (Mb) from M. albus. Whole blood was sampled using indwelling catheters for blood gas analysis and determination of O₂ equilibrium curves. Hb was purified to assess the effects of endogenous allosteric effectors, and Mb was isolated from heart and skeletal muscle to determine its O₂ binding properties. The blood of M. albus has a high O₂ carrying capacity [hematocrit (Hct) of 42.4±4.5%] and binds O₂ with an unusually high affinity (P₅₀=2.8±0.4mmHg at 27°C and pH7.7), correlating with insensitivity of the Hb to the anionic allosteric effectors that normally decrease Hb-O₂ affinity. In addition, Mb is present at high concentrations in both heart and muscle (5.16±0.99 and 1.08±0.19mg ∙ g wet tissue⁻¹, respectively). We suggest that the high Hct and high blood O₂ affinity serve to overcome the low diffusion capacity in the relatively inefficient respiratory surfaces, while high Hct and Mb concentration aid in increasing the O₂ flux from the blood to the muscles. Copyright © 2014 Elsevier Inc. All rights reserved.

Statistical removal of background signals from high-throughput 1H NMR line-broadening ligand-affinity screens

International Nuclear Information System (INIS)

Worley, Bradley; Sisco, Nicholas J.; Powers, Robert

2015-01-01

NMR ligand-affinity screens are vital to drug discovery, are routinely used to screen fragment-based libraries, and used to verify chemical leads from high-throughput assays and virtual screens. NMR ligand-affinity screens are also a highly informative first step towards identifying functional epitopes of unknown proteins, as well as elucidating the biochemical functions of protein–ligand interaction at their binding interfaces. While simple one-dimensional 1 H NMR experiments are capable of indicating binding through a change in ligand line shape, they are plagued by broad, ill-defined background signals from protein 1 H resonances. We present an uncomplicated method for subtraction of protein background in high-throughput ligand-based affinity screens, and show that its performance is maximized when phase-scatter correction is applied prior to subtraction

Selective induction of high-ouabain-affinity isoform of Na sup + -K sup + -ATPase by thyroid hormone

Energy Technology Data Exchange (ETDEWEB)

Haber, R.S.; Loeb, J.N. (Columbia Univ., New York, NY (USA))

1988-12-01

The administration of thyroid hormone is known to result in an induction of the Na{sup +}-K{sup +}-adenosinetriphosphatase (Na{sup +}-K{sup +}-ATPase) in rat skeletal muscle and other thyroid hormone-responsive tissues. Since the Na{sup +}-K{sup +}-ATPase in a variety of mammalian tissues has recently been reported to exist in at least two forms distinguishable by differing affinities for the inhibitory cardiac glycoside ouabain. The authors have studied the effects of 3,3{prime},5-triiodo-L-thyronine (T{sub 3}) treatment on these two forms of the enzyme in rat diaphragm. The inhibition of Na{sup +}-K{sup +}-ATPase activity in a crude membrane fraction by varying concentrations of ouabain conformed to a biphasic pattern consistent with the presence of two distinct isoforms with inhibition constants (K{sub I}s) for ouabain of {approximately}10{sup {minus}7} and 10{sup {minus}4} M, respectively. Measurement of the specific binding of ({sup 3}H)ouabain to these membranes confirmed the presence of a class of high-affinity ouabain binding sites with a dissociation constant (K{sub d}) of slightly less than 10{sup {minus}7}M, whose maximal binding capacity was increased by T{sub 3} treatment by 185%. Binding studies in unfractionated homogenates of diaphragm similarly demonstrated the presence of high-affinity sites whose maximal binding capacity was increased by T{sub 3} treatment. Quantitation of both the high- and low-ouabain-affinity forms of the Na{sup +}-K{sup +}-ATPase by ouabain-dependent phosphorylation from ({sup 32}P)orthophosphate confirmed that T{sub 3} treatment markedly increased the number of high-affinity sites while having little effect on the number of low-affinity sites. These observations provide, to our knowledge, the first demonstration that these two forms of the Na{sup +}-K{sup +}-ATPase are subject to selective hormonal induction.

Thyroid hormone deiodinase type 2 mRNA levels in sea lamprey (Petromyzon marinus) are regulated during metamorphosis and in response to a thyroid challenge.

Science.gov (United States)

Stilborn, S Salina M; Manzon, Lori A; Schauenberg, Jennifer D; Manzon, Richard G

2013-03-01

Thyroid hormones (THs) are crucial for normal vertebrate development and are the one obligate morphogen that drives amphibian metamorphosis. However, contrary to other metamorphosing vertebrates, lampreys exhibit a sharp drop in serum TH early in metamorphosis, and anti-thyroid agents such as potassium perchlorate (KClO(4)) induce metamorphosis. The type 2 deiodinase (D2) enzyme is a key regulator of TH availability during amphibian metamorphosis. We set out to determine how D2 may be involved in the regulation of lamprey metamorphosis and thyroid homeostasis. We cloned a 1.8Kb Petromyzon marinus D2 cDNA that includes the entire protein coding region and a selenocysteine (Sec) codon. Northern blotting indicated that the lamprey D2 mRNA is the longest reported to date (>9Kb). Using real-time PCR, we showed that intestinal and hepatic D2 mRNA levels were elevated prior to and during the early stages of metamorphosis and then declined dramatically to low levels that were sustained for the remainder of metamorphosis. These data are consistent with previously reported changes in serum TH levels and deiodinase activity. Treatment of larvae with either TH or KClO(4) significantly affected D2 mRNA levels in the intestine and liver. These D2 mRNA levels during metamorphosis and in response to thyroid challenges suggest that D2 may function in the regulation of TH levels during lamprey metamorphosis and the maintenance of TH homeostasis. Copyright © 2013 Elsevier Inc. All rights reserved.

Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity

Energy Technology Data Exchange (ETDEWEB)

Wang, Deng-Liang [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China); Song, Yan-Ling; Zhu, Zhi; Li, Xi-Lan; Zou, Yuan [State Key Laboratory for Physical Chemistry of Solid Surfaces, Key Laboratory for Chemical Biology of Fujian Province, Key Laboratory of Analytical Chemistry, and Department of Chemical Biology, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005 (China); Yang, Hai-Tao; Wang, Jiang-Jie [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Yao, Pei-Sen [Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China); Pan, Ru-Jun [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Yang, Chaoyong James, E-mail: cyyang@xmu.edu.cn [State Key Laboratory for Physical Chemistry of Solid Surfaces, Key Laboratory for Chemical Biology of Fujian Province, Key Laboratory of Analytical Chemistry, and Department of Chemical Biology, College of Chemistry and Chemical Engineering, Xiamen University, Xiamen 361005 (China); Kang, De-Zhi, E-mail: kdzy99988@163.com [The First Clinical Medical College of Fujian Medical University, Fuzhou (China); Department of Neurosurgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou (China)

2014-10-31

Highlights: • This is the first report of DNA aptamer against EGFR in vitro. • Aptamer can bind targets with high affinity and selectivity. • DNA aptamers are more stable, cheap and efficient than RNA aptamers. • Our selected DNA aptamer against EGFR has high affinity with K{sub d} 56 ± 7.3 nM. • Our selected DNA aptamer against EGFR has high selectivity. - Abstract: Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher’s attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with K{sub d} 56 ± 7.3 nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy.

Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity

International Nuclear Information System (INIS)

Wang, Deng-Liang; Song, Yan-Ling; Zhu, Zhi; Li, Xi-Lan; Zou, Yuan; Yang, Hai-Tao; Wang, Jiang-Jie; Yao, Pei-Sen; Pan, Ru-Jun; Yang, Chaoyong James; Kang, De-Zhi

2014-01-01

Highlights: • This is the first report of DNA aptamer against EGFR in vitro. • Aptamer can bind targets with high affinity and selectivity. • DNA aptamers are more stable, cheap and efficient than RNA aptamers. • Our selected DNA aptamer against EGFR has high affinity with K d 56 ± 7.3 nM. • Our selected DNA aptamer against EGFR has high selectivity. - Abstract: Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher’s attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with K d 56 ± 7.3 nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy

Effects of sex pheromones and sexual maturation on locomotor activity in female sea lamprey (Petromyzon marinus)

Science.gov (United States)

Walaszczyk, Erin J.; Johnson, Nicholas S.; Steibel, Juan Pedro; Li, Weiming

2013-01-01

Synchronization of male and female locomotor rhythmicity can play a vital role in ensuring reproductive success. Several physiological and environmental factors alter these locomotor rhythms. As sea lamprey, Petromyzon marinus, progress through their life cycle, their locomotor activity rhythm changes multiple times. The goal of this study was to elucidate the activity patterns of adult female sea lamprey during the sexual maturation process and discern the interactions of these patterns with exposure to male pheromones. During these stages, preovulated and ovulated adult females are exposed to sex pheromone compounds, which are released by spermiated males and attract ovulated females to the nest for spawning. The locomotor behavior of adult females was monitored in a natural stream with a passive integrated tag responder system as they matured, and they were exposed to a sex pheromone treatment (spermiated male washings) or a control (prespermiated male washings). Results showed that, dependent on the hour of day, male sex pheromone compounds reduce total activity (p reproductive synchrony of mature adults, thus increasing reproductive success in this species.

Two high-affinity ligand binding states of uterine estrogen receptor distinguished by modulation of hydrophobic environment

International Nuclear Information System (INIS)

Hutchens, T.W.; Li, C.M.; Zamah, N.M.; Besch, P.K.

1987-01-01

The steroid binding function of soluble (cytosolic) estrogen receptors from calf uteri was evaluated under conditions known to modify the extent of hydrophobic interaction with receptor-associated proteins. Receptor preparations were equilibrated into 6 M urea buffers and control buffers by chromatography through small columns of Sephadex G-25 or by dialysis at 0.6 0 C. Equilibrium dissociation constants (K/sub d/) and binding capacities (n) of experimental and control receptor preparations were determined by 13-point Scatchard analyses using concentrations of 17β-[ 3 H]estradiol from 0.05 to 10 nM. Nonspecific binding was determined at each concentration by parallel incubations with a 200-fold molar excess of the receptor-specific competitor diethylstilbestrol. The control receptor population was consistently found to be a single class of binding sites with a high affinity for estradiol which was unaffected by G-25 chromatography, by dialysis, by dilution, or by the presence of 0.4 M KCl. However, equilibration into 6 M urea induced a discrete (10-fold) reduction in receptor affinity to reveal a second, thermodynamically stable, high-affinity binding state. The presence of 0.4 M KCl did not significantly influence the discrete change in receptor affinity induced by urea. The effects of urea on both receptor affinity and binding capacity were reversible, suggesting a lack of covalent modification. These results demonstrate nonenzymatic means by which not only the binding capacity but also the affinity of receptor for estradiol can be reversibly controlled, suggesting that high concentrations of urea might be more effectively utilized during the physicochemical characterization and purification of steroid receptor proteins

Effectiveness of common fish screen materials for protecting lamprey ammocoetes—Influence of sweeping velocities and decreasing flows

Science.gov (United States)

Mesa, Matthew G.; Liedtke, Theresa L.; Weiland, Lisa K.; Christiansen, Helena E.

2017-12-14

In previous tests of the effectiveness of four common fish screen materials for excluding lamprey ammocoetes, we determined that woven wire (WW) allowed substantially more entrainment than perforated plate (PP), profile bar (PB), or Intralox (IL) material. These tests were simplistic because they used small vertically-oriented screens positioned perpendicular to the flow without a bypass or a sweeping velocity (SV). In the subsequent test discussed in this report, we exposed ammocoetes to much larger (2.5-m-wide) screen panels with flows up to 10 ft3 /s, a SV component, and a simulated bypass channel. The addition of a SV modestly improved protection of lamprey ammocoetes for all materials tested. A SV of 35 cm/s with an approach velocity (AV) of 12 cm/s, was able to provide protection for fish about 5–15 mm smaller than the protection provided by an AV of 12 cm/s without a SV component. The best-performing screen panels (PP, IL, and PB) provided nearly complete protection from entrainment for fish greater than 50-mm toal length, but the larger openings in the WW material only protected fish greater than 100-mm total length. Decreasing the AV and SV by 50 percent expanded the size range of protected lampreys by about 10–15 mm for those exposed to IL and WW screens, and it decreased the protective ability of PP screens by about 10 mm. Much of the improvement for IL and WW screens under the reduced flow conditions resulted from an increase in the number of lampreys swimming away from the screen. Fish of all sizes became impinged (that is, stuck on the screen surface for more than 1 s) on the screens, with the rate of impingement highest on PP (39– 72 percent) and lowest on WW (7–22 percent). Although impingements were common, injuries were rare, and 24-h post-test survival was greater than 99 percent. Our results refined the level of protection provided by these screen materials when both an AV and SV are present and confirmed our earlier recommendation that

Amphioxus and lamprey AP-2 genes: implications for neural crest evolution and migration patterns

Science.gov (United States)

Meulemans, Daniel; Bronner-Fraser, Marianne

2002-01-01

The neural crest is a uniquely vertebrate cell type present in the most basal vertebrates, but not in cephalochordates. We have studied differences in regulation of the neural crest marker AP-2 across two evolutionary transitions: invertebrate to vertebrate, and agnathan to gnathostome. Isolation and comparison of amphioxus, lamprey and axolotl AP-2 reveals its extensive expansion in the vertebrate dorsal neural tube and pharyngeal arches, implying co-option of AP-2 genes by neural crest cells early in vertebrate evolution. Expression in non-neural ectoderm is a conserved feature in amphioxus and vertebrates, suggesting an ancient role for AP-2 genes in this tissue. There is also common expression in subsets of ventrolateral neurons in the anterior neural tube, consistent with a primitive role in brain development. Comparison of AP-2 expression in axolotl and lamprey suggests an elaboration of cranial neural crest patterning in gnathostomes. However, migration of AP-2-expressing neural crest cells medial to the pharyngeal arch mesoderm appears to be a primitive feature retained in all vertebrates. Because AP-2 has essential roles in cranial neural crest differentiation and proliferation, the co-option of AP-2 by neural crest cells in the vertebrate lineage was a potentially crucial event in vertebrate evolution.

NK1 receptor fused to beta-arrestin displays a single-component, high-affinity molecular phenotype.

Science.gov (United States)

Martini, Lene; Hastrup, Hanne; Holst, Birgitte; Fraile-Ramos, Alberto; Marsh, Mark; Schwartz, Thue W

2002-07-01

Arrestins are cytosolic proteins that, upon stimulation of seven transmembrane (7TM) receptors, terminate signaling by binding to the receptor, displacing the G protein and targeting the receptor to clathrin-coated pits. Fusion of beta-arrestin1 to the C-terminal end of the neurokinin NK1 receptor resulted in a chimeric protein that was expressed to some extent on the cell surface but also accumulated in transferrin-labeled recycling endosomes independently of agonist stimulation. As expected, the fusion protein was almost totally silenced with respect to agonist-induced signaling through the normal Gq/G11 and Gs pathways. The NK1-beta-arrestin1 fusion construct bound nonpeptide antagonists with increased affinity but surprisingly also bound two types of agonists, substance P and neurokinin A, with high, normal affinity. In the wild-type NK1 receptor, neurokinin A (NKA) competes for binding against substance P and especially against antagonists with up to 1000-fold lower apparent affinity than determined in functional assays and in homologous binding assays. When the NK1 receptor was closely fused to G proteins, this phenomenon was eliminated among agonists, but the agonists still competed with low affinity against antagonists. In contrast, in the NK1-beta-arrestin1 fusion protein, all ligands bound with similar affinity independent of the choice of radioligand and with Hill coefficients near unity. We conclude that the NK1 receptor in complex with arrestin is in a high-affinity, stable, agonist-binding form probably best suited to structural analysis and that the receptor can display binding properties that are nearly theoretically ideal when it is forced to complex with only a single intracellular protein partner.

The high-affinity peptidoglycan binding domain of Pseudomonas phage endolysin KZ144

Energy Technology Data Exchange (ETDEWEB)

Briers, Yves [Division of Gene Technology, Department of Biosystems, Katholieke Universiteit Leuven, Kasteelpark Arenberg 21, B-3001 Leuven (Belgium); Schmelcher, Mathias; Loessner, Martin J. [Institute of Food Science and Nutrition, ETH Zuerich, Schmelzbergstrasse 7, CH-8092 Zuerich (Switzerland); Hendrix, Jelle; Engelborghs, Yves [Laboratory of Biomolecular Dynamics, Department of Chemistry, Katholieke Universiteit Leuven, Celestijnenlaan 200G, B-3001 Leuven (Belgium); Volckaert, Guido [Division of Gene Technology, Department of Biosystems, Katholieke Universiteit Leuven, Kasteelpark Arenberg 21, B-3001 Leuven (Belgium); Lavigne, Rob, E-mail: rob.lavigne@biw.kuleuven.be [Division of Gene Technology, Department of Biosystems, Katholieke Universiteit Leuven, Kasteelpark Arenberg 21, B-3001 Leuven (Belgium)

2009-05-29

The binding affinity of the N-terminal peptidoglycan binding domain of endolysin KZ144 (PBD{sub KZ}), originating from Pseudomonas aeruginosa bacteriophage {phi}KZ, has been examined using a fusion protein of PBD{sub KZ} and green fluorescent protein (PBD{sub KZ}-GFP). A fluorescence recovery after photobleaching analysis of bound PBD{sub KZ}-GFP molecules showed less than 10% fluorescence recovery in the bleached area within 15 min. Surface plasmon resonance analysis confirmed this apparent high binding affinity revealing an equilibrium affinity constant of 2.95 x 10{sup 7} M{sup -1} for the PBD{sub KZ}-peptidoglycan interaction. This unique domain, which binds to the peptidoglycan of all tested Gram-negative species, was harnessed to improve the specific activity of the peptidoglycan hydrolase domain KMV36C. The chimeric peptidoglycan hydrolase (PBD{sub KZ}-KMV36C) exhibits a threefold higher specific activity than the native catalytic domain (KMV36C). These results demonstrate that the modular assembly of functional domains is a rational approach to improve the specific activity of endolysins from phages infecting Gram-negatives.

High-affinity RNA aptamers to C-reactive protein (CRP): newly developed pre-elution methods for aptamer selection

International Nuclear Information System (INIS)

Orito, N; Umekage, S; Sakai, E; Tanaka, T; Kikuchi, Y; Sato, K; Kawauchi, S; Tanaka, H

2012-01-01

We have developed a modified SELEX (systematic evolution of ligands by exponential enrichment) method to obtain RNA aptamers with high affinity to C-reactive protein (CRP). CRP is a clinical biomarker present in plasma, the level of which increases in response to infections and noninfectious inflammation. The CRP level is also an important prognostic indicator in patients with several syndromes. At present, CRP content in blood is measured immunochemically using antibodies. To develop a more sensitive method using RNA aptamers, we have attempted to obtain high-affinity RNA aptamers to CRP. We succeeded in obtaining an RNA aptamer with high affinity to CRP using a CRP-immobilized Sepharose column and pre-elution procedure. Pre-elution is a method that removes the weak binding portion from a selected RNA population by washing for a short time with buffer containing CRP. By surface plasmon-resonance (SPR) analysis, the affinity constant of this aptamer for CRP was calculated to be K D = 2.25x10 -9 (M). The secondary structure, contact sites with CRP protein, and application of this aptamer will be described.

Isolation of Anti-Ricin Protective Antibodies Exhibiting High Affinity from Immunized Non-Human Primates

Directory of Open Access Journals (Sweden)

Tal Noy-Porat

2016-03-01

Full Text Available Ricin, derived from the castor bean plant Ricinus communis, is one of the most potent and lethal toxins known, against which there is no available antidote. To date, the use of neutralizing antibodies is the most promising post-exposure treatment for ricin intoxication. The aim of this study was to isolate high affinity anti-ricin antibodies that possess potent toxin-neutralization capabilities. Two non-human primates were immunized with either a ricin-holotoxin- or subunit-based vaccine, to ensure the elicitation of diverse high affinity antibodies. By using a comprehensive set of primers, immune scFv phage-displayed libraries were constructed and panned. A panel of 10 antibodies (five directed against the A subunit of ricin and five against the B subunit was isolated and reformatted into a full-length chimeric IgG. All of these antibodies were found to neutralize ricin in vitro, and several conferred full protection to ricin-intoxicated mice when given six hours after exposure. Six antibodies were found to possess exceptionally high affinity toward the toxin, with KD values below pM (koff < 1 × 10−7 s−1 that were well correlated with their ability to neutralize ricin. These antibodies, alone or in combination, could be used for the development of a highly-effective therapeutic preparation for post-exposure treatment of ricin intoxication.

A high affinity monoclonal antibody recognizing the light chain of human coagulating factor VII.

Science.gov (United States)

Sarial, Sheila; Asadi, Farzad; Jeddi-Tehrani, Mahmood; Hadavi, Reza; Bayat, Ali Ahmad; Mahmoudian, Jafar; Taghizadeh-Jahed, Masoud; Shokri, Fazel; Rabbani, Hodjattallah

2012-12-01

Factor VII (FVII) is a serine protease-coagulating element responsible for the initiation of an extrinsic pathway of clot formation. Here we generated and characterized a high affinity monoclonal antibody that specifically recognizes human FVII. Recombinant human FVII (rh-FVII) was used for the production of a monoclonal antibody using BALB/c mice. The specificity of the antibody was determined by Western blot using plasma samples from human, mouse, sheep, goat, bovine, rabbit, and rat. Furthermore, the antibody was used to detect transiently expressed rh-FVII in BHK21 cell line using Western blot and sandwich ELISA. A mouse IgG1 (kappa chain) monoclonal antibody clone 1F1-B11 was produced against rh-FVII. The affinity constant (K(aff)) of the antibody was calculated to be 6.4×10(10) M(-1). The antibody could specifically recognize an epitope on the light chain of hFVII, with no reactivity with factor VII from several other animals. In addition, transiently expressed rh-FVII in BHK21 cells was recognized by 1F1-B11. The high affinity as well as the specificity of 1F1-B11 for hFVII will facilitate the affinity purification of hFVII and also production of FVII deficient plasma and minimizes the risk of bovine FVII contamination when fetal bovine serum-supplemented media are used for production and subsequent purification of rh-FVII.

Affine and quasi-affine frames for rational dilations

DEFF Research Database (Denmark)

Bownik, Marcin; Lemvig, Jakob

2011-01-01

In this paper we extend the investigation of quasi-affine systems, which were originally introduced by Ron and Shen [J. Funct. Anal. 148 (1997), 408-447] for integer, expansive dilations, to the class of rational, expansive dilations. We show that an affine system is a frame if, and only if......, the corresponding family of quasi-affine systems are frames with uniform frame bounds. We also prove a similar equivalence result between pairs of dual affine frames and dual quasi-affine frames. Finally, we uncover some fundamental differences between the integer and rational settings by exhibiting an example...

Structure-based engineering to restore high affinity binding of an isoform-selective anti-TGFβ1 antibody

Science.gov (United States)

Honey, Denise M.; Best, Annie; Qiu, Huawei

2018-01-01

ABSTRACT Metelimumab (CAT192) is a human IgG4 monoclonal antibody developed as a TGFβ1-specific antagonist. It was tested in clinical trials for the treatment of scleroderma but later terminated due to lack of efficacy. Subsequent characterization of CAT192 indicated that its TGFβ1 binding affinity was reduced by ∼50-fold upon conversion from the parental single-chain variable fragment (scFv) to IgG4. We hypothesized this result was due to decreased conformational flexibility of the IgG that could be altered via engineering. Therefore, we designed insertion mutants in the elbow region and screened for binding and potency. Our results indicated that increasing the elbow region linker length in each chain successfully restored the isoform-specific and high affinity binding of CAT192 to TGFβ1. The crystal structure of the high binding affinity mutant displays large conformational rearrangements of the variable domains compared to the wild-type antigen-binding fragment (Fab) and the low binding affinity mutants. Insertion of two glycines in both the heavy and light chain elbow regions provided sufficient flexibility for the variable domains to extend further apart than the wild-type Fab, and allow the CDR3s to make additional interactions not seen in the wild-type Fab structure. These interactions coupled with the dramatic conformational changes provide a possible explanation of how the scFv and elbow-engineered Fabs bind TGFβ1 with high affinity. This study demonstrates the benefits of re-examining both structure and function when converting scFv to IgG molecules, and highlights the potential of structure-based engineering to produce fully functional antibodies. PMID:29333938

Energy-dependent dissociation of ATP from high affinity catalytic sites of beef heart mitochondrial adenosine triphosphatase

International Nuclear Information System (INIS)

Penefsky, H.S.

1985-01-01

Incubation of [gamma- 32 P]ATP with a molar excess of the membrane-bound form of mitochondrial ATPase (F1) results in binding of the bulk of the radioactive nucleotide in high affinity catalytic sites (Ka = 10(12) M-1). Subsequent initiation of respiration by addition of succinate or NADH is accompanied by a profound decrease in the affinity for ATP. About one-third of the bound radioactive ATP appears to dissociate, that is, the [gamma- 32 P]ATP becomes accessible to hexokinase. The NADH-stimulated dissociation of [gamma- 32 P]ATP is energy-dependent since the stimulation is inhibited by uncouplers of oxidative phosphorylation and is prevented by respiratory chain inhibitors. The rate of the energy-dependent dissociation of ATP that occurs in the presence of NADH, ADP, and Pi is commensurate with the measured initial rate of ATP synthesis in NADH-supported oxidative phosphorylation catalyzed by the same submitochondrial particles. Thus, the rate of dissociation of ATP from the high affinity catalytic site of submitochondrial particles meets the criterion of kinetic competency under the conditions of oxidative phosphorylation. These experiments provide evidence in support of the argument that energy conserved during the oxidation of substrates by the respiratory chain can be utilized to reduce the very tight binding of product ATP in high affinity catalytic sites and to promote dissociation of the nucleotide

Traumatic injury induces changes in the expression of the serotonin 1A receptor in the spinal cord of lampreys.

Science.gov (United States)

Cornide-Petronio, María Eugenia; Fernández-López, Blanca; Barreiro-Iglesias, Antón; Rodicio, María Celina

2014-02-01

After spinal cord injury (SCI) in mammals, the loss of serotonin coming from the brainstem reduces the excitability of motor neurons and leads to a compensatory overexpression of serotonin receptors. Despite the key role of the serotonin receptor 1a in the control of locomotion, little attention has been put in the study of this receptor after SCI. In contrast to mammals, lampreys recover locomotion after a complete SCI, so, studies in this specie could help to understand events that lead to recovery of function. Here, we showed that in lampreys there is an acute increase in the expression of the serotonin 1A receptor transcript (5-ht1a) after SCI and a few weeks later expression levels go back to normal rostrally and caudally to the lesion. Overexpression of the 5-ht1a in rostral levels after SCI has not been reported in mammals, suggesting that this could be part of the plastic events that lead to the recovery of function in lampreys. The analysis of changes in 5-ht1a expression by zones (periventricular region and horizontally extended grey matter) showed that they followed the same pattern of changes detected in the spinal cord as a whole, with the exception of the caudal periventricular layer, where no significant differences were observed between control and experimental animals at any time post lesion. This suggests that different molecular signals act on the periventricular cells of the rostral and caudal regions to injury site and thus affecting their response to the injury in terms of expression of the 5-ht1a.

New Synthesis and Tritium Labeling of a Selective Ligand for Studying High-affinity γ-Hydroxybutyrate (GHB) Binding Sites

Science.gov (United States)

Vogensen, Stine B.; Marek, Aleš; Bay, Tina; Wellendorph, Petrine; Kehler, Jan; Bundgaard, Christoffer; Frølund, Bente; Pedersen, Martin H.F.; Clausen, Rasmus P.

2013-01-01

3-Hydroxycyclopent-1-enecarboxylic acid (HOCPCA, 1) is a potent ligand for the high-affinity GHB binding sites in the CNS. An improved synthesis of 1 together with a very efficient synthesis of [3H]-1 is described. The radiosynthesis employs in situ generated lithium trimethoxyborotritide. Screening of 1 against different CNS targets establishes a high selectivity and we demonstrate in vivo brain penetration. In vitro characterization of [3H]-1 binding shows high specificity to the high-affinity GHB binding sites. PMID:24053696

ISOLATION AND CHARACTERIZATION OF THE HIGH-AFFINITY K+-TRANSLOCATING ATPASE FROM RHODOBACTER-SPHAEROIDES

NARCIS (Netherlands)

ABEE, T; SIEBERS, A; ALTENDORF, K; KONINGS, WN

1992-01-01

Cells of the purple nonsulfur bacterium Rhodobacter sphaeroides express a high-affinity K+ uptake system when grown in media with low K+ concentrations. A vanadate-sensitive, K+-stimulated and Mg2+-stimulated ATPase was purified from membranes of these cells by solubilization with

Affinity selection-mass spectrometry and its emerging application to the high throughput screening of G protein-coupled receptors.

Science.gov (United States)

Whitehurst, Charles E; Annis, D Allen

2008-07-01

Advances in combinatorial chemistry and genomics have inspired the development of novel affinity selection-based screening techniques that rely on mass spectrometry to identify compounds that preferentially bind to a protein target. Of the many affinity selection-mass spectrometry techniques so far documented, only a few solution-based implementations that separate target-ligand complexes away from unbound ligands persist today as routine high throughput screening platforms. Because affinity selection-mass spectrometry techniques do not rely on radioactive or fluorescent reporters or enzyme activities, they can complement traditional biochemical and cell-based screening assays and enable scientists to screen targets that may not be easily amenable to other methods. In addition, by employing mass spectrometry for ligand detection, these techniques enable high throughput screening of massive library collections of pooled compound mixtures, vastly increasing the chemical space that a target can encounter during screening. Of all drug targets, G protein coupled receptors yield the highest percentage of therapeutically effective drugs. In this manuscript, we present the emerging application of affinity selection-mass spectrometry to the high throughput screening of G protein coupled receptors. We also review how affinity selection-mass spectrometry can be used as an analytical tool to guide receptor purification, and further used after screening to characterize target-ligand binding interactions, enabling the classification of orthosteric and allosteric binders.

Opioid receptor subtypes mediating the noise-induced decreases in high-affinity choline uptake in the rat brain.

Science.gov (United States)

Lai, H; Carino, M A

1992-07-01

Acute (20 min) exposure to 100-dB white noise elicits a naltrexone-sensitive decrease in sodium-dependent high-affinity choline uptake in the frontal cortex and hippocampus of the rat. In the present study, the subtypes of opioid receptors involved were investigated by pretreating rats with microinjection of specific opioid-receptor antagonists into the lateral cerebroventricle before noise exposure. We found that the noise-induced decrease in high-affinity choline uptake in the hippocampus was blocked by pretreatment with either mu-, delta-, or kappa-opioid-receptor antagonists, whereas the effect of noise on frontal cortical high-affinity choline uptake was blocked by a mu- and delta- but not by a kappa-antagonist. These data further confirm the role of endogenous opioids in mediating the effects of noise on central cholinergic activity and indicate that different neural mechanisms are involved in the effects of noise on the frontal cortical and hippocampal cholinergic systems.

Pre-spawning migration of adult Pacific lamprey, Entosphenus tridentatus, in the Willamette River, Oregon, U.S.A.

Science.gov (United States)

Clemens, Benjamin J.; Mesa, Matthew G.; Magie, Robert J.; Young, Douglas A.; Schreck, Carl B.

2012-01-01

We describe the migration distances and timing of the adult Pacific lamprey, Entosphenus tridentatus, in the Willamette River Basin (Oregon, U.S.A.). We conducted aerial surveys to track radio-tagged fish upstream of a major waterfall and hydropower complex en route to spawning areas. We detected 24 out of the 43 fish that passed the waterfall-hydropower complex. Of the detected fish, 17 were detected multiple times. Their maximum migration distance upstream in the mainstem Willamette approximated a normal distribution. The maximum distance migrated upstream did not significantly correlate with total body length (r = −0.186, P = 0.385) or date that the fish passed Willamette Falls (r = −0.118, P = 0.582). Fish migrated primarily during the spring to early summer period before stopping during the summer, when peak river temperatures (≥20°C). However, at least three fish continued to migrate upstream after September. Behavior ranged from relatively slow migration, followed by holding; to rapid migration, followed by slow migration further up in the basin. This study provides a basis for informing more detailed research on Pacific lamprey in the future.

Surgical wound healing in radio-tagged adult Pacific lamprey Entosphenus tridentatus held on different substrata

Science.gov (United States)

Mesa, M.G.; Magie, R.J.; Copeland, E.S.; Christiansen, H.E.

2011-01-01

Radio-tagged adult Pacific lamprey Entosphenus tridentatus held in a raceway with Plexiglas-lined walls and bottom healed more slowly and retained sutures longer than fish held in an all-concrete raceway or one with Plexiglas walls and a cobble-lined bottom. On all substrata, healing depended on when sutures were lost, and fish that lost their sutures in healed faster than those that kept sutures longer. Long-term suture retention led to tissue trauma, infection and poor survival.

Irreversible blockade of the high and low affinity (3H) naloxone binding sites by C-6 derivatives of morphinane-6-ones

International Nuclear Information System (INIS)

Krizsan, D.; Varga, E.; Benyhe, S.; Szucs, M.; Borsodi, A.; Hosztafi, S.

1991-01-01

C-6 derivatives-hydrazones, phenylhydrazones, dinitrophenylhydrazones, oximes and semicarbazones - of morphinane-6-ones were synthesized and their binding characteristics were studied on rat brain membranes. The dihydromorphinone and oxymorphone derivatives compete for the ( 3 H)naloxone binding sites with high affinity, while the dihydrocodeinone and oxycodone derivatives are less potent. The affinity of the new compounds is decreased for the delta sites as compared to the parent ligands. The ligands bearing bulky substituents also bind with low affinity to the kappa sites. The modification decreased the Na + -index of compounds indicating their mixed agonist-antagonist character. The dihydromorphinone derivatives are all capable to block irreversibly the high affinity binding site of ( 3 H)naloxone, whereas the dihydrocodeinone derivatives block irreversibly the low affinity site. A possible mechanism for the inhibition is suggested

A novel lentiviral scFv display library for rapid optimization and selection of high affinity antibodies.

Science.gov (United States)

Qudsia, Sehar; Merugu, Siva B; Mangukiya, Hitesh B; Hema, Negi; Wu, Zhenghua; Li, Dawei

2018-04-30

Antibody display libraries have become a popular technique to screen monoclonal antibodies for therapeutic purposes. An important aspect of display technology is to generate an optimization library by changing antibody affinity to antigen through mutagenesis and screening the high affinity antibody. In this study, we report a novel lentivirus display based optimization library antibody in which Agtuzumab scFv is displayed on cell membrane of HEK-293T cells. To generate an optimization library, hotspot mutagenesis was performed to achieve diverse antibody library. Based on sequence analysis of randomly selected clones, library size was estimated approximately to be 1.6 × 10 6 . Lentivirus display vector was used to display scFv antibody on cell surface and flow cytometery was performed to check the antibody affinity to antigen. Membrane bound scFv antibodies were then converted to secreted antibody through cre/loxP recombination. One of the mutant clones, M8 showed higher affinity to antigen in flow cytometery analysis. Further characterization of cellular and secreted scFv through western blot showed that antibody affinity was increased by three fold after mutagenesis. This study shows successful construction of a novel antibody library and suggests that hotspot mutagenesis could prove a useful and rapid optimization tool to generate similar libraries with various degree of antigen affinity. Copyright © 2018 Elsevier Inc. All rights reserved.

Nitrogen Saturation in Highly Retentive Watersheds?

Science.gov (United States)

Daley, M. L.; McDowell, W. H.

2009-12-01

Watershed managers are often concerned with minimizing the amount of N delivered to N-limited estuaries and coastal zones. A major concern is that watersheds might reach N saturation, in which N delivered to coastal zones increases due to declines in the efficiency of N retention despite constant or even reduced N inputs. We have quantified long-term changes in N inputs (atmospheric deposition, imported food and agricultural fertilizers), outputs (N concentration and export) and retention in the urbanizing Lamprey River watershed in coastal NH. Overall, the Lamprey watershed is 70% forested, receives about 13.5 kg N/ha/yr and has a high rate of annual N retention (85%). Atmospheric deposition (8.7 kg/ha/yr) is the largest N input to the watershed. Of the 2.2 kg N/ha/yr exported in the Lamprey River, dissolved organic N (DON) is the dominant form (50% of total) and it varies spatially throughout the watershed with wetland cover. Nitrate accounts for 30% of the N exported, shows a statistically significant increase from 1999 to 2009, and its spatial variability in both concentration and export is related to human population density. In sub-basins throughout the Lamprey, inorganic N retention is high (85-99%), but the efficiency of N retention declines sharply with increased human population density and associated anthropogenic N inputs. N assimilation in the vegetation, denitrification to the atmosphere and storage in the groundwater pool could all be important contributors to the current high rates of N retention. The temporal and spatial patterns that we have observed in nitrate concentration and export are driven by increases in N inputs and impervious surfaces over time, but the declining efficiency of N retention suggests that the watershed may also be reaching N saturation. The downstream receiving estuary, Great Bay, already suffers from low dissolved oxygen levels and eelgrass loss in part due to N loading from the Lamprey watershed. Targeting and reducing

The sea lamprey meiotic map improves resolution of ancient vertebrate genome duplications.

Science.gov (United States)

Smith, Jeramiah J; Keinath, Melissa C

2015-08-01

It is generally accepted that many genes present in vertebrate genomes owe their origin to two whole-genome duplications that occurred deep in the ancestry of the vertebrate lineage. However, details regarding the timing and outcome of these duplications are not well resolved. We present high-density meiotic and comparative genomic maps for the sea lamprey (Petromyzon marinus), a representative of an ancient lineage that diverged from all other vertebrates ∼550 million years ago. Linkage analyses yielded a total of 95 linkage groups, similar to the estimated number of germline chromosomes (1n ∼ 99), spanning a total of 5570.25 cM. Comparative mapping data yield strong support for the hypothesis that a single whole-genome duplication occurred in the basal vertebrate lineage, but do not strongly support a hypothetical second event. Rather, these comparative maps reveal several evolutionarily independent segmental duplications occurring over the last 600+ million years of chordate evolution. This refined history of vertebrate genome duplication should permit more precise investigations of vertebrate evolution. © 2015 Smith and Keinath; Published by Cold Spring Harbor Laboratory Press.

Hemoglobin affinity in Andean rodents

Directory of Open Access Journals (Sweden)

HRVOJ OSTOJIC

2002-01-01

Full Text Available Blood hemoglobin oxygen affinity (P50 was measured in three Andean species and in the laboratory rat (control, all raised near sea level. Chinchilla lanigera (Molina, 1792 has an altitudinal habitat range from low Andean slopes up to 3000 m., while Chinchilla brevicaudata (Waterhouse, 1848 has an altitudinal range from 3000 to 5000 m. The laboratory type guinea pig, wild type guinea pig (Cavia porcellus, (Waterhouse, 1748, and laboratory rat (Rattus norvegicus were also raised at sea level. The Andean species had high hemoglobin oxygen affinities (low P50 compared with the rat. Chinchilla brevicaudata had a higher affinity than Chinchilla lanigera. The wild type guinea pig had a higher affinity than the laboratory type. As has been shown in other species, this is another example of an inverse correlation between the altitude level and the P50 values. This is the first hemoglobin oxygen affinity study in Chinchilla brevicaudata.

High-affinity receptors for bombesin-like peptides in normal guinea pig lung membranes

International Nuclear Information System (INIS)

Lach, E.; Trifilieff, A.; Landry, Y.; Gies, J.P.

1991-01-01

The binding of the radiolabeled bombesin analogue [ 125 I-Tyr 4 ]bombesin to guinea-pig lung membranes was investigated. Binding of [ 125 I-Tyr 4 ]bombesin was specific, saturable, reversible and linearly related to the protein concentration. Scatchard analysis of equilibrium binding data at 25C indicated the presence of a single class of non-interacting binding sites for bombesin (B max = 7.7 fmol/mg protein). The value of the equilibrium dissociation constant (K D = 90 pM) agrees with a high-affinity binding site. Bombesin and structurally related peptides such as [ 125 I-Tyr 4 ]bombesin, neuromedin B and neuromedin C inhibited the binding of [ 125 I-Tyr 4 ]bombesin in an order of potencies as follows: [ 125 I-Tyr 4 ]bombesin > bombesin ≥ neuromedin C much-gt neuromedin B. These results indicate that guinea-pig lung membranes possess a single class of bombesin receptors with a high affinity for bombesin and a lower one for neuromedin B

High-throughput bioscreening system utilizing high-performance affinity magnetic carriers exhibiting minimal non-specific protein binding

International Nuclear Information System (INIS)

Hanyu, Naohiro; Nishio, Kosuke; Hatakeyama, Mamoru; Yasuno, Hiroshi; Tanaka, Toshiyuki; Tada, Masaru; Nakagawa, Takashi; Sandhu, Adarsh; Abe, Masanori; Handa, Hiroshi

2009-01-01

For affinity purification of drug target protein we have developed magnetic carriers, narrow in size distribution (184±9 nm), which exhibit minimal non-specific binding of unwanted proteins. The carriers were highly dispersed in aqueous solutions and highly resistant to organic solvents, which enabled immobilization of various hydrophobic chemicals as probes on the carrier surfaces. Utilizing the carriers we have automated the process of separation and purification of the target proteins that had been done by manual operation previously.

A mix-and-read drop-based in vitro two-hybrid method for screening high-affinity peptide binders

Science.gov (United States)

Cui, Naiwen; Zhang, Huidan; Schneider, Nils; Tao, Ye; Asahara, Haruichi; Sun, Zhiyi; Cai, Yamei; Koehler, Stephan A.; de Greef, Tom F. A.; Abbaspourrad, Alireza; Weitz, David A.; Chong, Shaorong

2016-01-01

Drop-based microfluidics have recently become a novel tool by providing a stable linkage between phenotype and genotype for high throughput screening. However, use of drop-based microfluidics for screening high-affinity peptide binders has not been demonstrated due to the lack of a sensitive functional assay that can detect single DNA molecules in drops. To address this sensitivity issue, we introduced in vitro two-hybrid system (IVT2H) into microfluidic drops and developed a streamlined mix-and-read drop-IVT2H method to screen a random DNA library. Drop-IVT2H was based on the correlation between the binding affinity of two interacting protein domains and transcriptional activation of a fluorescent reporter. A DNA library encoding potential peptide binders was encapsulated with IVT2H such that single DNA molecules were distributed in individual drops. We validated drop-IVT2H by screening a three-random-residue library derived from a high-affinity MDM2 inhibitor PMI. The current drop-IVT2H platform is ideally suited for affinity screening of small-to-medium-sized libraries (103–106). It can obtain hits within a single day while consuming minimal amounts of reagents. Drop-IVT2H simplifies and accelerates the drop-based microfluidics workflow for screening random DNA libraries, and represents a novel alternative method for protein engineering and in vitro directed protein evolution. PMID:26940078

Synthesis of site-heterologous haptens for high-affinity anti-pyraclostrobin antibody generation.

Science.gov (United States)

Mercader, Josep V; Agulló, Consuelo; Abad-Somovilla, Antonio; Abad-Fuentes, Antonio

2011-03-07

The design and synthesis of functional chemical derivatives of small organic molecules is usually a key step for the intricate production of a variety of bioconjugates. In this respect, the derivatization site at which the spacer arm is introduced in immunizing conjugates constitutes a highly critical parameter for the generation of high-affinity and selective antibodies. However, due to the usual complexity of the required synthetic procedures, the appropriate comparison of alternative tethering positions has often been neglected. In the present study, meticulous strategies were followed to prepare synthetic derivatives of pyraclostrobin with the same linkers located at diverse rationally-chosen sites. Activity appraisal of antibodies and bioconjugates was carried out by bidimensional competitive direct and indirect immunoassays, and a superior performance of two of the three synthesized haptens was found. Finally, a detailed analysis of the conformations of the target molecule and the synthesized haptens in aqueous solution was done using computer assisted molecular modeling techniques. This study suggested that the lower titers and affinities of one set of antibodies are most probably due to conformational effects of the spacer arm in the immunizing bioconjugate.

Affinity imaging mass spectrometry (AIMS): high-throughput screening for specific small molecule interactions with frozen tissue sections.

Science.gov (United States)

Yoshimi, T; Kawabata, S; Taira, S; Okuno, A; Mikawa, R; Murayama, S; Tanaka, K; Takikawa, O

2015-11-07

A novel screening system, using affinity imaging mass spectrometry (AIMS), has been developed to identify protein aggregates or organ structures in unfixed human tissue. Frozen tissue sections are positioned on small (millimetre-scale) stainless steel chips and incubated with an extensive library of small molecules. Candidate molecules showing specific affinity for the tissue section are identified by imaging mass spectrometry (IMS). As an example application, we screened over a thousand compounds against Alzheimer's disease (AD) brain tissue and identified several compounds with high affinity for AD brain sections containing tau deposits compared to age-matched controls. It should also be possible to use AIMS to isolate chemical compounds with affinity for tissue structures or components that have been extensively modified by events such as oxidation, phosphorylation, acetylation, aggregation, racemization or truncation, for example, due to aging. It may also be applicable to biomarker screening programs.

Molecular electron affinities

International Nuclear Information System (INIS)

Fukuda, E.K.

1983-01-01

Molecular electron affinities have historically been difficult quantities to measure accurately. These difficulties arise from differences in structure between the ion and neutral as well as the existence of excited negative ion states. To circumvent these problems, relative electron affinities were determined in this dissertation by studying equilibrium electron transfer reactions using a pulsed ion cyclotron resonance (ICR) spectrometer. Direct measurement of ion and neutral concentrations for reactions of the general type, A - + B = B - + A, allow calculation of the equilibrium constant and, therefore, the free energy change. The free energy difference is related to the difference in electron affinities between A and B. A relative electron affinity scale covering a range of about 45 kcal/mol was constructed with various substituted p-benzoquinones, nitrobenzenes, anhydrides, and benzophenones. To assign absolute electron affinities, various species with accurately known electron affinities are tied to the scale via ion-cyclotron double resonance bracketing techniques. After the relative scale is anchored to these species with well-known electron affinities, the scale is then used as a check on other electron affinity values as well as generating new electron affinity values. Many discrepancies were found between the electron affinities measured using the ICR technique and previous literature determinations

High affinity, ligand specific uptake of complexed copper-67 by brain tissue incubated in vitro

International Nuclear Information System (INIS)

Barnea, A.; Hartter, D.E.

1987-01-01

Copper is an essential metal that is highly concentrated in the brain. The blood, the sole source of tissue Cu, contains 16-20 μM Cu, of which >95% is complexed to proteins and 2 was 10 times greater than that of CuAlbumin or Cu(II). Within the range of 0.2-150μM Cu, multiple uptake sites for CuHis were apparent. Increasing the molar ratio of His:Cu had a differential effect on Cu uptake: enhancing uptake at [Cu] 1 μM. Thus, using a His:Cu ratio of 1000, they observed a high affinity process exhibiting saturating and half saturating values of 5 μM and 1.5 μM Cu, respectively; using a His:Cu ratio of 2, they observed a low affinity process exhibiting saturating and half-saturating values of 100 μM and 40 μM Cu, respectively. Both processes required thermic but not metabolic energy, suggestive of facilitated diffusion. Considering the blood brain barrier for proteins, CuHis appears to be the major substrate for Cu uptake by neuronal tissue. They demonstrate the existence of a ligand specific, high affinity (apparent Km about 1.5 μM Cu) uptake process for CuHis in the brain, operative at the physiological concentration range of CuHis and histidine

Annual sex steroid and other physiological profiles of Pacific lampreys (Entosphenus tridentatus)

Science.gov (United States)

Mesa, Matthew G.; Bayer, Jennifer M.; Bryan, Mara B.; Sower, Stacia A.

2010-01-01

We documented changes in plasma levels of estradiol 17-β (E2), progesterone (P), 15α-hydroxytestosterone (15α-T), thyroxine (T4), triiodothyronine (T3), protein, triglycerides (TGs), and glucose in adult Pacific lampreys (Entosphenus tridentatus) held in the laboratory in two different years. Levels of E2 in both sexes ranged from 0.5 to 2 ng/mL from September to March, peaked in late April (2–4 ng/mL), and decreased in May, with levels higher in males than in females. Levels of P were low from September through April, but then increased substantially during May (2–4 ng/mL), with levels again highest in males. Levels of 15α-T in males were around 0.75 ng/mL through the winter before exceeding 1 ng/mL in April and decreasing thereafter, whereas females showed a gradual increase from 0.25 ng/mL in November to 0.5 ng/mL in April before decreasing. Thyroxine concentrations differed between fish in each year, with most having levels ranging from 0.75 to 2.5 ng/mL in the fall and winter, and only fish in 2003 showing distinct peaks (3–4 ng/mL) in early April or May. Plasma T3 was undetectable from November through mid-March before surging dramatically in April (ca. 150 ng/mL) and decreasing thereafter. Levels of protein, TGs, and glucose decreased or were stable during the fall and winter with TGs and glucose surging in late April to early May for some fish. Our study is the first to document long-term physiological changes in Pacific lampreys during overwintering and sexual maturation and increases our understanding of the life history of this unique fish.

Amyloid-beta binds catalase with high affinity and inhibits hydrogen peroxide breakdown.

OpenAIRE

Milton, N G

1999-01-01

Amyloid-beta (Abeta) specifically bound purified catalase with high affinity and inhibited catalase breakdown of H(2)O(2). The Abeta-induced catalase inhibition involved formation of the inactive catalase Compound II and was reversible. CatalaseAbeta interactions provide rapid functional assays for the cytotoxic domain of Abeta and suggest a mechanism for some of the observed actions of Abeta plus catalase in vitro.

Mobile Technology Affinity in Renal Transplant Recipients.

Science.gov (United States)

Reber, S; Scheel, J; Stoessel, L; Schieber, K; Jank, S; Lüker, C; Vitinius, F; Grundmann, F; Eckardt, K-U; Prokosch, H-U; Erim, Y

Medication nonadherence is a common problem in renal transplant recipients (RTRs). Mobile health approaches to improve medication adherence are a current trend, and several medication adherence apps are available. However, it is unknown whether RTRs use these technologies and to what extent. In the present study, the mobile technology affinity of RTRs was analyzed. We hypothesized significant age differences in mobile technology affinity and that mobile technology affinity is associated with better cognitive functioning as well as higher educational level. A total of 109 RTRs (63% male) participated in the cross-sectional study, with an overall mean age of 51.8 ± 14.2 years. The study included the Technology Experience Questionnaire (TEQ) for the assessment of mobile technology affinity, a cognitive test battery, and sociodemographic data. Overall, 57.4% of the patients used a smartphone or tablet and almost 45% used apps. The TEQ sum score was 20.9 in a possible range from 6 (no affinity to technology) to 30 (very high affinity). Younger patients had significantly higher scores in mobile technology affinity. The only significant gender difference was found in having fun with using electronic devices: Men enjoyed technology more than women did. Mobile technology affinity was positively associated with cognitive functioning and educational level. Young adult patients might profit most from mobile health approaches. Furthermore, high educational level and normal cognitive functioning promote mobile technology affinity. This should be kept in mind when designing mobile technology health (mHealth) interventions for RTRs. For beneficial mHealth interventions, further research on potential barriers and desired technologic features is necessary to adapt apps to patients' needs. Copyright © 2017 Elsevier Inc. All rights reserved.

Identification of high-affinity calmodulin-binding proteins in rat liver

International Nuclear Information System (INIS)

Hanley, R.M.; Dedman, J.R.; Shenolikar, S.

1987-01-01

The Ca 2+ -dependent binding of [ 125 I] calmodulin (CaM) to hepatic proteins separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was utilized to identify CaM binding or acceptor proteins or CAPs. Two proteins of apparent molecular weight of 60,000 (CAP-60) and 45,000 (CAP-45) comprised > 80% of the Ca 2+ -dependent CaM binding in rat liver cytosol. CAP-60 and CAP-45 were partially purified by a variety of chromatographic steps, including affinity chromatography on CaM Sepharose. CAP-60 possessed a native molecular size of 400,000, indicating it to be the CaM-binding subunit of a larger oligomeric complex. In contrast, CAP-45 was monomeric as judged by gel filtration. Neither CAP-60 nor CAP-45 possessed chromatographic properties consistent with known CaM-dependent enzymes reported in the literature. Two-dimensional peptide mapping provided convincing evidence that CAP-60 and CAP-45 were unrelated to other well-characterized CAPs, namely Ca 2+ (CaM)-dependent protein kinase II, calcineurin, or the CaM-dependent cyclic nucleotide phosphodiesterase. The relative abundance and high affinity for CaM could suggest that these novel target proteins, CAP-60 and CAP-45, represent a dominant pathway for CaM action in the mammalian liver

Antibody Binding Selectivity: Alternative Sets of Antigen Residues Entail High-Affinity Recognition.

Directory of Open Access Journals (Sweden)

Yves Nominé

Full Text Available Understanding the relationship between protein sequence and molecular recognition selectivity remains a major challenge. The antibody fragment scFv1F4 recognizes with sub nM affinity a decapeptide (sequence 6TAMFQDPQER15 derived from the N-terminal end of human papilloma virus E6 oncoprotein. Using this decapeptide as antigen, we had previously shown that only the wild type amino-acid or conservative replacements were allowed at positions 9 to 12 and 15 of the peptide, indicating a strong binding selectivity. Nevertheless phenylalanine (F was equally well tolerated as the wild type glutamine (Q at position 13, while all other amino acids led to weaker scFv binding. The interfaces of complexes involving either Q or F are expected to diverge, due to the different physico-chemistry of these residues. This would imply that high-affinity binding can be achieved through distinct interfacial geometries. In order to investigate this point, we disrupted the scFv-peptide interface by modifying one or several peptide positions. We then analyzed the effect on binding of amino acid changes at the remaining positions, an altered susceptibility being indicative of an altered role in complex formation. The 23 starting variants analyzed contained replacements whose effects on scFv1F4 binding ranged from minor to drastic. A permutation analysis (effect of replacing each peptide position by all other amino acids except cysteine was carried out on the 23 variants using the PEPperCHIP® Platform technology. A comparison of their permutation patterns with that of the wild type peptide indicated that starting replacements at position 11, 12 or 13 modified the tolerance to amino-acid changes at the other two positions. The interdependence between the three positions was confirmed by SPR (Biacore® technology. Our data demonstrate that binding selectivity does not preclude the existence of alternative high-affinity recognition modes.

Bayesian phylogeny analysis of vertebrate serpins illustrates evolutionary conservation of the intron and indels based six groups classification system from lampreys for ∼500 MY

Directory of Open Access Journals (Sweden)

Abhishek Kumar

2015-06-01

Full Text Available The serpin superfamily is characterized by proteins that fold into a conserved tertiary structure and exploits a sophisticated and irreversible suicide-mechanism of inhibition. Vertebrate serpins are classified into six groups (V1–V6, based on three independent biological features—genomic organization, diagnostic amino acid sites and rare indels. However, this classification system was based on the limited number of mammalian genomes available. In this study, several non-mammalian genomes are used to validate this classification system using the powerful Bayesian phylogenetic method. This method supports the intron and indel based vertebrate classification and proves that serpins have been maintained from lampreys to humans for about 500 MY. Lampreys have fewer than 10 serpins, which expand into 36 serpins in humans. The two expanding groups V1 and V2 have SERPINB1/SERPINB6 and SERPINA8/SERPIND1 as the ancestral serpins, respectively. Large clusters of serpins are formed by local duplications of these serpins in tetrapod genomes. Interestingly, the ancestral HCII/SERPIND1 locus (nested within PIK4CA possesses group V4 serpin (A2APL1, homolog of α2-AP/SERPINF2 of lampreys; hence, pointing to the fact that group V4 might have originated from group V2. Additionally in this study, details of the phylogenetic history and genomic characteristics of vertebrate serpins are revisited.

Identification and properties of very high affinity brain membrane-binding sites for a neurotoxic phospholipase from the taipan venom

International Nuclear Information System (INIS)

Lambeau, G.; Barhanin, J.; Schweitz, H.; Qar, J.; Lazdunski, M.

1989-01-01

Four new monochain phospholipases were purified from the Oxyuranus scutellatus (taipan) venom. Three of them were highly toxic when injected into mice brain. One of these neurotoxic phospholipases, OS2, was iodinated and used in binding experiments to demonstrate the presence of two families of specific binding sites in rat brain synaptic membranes. The affinities were exceptionally high, Kd1 = 1.5 +/- 0.5 pM and Kd2 = 45 +/- 10 pM, and the maximal binding capacities were Bmax 1 = 1 +/- 0.4 and Bmax 2 = 3 +/- 0.5 pmol/mg of protein. Both binding sites were sensitive to proteolysis and demonstrated to be located on proteins of Mr 85,000-88,000 and 36,000-51,000 by cross-linking and photoaffinity labeling techniques. The binding of 125 I-OS2 to synaptic membranes was dependent on Ca2+ ions and enhanced by Zn2+ ions which inhibit phospholipase activity. Competition experiments have shown that, except for beta-bungarotoxin, a number of known toxic snake or bee phospholipases have very high affinities for the newly identified binding sites. A good correlation (r = 0.80) was observed between toxicity and affinity but not between phospholipase activity and affinity

The utility of affine variables and affine coherent states

International Nuclear Information System (INIS)

Klauder, John R

2012-01-01

Affine coherent states are generated by affine kinematical variables much like canonical coherent states are generated by canonical kinematical variables. Although all classical and quantum formalisms normally entail canonical variables, it is shown that affine variables can serve equally well for many classical and quantum studies. This general purpose analysis provides tools to discuss two major applications: (1) the completely successful quantization of a nonrenormalizable scalar quantum field theory by affine techniques, in complete contrast to canonical techniques which only offer triviality; and (2) a formulation of the kinematical portion of quantum gravity that favors affine kinematical variables over canonical kinematical variables, and which generates a framework in which a favorable analysis of the constrained dynamical issues can take place. All this is possible because of the close connection between the affine and the canonical stories, while the few distinctions can be used to advantage when appropriate. This article is part of a special issue of Journal of Physics A: Mathematical and Theoretical devoted to ‘Coherent states: mathematical and physical aspects’. (review)

Selection of imprinted nanoparticles by affinity chromatography.

Science.gov (United States)

Guerreiro, António R; Chianella, Iva; Piletska, Elena; Whitcombe, Michael J; Piletsky, Sergey A

2009-04-15

Soluble molecularly imprinted nanoparticles were synthesised via iniferter initiated polymerisation and separated by size via gel permeation chromatography. Subsequent fractionation of these particles by affinity chromatography allowed the separation of high affinity fractions from the mixture of nanoparticles. Fractions selected this way possess affinity similar to that of natural antibodies (K(d) 6.6x10(-8)) M and were also able to discriminate between related functional analogues of the template.

Structural insights into a high affinity nanobody:antigen complex by homology modelling

DEFF Research Database (Denmark)

Skottrup, Peter Durand

2017-01-01

Porphyromonas gingivalis is a major periodontitis-causing pathogens. P. gingivalis secrete a cysteine protease termed RgpB, which is specific for Arg-Xaa bonds in substrates. Recently, a nanobody-based assay was used to demonstrate that RgpB could represent a novel diagnostic target, thereby...... simplifying. P. gingivalis detection. The nanobody, VHH7, had a high binding affinity and was specific for RgpB, when tested towards the highly identical RgpA. In this study a homology model of VHH7 was build. The complementarity determining regions (CDR) comprising the paratope residues responsible for Rgp...

Response of larval sea lampreys (Petromyzon marinus) to pulsed DC electrical stimuli in laboratory experiments

Science.gov (United States)

Bowen, Anjanette K.; Weisser, John W.; Bergstedt, Roger A.; Famoye, Felix

2003-01-01

Four electrical factors that are used in pulsed DC electrofishing for larval sea lampreys (Petromyzon marinus) were evaluated in two laboratory studies to determine the optimal values to induce larval emergence over a range of water temperatures and conductivities. Burrowed larvae were exposed to combinations of pulsed DC electrical factors including five pulse frequencies, three pulse patterns, and two levels of duty cycle over a range of seven voltage gradients in two separate studies conducted at water temperatures of 10, 15, and 20°C and water conductivities of 25, 200, and 900 μS/cm. A four-way analysis of variance was used to determine significant (α = 0.05) influences of each electrical factor on larval emergence. Multiple comparison tests with Bonferroni adjustments were used to determine which values of each factor resulted in significantly higher emergence at each temperature and conductivity. Voltage gradient and pulse frequency significantly affected emergence according to the ANOVA model at each temperature and conductivity tested. Duty cycle and pulse pattern generally did not significantly influence the model. Findings suggest that a setting of 2.0 V/cm, 3 pulses/sec, 10% duty, and 2:2 pulse pattern seems the most promising in waters of medium conductivity and across a variety of temperatures. This information provides a basis for understanding larval response to pulsed DC electrofishing gear factors and identifies electrofisher settings that show promise to increase the efficiency of the gear during assessments for burrowed sea lamprey larvae.

Alternative affinity tools: more attractive than antibodies?

NARCIS (Netherlands)

Ruigrok, V.J.B.; Levisson, M.; Eppink, M.H.M.; Smidt, H.; Oost, van der J.

2011-01-01

Antibodies are the most successful affinity tools used today, in both fundamental and applied research (diagnostics, purification and therapeutics). Nonetheless, antibodies do have their limitations, including high production costs and low stability. Alternative affinity tools based on nucleic acids

Trace element affinities in two high-Ge coals from China

Energy Technology Data Exchange (ETDEWEB)

Jing Li; Xinguo Zhuang; Xavier Querol [China University of Geosciences, Wuhan (China). Faculty of Earth Resources

2011-01-15

The Lincang (Yunnan Province, Southwest China) and Wulantuga (Inner Mongolia, Northeast China) coal deposits are known because of the high-Ge content. These coals have also a high concentration of a number of other elements. To determine the mode of occurrence of the enriched elements in both coals, six density fractions from {lt} 1.43 to {gt} 2.8 g/cm{sup 3} were obtained from two representative samples using heavy-liquids. A number of peculiar geochemical patterns characterize these high-Ge coals. Thus, the results of the chemical analysis of these density fractions showed that both coals (very distant and of a different geological age) are highly enriched (compared with the usual worldwide coal concentration ranges) in Ge, As, Sb, W, Be, and Tl. This may be due to similar geochemistry of hydrothermal fluids influencing the Earth Crust in these regions of China. Moreover, Wulantuga coal (Early Cretaceous subbituminous coal) is also enriched in Ca, Mg, and Na, and Lincang coal (Neogene subbituminous coal) in K, Rb, Nb, Mo, Sn, Cs, and U. A group of elements consisting of Ge, W, B, Nb, and Sb mostly occur with an organic affinity in both coals. Additionally, Be, U, and Mo (and partially Mn and Zn) in Lincang, and Na and Mg in Wulantuga occur also with a major organic affinity. Both coals have sulfide-arsenide mineral assemblages (Fe, S, As, Sn, and Pb, and in addition to Tl, Ta, and Cs in the Lincang coal). The occurrence of Al, P, Li, Sc, Ti, V, Cr, and Zr in both coals, and Ba in Lincang, are associated with the mineral assemblage of silico-aluminates and minor heavy minerals. Furthermore, P, Na, Li, Sc, Ti, Ga, Rb, Zr, Cr, Ba, Th, and LREE (La, Ce, Pr, Nd, and Gd) in Lincang are associated with mineral assemblages of phosphates and minor heavy minerals. The two later mineral assemblages are derived from the occurrence of detrital minerals. 34 refs., 7 figs., 3 tabs.

A study of the uptake of chloroquine in malaria-infected erythrocytes. High and low affinity uptake and the influence of glucose and its analogues.

Science.gov (United States)

Diribe, C O; Warhurst, D C

1985-09-01

A study of concentration- and substrate-dependence of chloroquine uptake has been carried out on mouse erythrocytes infected with the chloroquine-sensitive NK65 and the chloroquine-resistant RC strains of Plasmodium berghei. The presence of drug binding sites of high and low affinity in such strains of P. berghei was confirmed. High affinity uptake sites in cells parasitized with chloroquine-sensitive and chloroquine-resistant parasites have similar characteristics, but in the sensitive strain the major component of chloroquine-uptake is at high affinity and dependent on the availability of ATP whilst in the resistant strain the major component of uptake is at low affinity and independent of energy. An absolute increase in the quantity of the low affinity site in erythrocytes parasitized with chloroquine-resistant P. berghei was noted, which may be related to an increase in quantity of parasite membrane.

Characterization of a high affinity cocaine binding site in rat brain

International Nuclear Information System (INIS)

Calligaro, D.; Eldefrawi, M.

1986-01-01

Binding of [ 3 H]cocaine to synaptic membranes from whole rat brain was reversible and saturable. Nonlinear regression analysis of binding isotherms indicated two binding affinities: one with k/sub d/ = 16 nM, B/sub max/ = 0.65 pmoles/mg protein and the other with K/sub d/ = 660 nM, B/sub max/ = 5.1 pmoles/mg protein. The high-affinity binding of [ 3 H]cocaine was sensitive to the actions of trypsin and chymotrypsin but not carboxypeptidase, and was eliminated by exposure of the membranes to 95 0 C for 5 min. Specific binding at 2 nM was higher at pH 8.8 than at pH 7.0. Binding of [ 3 H]cocaine (15 nM) was inhibited by increasing concentrations of Na + ions. Several cocaine analogues, neurotransmitter uptake inhibitors and local anesthetics displaced specific [ 3 H]cocaine binding at 2 nM with various potencies. The cocaine analogue (-)-norcocaine was the most potent (IC 50 = 10 nM), while the local anesthetic tetracaine was the least potent in inhibiting [ 3 H]cocaine binding. Several biogenic amine uptake inhibitors, including tricyclic antidepressants and phencyclidine, had IC 50 values below μM concentrations

Report: Affinity Chromatography.

Science.gov (United States)

Walters, Rodney R.

1985-01-01

Supports, affinity ligands, immobilization, elution methods, and a number of applications are among the topics considered in this discussion of affinity chromatography. An outline of the basic principles of affinity chromatography is included. (JN)

Discovery of PF-06928215 as a high affinity inhibitor of cGAS enabled by a novel fluorescence polarization assay.

Science.gov (United States)

Hall, Justin; Brault, Amy; Vincent, Fabien; Weng, Shawn; Wang, Hong; Dumlao, Darren; Aulabaugh, Ann; Aivazian, Dikran; Castro, Dana; Chen, Ming; Culp, Jeffrey; Dower, Ken; Gardner, Joseph; Hawrylik, Steven; Golenbock, Douglas; Hepworth, David; Horn, Mark; Jones, Lyn; Jones, Peter; Latz, Eicke; Li, Jing; Lin, Lih-Ling; Lin, Wen; Lin, David; Lovering, Frank; Niljanskul, Nootaree; Nistler, Ryan; Pierce, Betsy; Plotnikova, Olga; Schmitt, Daniel; Shanker, Suman; Smith, James; Snyder, William; Subashi, Timothy; Trujillo, John; Tyminski, Edyta; Wang, Guoxing; Wong, Jimson; Lefker, Bruce; Dakin, Leslie; Leach, Karen

2017-01-01

Cyclic GMP-AMP synthase (cGAS) initiates the innate immune system in response to cytosolic dsDNA. After binding and activation from dsDNA, cGAS uses ATP and GTP to synthesize 2', 3' -cGAMP (cGAMP), a cyclic dinucleotide second messenger with mixed 2'-5' and 3'-5' phosphodiester bonds. Inappropriate stimulation of cGAS has been implicated in autoimmune disease such as systemic lupus erythematosus, thus inhibition of cGAS may be of therapeutic benefit in some diseases; however, the size and polarity of the cGAS active site makes it a challenging target for the development of conventional substrate-competitive inhibitors. We report here the development of a high affinity (KD = 200 nM) inhibitor from a low affinity fragment hit with supporting biochemical and structural data showing these molecules bind to the cGAS active site. We also report a new high throughput cGAS fluorescence polarization (FP)-based assay to enable the rapid identification and optimization of cGAS inhibitors. This FP assay uses Cy5-labelled cGAMP in combination with a novel high affinity monoclonal antibody that specifically recognizes cGAMP with no cross reactivity to cAMP, cGMP, ATP, or GTP. Given its role in the innate immune response, cGAS is a promising therapeutic target for autoinflammatory disease. Our results demonstrate its druggability, provide a high affinity tool compound, and establish a high throughput assay for the identification of next generation cGAS inhibitors.

High-Affinity Methanotrophy Informed by Genome-Wide Analysis of Upland Soil Cluster Alpha (USCα) from Axel Heiberg Island, Canadian High Arctic

Science.gov (United States)

Rusley, C.; Onstott, T. C.; Lau, M.

2017-12-01

Methane (CH4) is a potent greenhouse gas whose proper budgeting is vital to climate predictions. Recent studies have identified upland Arctic mineral cryosols as consistent CH4 sinks, drawing CH4 from both the atmosphere and underlying anaerobic soil layers. Global atmospheric CH4 uptake is proposed to be mediated by high-affinity methanotrophs based on the detection of the marker gene pmoA (particulate methane monooxygenase beta subunit). However, a lack of pure cultures and scarcity of genomic information have hindered our understanding of their metabolic capabilities and versatility. Together with the missing genetic linkage between its pmoA and 16S ribosomal RNA (rRNA) gene, the factors that control the distribution and magnitude of high-affinity methanotrophy in the Arctic permafrost-affected region have remained elusive. Using 21 metagenomic datasets of surface soils obtained from long-term core incubation experiments,1 this bioinformatics study aimed to reconstruct the draft genome of the Upland Soil Cluster α-proteobacteria (USCα), the high-affinity methanotroph previously detected in the samples,2 and to determine its phylogeny and metabolic requirements. We obtained a genome bin containing the high-affinity form of the USCα-like pmoA gene. The 3.03 Mbp assembly is 91.6% complete with a unique set of single-copy marker genes. The 16S rRNA gene fragment of USCα belongs to the α-proteobacterial family Beijerinckiaceae. Genome annotation indicates possible formaldehyde oxidation via tetrahydromethanopterin-linked C1 transfer pathways, acetate utilization, carbon fixation via the Calvin-Benson-Bassham cycle, and glycogen production. Notably, the key enzymes for formaldehyde assimilation via the serine and ribulose monophosphate pathways are missing. The presence of genes encoding nitrate reductase and hemoglobin suggests adaptation to low O2 under water-logged conditions. Since USCα has versatile carbon metabolisms, it may not be an obligate methanotroph

Identification and properties of very high affinity brain membrane-binding sites for a neurotoxic phospholipase from the taipan venom

Energy Technology Data Exchange (ETDEWEB)

Lambeau, G.; Barhanin, J.; Schweitz, H.; Qar, J.; Lazdunski, M. (Centre de Biochimie, Nice (France))

1989-07-05

Four new monochain phospholipases were purified from the Oxyuranus scutellatus (taipan) venom. Three of them were highly toxic when injected into mice brain. One of these neurotoxic phospholipases, OS2, was iodinated and used in binding experiments to demonstrate the presence of two families of specific binding sites in rat brain synaptic membranes. The affinities were exceptionally high, Kd1 = 1.5 +/- 0.5 pM and Kd2 = 45 +/- 10 pM, and the maximal binding capacities were Bmax 1 = 1 +/- 0.4 and Bmax 2 = 3 +/- 0.5 pmol/mg of protein. Both binding sites were sensitive to proteolysis and demonstrated to be located on proteins of Mr 85,000-88,000 and 36,000-51,000 by cross-linking and photoaffinity labeling techniques. The binding of {sup 125}I-OS2 to synaptic membranes was dependent on Ca2+ ions and enhanced by Zn2+ ions which inhibit phospholipase activity. Competition experiments have shown that, except for beta-bungarotoxin, a number of known toxic snake or bee phospholipases have very high affinities for the newly identified binding sites. A good correlation (r = 0.80) was observed between toxicity and affinity but not between phospholipase activity and affinity.

Structural insights into a high affinity nanobody:antigen complex by homology modelling.

Science.gov (United States)

Skottrup, Peter Durand

2017-09-01

Porphyromonas gingivalis is a major periodontitis-causing pathogens. P. gingivalis secrete a cysteine protease termed RgpB, which is specific for Arg-Xaa bonds in substrates. Recently, a nanobody-based assay was used to demonstrate that RgpB could represent a novel diagnostic target, thereby simplifying. P. gingivalis detection. The nanobody, VHH7, had a high binding affinity and was specific for RgpB, when tested towards the highly identical RgpA. In this study a homology model of VHH7 was build. The complementarity determining regions (CDR) comprising the paratope residues responsible for RgpB binding were identified and used as input to the docking. Furthermore, residues likely involved in the RgpB epitope was identified based upon RgpB:RgpA alignment and analysis of residue surface accessibility. CDR residues and putitative RgpB epitope residues were used as input to an information-driven flexible docking approach using the HADDOCK server. Analysis of the VHH7:RgpB model demonstrated that the epitope was found in the immunoglobulin-like domain and residue pairs located at the molecular paratope:epitope interface important for complex stability was identified. Collectively, the VHH7 homology model and VHH7:RgpB docking supplies knowledge of the residues involved in the high affinity interaction. This information could prove valuable in the design of an antibody-drug conjugate for specific RgpB targeting. Copyright © 2017 Elsevier Inc. All rights reserved.

Characterization of high affinity [3H]triazolam binding in rat brain

International Nuclear Information System (INIS)

Earle, M.; Concas, A.; Yamamura, H.I.

1986-01-01

The hypnotic Triazolam (TZ), a triazolo (1,4)-benzodiazepine, displays a short physiological half life and has been used for the treatment of insomnia related to anxiety states. Specific binding properties of this recently tritiated TZ were characterized. The authors major objectives were the direct measurement of the temperature dependence and the GABA effect on [ 3 H]TZ binding. Saturation studies showed a shift to lower affinity at 37 0 C (K/sub d/ = 0.25 +/- 0.01 nM at O 0 C; K/sub d/ = 1.46 +/- 0.03 nM at 37 0 C) while the B/sub max/ values remained unchanged (1003 +/- 37 fmoles/mg prot. at 0 0 C and 1001 +/- 43 fmoles/mg prot. at 37 0 C). Inhibition studies showed that [ 3 H]TZ binding displayed no GABA shift at 0 0 C(K/sub i/ 0.37 +/- 0.03 nM/- GABA and K/sub i/ = 0.55 +/- 0.13 nM/+GABA) but a nearly two-fold shift was apparent at 37 0 C (K/sub i/ = 2.92 +/- 0.2 nM/-GABA; K/sub i/ = 1.37 +/- 0.11 mM/+GABA). These results were also confirmed by saturation studies in the presence or absence of GABA showing a shift to higher affinity in the presence of GABA only at 37 0 C. In Ro 15-1788/[ 3 H]TZ competition experiments the presence of GABA did not affect the inhibitory potency of Ro 15-1788 on [ 3 H]TZ binding at both temperatures. In conclusion [ 3 H]TZ binding showed an extremely high affinity for benzodiazepine receptors. In contrast to reported literature, the findings suggest that TZ interacts with benzodiazepine receptors similar to other benzodiazepine agonists

Glucose uptake and growth of glucose-limited chemostat cultures of Aspergillus niger and a disruptant lacking MstA, a high-affinity glucose transporter

DEFF Research Database (Denmark)

Jørgensen, Thomas R; vanKuyk, Patricia A; Poulsen, Bjarne R

2007-01-01

This is a study of high-affinity glucose uptake in Aspergillus niger and the effect of disruption of a high-affinity monosaccharide-transporter gene, mstA. The substrate saturation constant (K(s)) of a reference strain was about 15 microM in glucose-limited chemostat culture. Disruption of mst......-affinity uptake system of A. niger. The mstA disruptant and a reference strain were cultivated in glucose-limited chemostat cultures at low, intermediate and high dilution rate (D=0.07 h(-1), 0.14 h(-1) and 0.20 h(-1)). Mycelium harvested from steady-state cultures was subjected to glucose uptake assays...

A Novel Recombinant DNA System for High Efficiency Affinity Purification of Proteins in Saccharomyces cerevisiae

Directory of Open Access Journals (Sweden)

Brian H. Carrick

2016-03-01

Full Text Available Isolation of endogenous proteins from Saccharomyces cerevisiae has been facilitated by inserting encoding polypeptide affinity tags at the C-termini of chromosomal open reading frames (ORFs using homologous recombination of DNA fragments. Tagged protein isolation is limited by a number of factors, including high cost of affinity resins for bulk isolation and low concentration of ligands on the resin surface, leading to low isolation efficiencies and trapping of contaminants. To address this, we have created a recombinant “CelTag” DNA construct from which PCR fragments can be created to easily tag C-termini of S. cerevisiae ORFs using selection for a nat1 marker. The tag has a C-terminal cellulose binding module to be used in the first affinity step. Microgranular cellulose is very inexpensive and has an effectively continuous ligand on its surface, allowing rapid, highly efficient purification with minimal background. Cellulose-bound proteins are released by specific cleavage of an included site for TEV protease, giving nearly pure product. The tag can be lifted from the recombinant DNA construct either with or without a 13x myc epitope tag between the target ORF and the TEV protease site. Binding of CelTag protein fusions to cellulose is stable to high salt, nonionic detergents, and 1 M urea, allowing stringent washing conditions to remove loosely associated components, as needed, before specific elution. It is anticipated that this reagent could allow isolation of protein complexes from large quantities of yeast extract, including soluble, membrane-bound, or nucleic acid-associated assemblies.

Discovery of PF-06928215 as a high affinity inhibitor of cGAS enabled by a novel fluorescence polarization assay

Energy Technology Data Exchange (ETDEWEB)

Hall, Justin; Brault, Amy; Vincent, Fabien; Weng, Shawn; Wang, Hong; Dumlao, Darren; Aulabaugh, Ann; Aivazian, Dikran; Castro, Dana; Chen, Ming; Culp, Jeffrey; Dower, Ken; Gardner, Joseph; Hawrylik, Steven; Golenbock, Douglas; Hepworth, David; Horn, Mark; Jones, Lyn; Jones, Peter; Latz, Eicke; Li, Jing; Lin, Lih-Ling; Lin, Wen; Lin, David; Lovering, Frank; Niljanskul, Nootaree; Nistler, Ryan; Pierce, Betsy; Plotnikova, Olga; Schmitt, Daniel; Shanker, Suman; Smith, James; Snyder, William; Subashi, Timothy; Trujillo, John; Tyminski, Edyta; Wang, Guoxing; Wong, Jimson; Lefker, Bruce; Dakin, Leslie; Leach, Karen (UMASS, MED); (Pfizer)

2017-09-21

Cyclic GMP-AMP synthase (cGAS) initiates the innate immune system in response to cytosolic dsDNA. After binding and activation from dsDNA, cGAS uses ATP and GTP to synthesize 2', 3' -cGAMP (cGAMP), a cyclic dinucleotide second messenger with mixed 2'-5' and 3'-5' phosphodiester bonds. Inappropriate stimulation of cGAS has been implicated in autoimmune disease such as systemic lupus erythematosus, thus inhibition of cGAS may be of therapeutic benefit in some diseases; however, the size and polarity of the cGAS active site makes it a challenging target for the development of conventional substrate-competitive inhibitors. We report here the development of a high affinity (K_D = 200 nM) inhibitor from a low affinity fragment hit with supporting biochemical and structural data showing these molecules bind to the cGAS active site. We also report a new high throughput cGAS fluorescence polarization (FP)-based assay to enable the rapid identification and optimization of cGAS inhibitors. This FP assay uses Cy5-labelled cGAMP in combination with a novel high affinity monoclonal antibody that specifically recognizes cGAMP with no cross reactivity to cAMP, cGMP, ATP, or GTP. Given its role in the innate immune response, cGAS is a promising therapeutic target for autoinflammatory disease. Our results demonstrate its druggability, provide a high affinity tool compound, and establish a high throughput assay for the identification of next generation cGAS inhibitors.

A role for tight junction-associated MARVEL proteins in larval sea lamprey (Petromyzon marinus) osmoregulation.

Science.gov (United States)

Kolosov, Dennis; Bui, Phuong; Donini, Andrew; Wilkie, Mike P; Kelly, Scott P

2017-10-15

This study reports on tight junction-associated MARVEL proteins of larval sea lamprey ( Petromyzon marinus ) and their potential role in ammocoete osmoregulation. Two occludin isoforms (designated Ocln and Ocln-a) and a tricellulin (Tric) were identified. Transcripts encoding ocln , ocln-a and tric were broadly expressed in larval lamprey, with the greatest abundance of ocln in the gut, liver and kidney, ocln-a in the gill and skin, and tric in the kidney. Ocln and Ocln-a resolved as ∼63 kDa and ∼35 kDa MW proteins, respectively, while Tric resolved as a ∼50 kDa protein. Ocln immunolocalized to the gill vasculature and in gill mucous cells while Ocln-a localized to the gill pouch and gill epithelium. Both Ocln and Ocln-a localized in the nephron, the epidermis and the luminal side of the gut. In branchial tissue, Tric exhibited punctate localization, consistent with its presence at regions of tricellular contact. Following ion-poor water (IPW) acclimation of ammocoetes, serum [Na + ] and [Cl - ] decreased, but not [Ca 2+ ], and carcass moisture content increased. In association, Ocln abundance increased in the skin and kidney, but reduced in the gill of IPW-acclimated ammocoetes while Ocln-a abundance reduced in the kidney only. Tric abundance increased in the gill. Region-specific alterations in ocln , ocln-a and tric mRNA abundance were also observed in the gut. Data support a role for Ocln, Ocln-a and Tric in the osmoregulatory strategies of a basal vertebrate. © 2017. Published by The Company of Biologists Ltd.

Topography of the high-affinity lysine binding site of plasminogen as defined with a specific antibody probe

International Nuclear Information System (INIS)

Miles, L.A.; Plow, E.F.

1986-01-01

An antibody population that reacted with the high-affinity lysine binding site of human plasminogen was elicited by immunizing rabbits with an elastase degradation product containing kringles 1-3 (EDP I). This antibody was immunopurified by affinity chromatography on plasminogen-Sepharose and elution with 0.2 M 6-aminohexanoic acid. The eluted antibodies bound [ 125 I]EDP I, [ 125 I]Glu-plasminogen, and [ 125 I]Lys-plasminogen in radioimmunoassays, and binding of each ligand was at least 99% inhibited by 0.2 M 6-aminohexanoic acid. The concentrations for 50% inhibition of [ 125 I]EDP I binding by tranexamic acid, 6-aminohexanoic acid, and lysine were 2.6, 46, and l730 μM, respectively. Similar values were obtained with plasminogen and suggested that an unoccupied high-affinity lysine binding site was required for antibody recognition. The antiserum reacted exclusively with plasminogen derivatives containing the EDP I region and did not react with those lacking an EDP I region, or with tissue plasminogen activator or prothrombin, which also contains kringles. By immunoblotting analyses, a chymotryptic degradation product of M/sub r/ 20,000 was derived from EDP I that retained reactivity with the antibody. α 2 -Antiplasmin inhibited the binding of radiolabeled EDP I, Glu-plasminogen, or Lys-plasminogen by the antiserum, suggesting that the recognized site is involved in the noncovalent interaction of the inhibitor with plasminogen. The binding of [ 125 I]EDP I to fibrin was also inhibited by the antiserum. The observations provide independent evidence for the role of the high-affinity lysine binding site in the functional interactions of plasminogen with its primary substrate and inhibitor

Visual and Plasmon Resonance Absorption Sensor for Adenosine Triphosphate Based on the High Affinity between Phosphate and Zr(IV)

OpenAIRE

Qi, Wenjing; Liu, Zhongyuan; Zhang, Wei; Halawa, Mohamed Ibrahim; Xu, Guobao

2016-01-01

Zr(IV) can form phosphate and Zr(IV) (?PO3 2??Zr4+?) complex owing to the high affinity between Zr(IV) with phosphate. Zr(IV) can induce the aggregation of gold nanoparticles (AuNPs), while adenosine triphosphate(ATP) can prevent Zr(IV)-induced aggregation of AuNPs. Herein, a visual and plasmon resonance absorption (PRA)sensor for ATP have been developed using AuNPs based on the high affinity between Zr(IV)with ATP. AuNPs get aggregated in the presence of certain concentrations of Zr(IV). Aft...

14-O-Methylmorphine: A Novel Selective Mu-Opioid Receptor Agonist with High Efficacy and Affinity.

Science.gov (United States)

Zádor, Ferenc; Balogh, Mihály; Váradi, András; Zádori, Zoltán S; Király, Kornél; Szűcs, Edina; Varga, Bence; Lázár, Bernadette; Hosztafi, Sándor; Riba, Pál; Benyhe, Sándor; Fürst, Susanna; Al-Khrasani, Mahmoud

2017-11-05

14-O-methyl (14-O-Me) group in morphine-6-O-sulfate (M6SU) or oxymorphone has been reported to be essential for enhanced affinity, potency and antinociceptive effect of these opioids. Herein we report on the pharmacological properties (potency, affinity and efficacy) of the new compound, 14-O-methylmorphine (14-O-MeM) in in vitro. Additionally, we also investigated the antinociceptive effect of the novel compound, as well as its inhibitory action on gastrointestinal transit in in vivo. The potency and efficacy of test compound were measured by [ 35 S]GTPγS binding, isolated mouse vas deferens (MVD) and rat vas deferens (RVD) assays. The affinity of 14-O-MeM for opioid receptors was assessed by radioligand binding and MVD assays. The antinociceptive and gastrointestinal effects of the novel compound were evaluated in the rat tail-flick test and charcoal meal test, respectively. Morphine, DAMGO, Ile 5,6 deltorphin II, deltorphin II and U-69593 were used as reference compounds. 14-O-MeM showed higher efficacy (E max ) and potency (EC 50 ) than morphine in MVD, RVD or [ 35 S]GTPγS binding. In addition, 14-O-MeM compared to morphine showed higher affinity for μ-opioid receptor (MOR). In vivo, in rat tail-flick test 14-O-MeM proved to be stronger antinociceptive agent than morphine after peripheral or central administration. Additionally, both compounds inhibited the gastrointestinal peristalsis. However, when the antinociceptive and antitransit doses for each test compound are compared, 14-O-MeM proved to have slightly more favorable pharmacological profile. Our results affirm that 14-O-MeM, an opioid of high efficacy and affinity for MOR can be considered as a novel analgesic agent of potential clinical value. Copyright © 2017 Elsevier B.V. All rights reserved.

Botulinum neurotoxin B recognizes its protein receptor with high affinity and specificity.

Science.gov (United States)

Jin, Rongsheng; Rummel, Andreas; Binz, Thomas; Brunger, Axel T

2006-12-21

Botulinum neurotoxins (BoNTs) are produced by Clostridium botulinum and cause the neuroparalytic syndrome of botulism. With a lethal dose of 1 ng kg(-1), they pose a biological hazard to humans and a serious potential bioweapon threat. BoNTs bind with high specificity at neuromuscular junctions and they impair exocytosis of synaptic vesicles containing acetylcholine through specific proteolysis of SNAREs (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptors), which constitute part of the synaptic vesicle fusion machinery. The molecular details of the toxin-cell recognition have been elusive. Here we report the structure of a BoNT in complex with its protein receptor: the receptor-binding domain of botulinum neurotoxin serotype B (BoNT/B) bound to the luminal domain of synaptotagmin II, determined at 2.15 A resolution. On binding, a helix is induced in the luminal domain which binds to a saddle-shaped crevice on a distal tip of BoNT/B. This crevice is adjacent to the non-overlapping ganglioside-binding site of BoNT/B. Synaptotagmin II interacts with BoNT/B with nanomolar affinity, at both neutral and acidic endosomal pH. Biochemical and neuronal ex vivo studies of structure-based mutations indicate high specificity and affinity of the interaction, and high selectivity of BoNT/B among synaptotagmin I and II isoforms. Synergistic binding of both synaptotagmin and ganglioside imposes geometric restrictions on the initiation of BoNT/B translocation after endocytosis. Our results provide the basis for the rational development of preventive vaccines or inhibitors against these neurotoxins.

Acylated heptapeptide binds albumin with high affinity and application as tag furnishes long-acting peptides.

Science.gov (United States)

Zorzi, Alessandro; Middendorp, Simon J; Wilbs, Jonas; Deyle, Kaycie; Heinis, Christian

2017-07-17

The rapid renal clearance of peptides in vivo limits this attractive platform for the treatment of a broad range of diseases that require prolonged drug half-lives. An intriguing approach for extending peptide circulation times works through a 'piggy-back' strategy in which peptides bind via a ligand to the long-lived serum protein albumin. In accordance with this strategy, we developed an easily synthesized albumin-binding ligand based on a peptide-fatty acid chimera that has a high affinity for human albumin (K d =39 nM). This ligand prolongs the elimination half-life of cyclic peptides in rats 25-fold to over seven hours. Conjugation to a peptide factor XII inhibitor developed for anti-thrombotic therapy extends the half-life from 13 minutes to over five hours, inhibiting coagulation for eight hours in rabbits. This high-affinity albumin ligand could potentially extend the half-life of peptides in human to several days, substantially broadening the application range of peptides as therapeutics.

Acylated heptapeptide binds albumin with high affinity and application as tag furnishes long-acting peptides

Science.gov (United States)

Zorzi, Alessandro; Middendorp, Simon J.; Wilbs, Jonas; Deyle, Kaycie; Heinis, Christian

2017-07-01

The rapid renal clearance of peptides in vivo limits this attractive platform for the treatment of a broad range of diseases that require prolonged drug half-lives. An intriguing approach for extending peptide circulation times works through a `piggy-back' strategy in which peptides bind via a ligand to the long-lived serum protein albumin. In accordance with this strategy, we developed an easily synthesized albumin-binding ligand based on a peptide-fatty acid chimera that has a high affinity for human albumin (Kd=39 nM). This ligand prolongs the elimination half-life of cyclic peptides in rats 25-fold to over seven hours. Conjugation to a peptide factor XII inhibitor developed for anti-thrombotic therapy extends the half-life from 13 minutes to over five hours, inhibiting coagulation for eight hours in rabbits. This high-affinity albumin ligand could potentially extend the half-life of peptides in human to several days, substantially broadening the application range of peptides as therapeutics.

Structural implications of hERG K+ channel block by a high-affinity minimally structured blocker

Science.gov (United States)

Helliwell, Matthew V.; Zhang, Yihong; El Harchi, Aziza; Du, Chunyun; Hancox, Jules C.; Dempsey, Christopher E.

2018-01-01

Cardiac potassium channels encoded by human ether-à-go-go–related gene (hERG) are major targets for structurally diverse drugs associated with acquired long QT syndrome. This study characterized hERG channel inhibition by a minimally structured high-affinity hERG inhibitor, Cavalli-2, composed of three phenyl groups linked by polymethylene spacers around a central amino group, chosen to probe the spatial arrangement of side chain groups in the high-affinity drug-binding site of the hERG pore. hERG current (IhERG) recorded at physiological temperature from HEK293 cells was inhibited with an IC50 of 35.6 nm with time and voltage dependence characteristic of blockade contingent upon channel gating. Potency of Cavalli-2 action was markedly reduced for attenuated inactivation mutants located near (S620T; 54-fold) and remote from (N588K; 15-fold) the channel pore. The S6 Y652A and F656A mutations decreased inhibitory potency 17- and 75-fold, respectively, whereas T623A and S624A at the base of the selectivity filter also decreased potency (16- and 7-fold, respectively). The S5 helix F557L mutation decreased potency 10-fold, and both F557L and Y652A mutations eliminated voltage dependence of inhibition. Computational docking using the recent cryo-EM structure of an open channel hERG construct could only partially recapitulate experimental data, and the high dependence of Cavalli-2 block on Phe-656 is not readily explainable in that structure. A small clockwise rotation of the inner (S6) helix of the hERG pore from its configuration in the cryo-EM structure may be required to optimize Phe-656 side chain orientations compatible with high-affinity block. PMID:29545312

Adult Pacific Lamprey Migration Behavior and Escapement in the Bonneville Reservoir and Lower Columbia River Monitored Using the Juvenile Salmonid Acoustic Telemetry System (JSATS), 2011

Science.gov (United States)

2012-01-01

sea lions (Zalophus californicus, Eumetopias jubatus) or white sturgeon (Acipenser transmontanus) in the tailrace, moved to spawning tributaries...and management of three parasitic lampreys of North America. Fisheries 35:580-594. Close, D. A., M. Fitzpatrick, and H. Li. 2002. The ecological

Hirota's solitons in the affine and the conformal affine Toda models

International Nuclear Information System (INIS)

Aratyn, H.; Constantinidis, C.P.; Ferreira, L.A.; Gomes, J.F.; Zimerman, A.H.

1993-01-01

We use Hirota's method formulated as a recursive scheme to construct a complete set of soliton solutions for the affine Toda field theory based on an arbitrary Lie algebra. Our solutions include a new class of solitons connected with two different types of degeneracies encountered in Hirota's perturbation approach. We also derive an universal mass formula for all Hirota's solutions to the affine Toda model valid for all underlying Lie groups. Embedding of the affine Toda model in the conformal affine Toda model plays a crucial role in this analysis. (orig.)

Improving image segmentation by learning region affinities

Energy Technology Data Exchange (ETDEWEB)

Prasad, Lakshman [Los Alamos National Laboratory; Yang, Xingwei [TEMPLE UNIV.; Latecki, Longin J [TEMPLE UNIV.

2010-11-03

We utilize the context information of other regions in hierarchical image segmentation to learn new regions affinities. It is well known that a single choice of quantization of an image space is highly unlikely to be a common optimal quantization level for all categories. Each level of quantization has its own benefits. Therefore, we utilize the hierarchical information among different quantizations as well as spatial proximity of their regions. The proposed affinity learning takes into account higher order relations among image regions, both local and long range relations, making it robust to instabilities and errors of the original, pairwise region affinities. Once the learnt affinities are obtained, we use a standard image segmentation algorithm to get the final segmentation. Moreover, the learnt affinities can be naturally unutilized in interactive segmentation. Experimental results on Berkeley Segmentation Dataset and MSRC Object Recognition Dataset are comparable and in some aspects better than the state-of-art methods.

The solutions of affine and conformal affine Toda field theory

International Nuclear Information System (INIS)

Papadopoulos, G.; Spence, B.

1994-02-01

We give new formulations of the solutions of the field equations of the affine Toda and conformal affine Toda theories on a cylinder and two-dimensional Minkowski space-time. These solutions are parameterised in terms of initial data and the resulting covariant phase spaces are diffeomorphic to the Hamiltonian ones. We derive the fundamental Poisson brackets of the parameters of the solutions and give the general static solutions for the affine theory. (authors). 10 refs

Influence of self-affine roughness on the friction coefficient of rubber at high sliding velocity

NARCIS (Netherlands)

Palasantzas, G

2004-01-01

In this work we investigate the influence of self-affine roughness on the friction coefficient of a rubber body onto a solid surface at high speeds. The roughness is characterized by the rms amplitude w, the correlation length xi, and the roughness exponent H. It is shown that the friction

Lamprey Prohibitin2 Arrest G2/M Phase Transition of HeLa Cells through Down-regulating Expression and Phosphorylation Level of Cell Cycle Proteins.

Science.gov (United States)

Shi, Ying; Guo, Sicheng; Wang, Ying; Liu, Xin; Li, Qingwei; Li, Tiesong

2018-03-02

Prohibitin 2(PHB2) is a member of the SFPH trans-membrane family proteins. It is a highly conserved and functionally diverse protein that plays an important role in preserving the structure and function of the mitochondria. In this study, the lamprey PHB2 gene was expressed in HeLa cells to investigate its effect on cell proliferation. The effect of Lm-PHB2 on the proliferation of HeLa cells was determined by treating the cells with pure Lm-PHB2 protein followed by MTT assay. Using the synchronization method with APC-BrdU and PI double staining revealed rLm-PHB2 treatment induced the decrease of both S phase and G0/G1 phase and then increase of G2/M phase. Similarly, cells transfected with pEGFP-N1-Lm-PHB2 also exhibited remarkable reduction in proliferation. Western blot and quantitative real-time PCR(qRT-PCR) assays suggested that Lm-PHB2 caused cell cycle arrest in HeLa cells through inhibition of CDC25C and CCNB1 expression. According to our western blot analysis, Lm-PHB2 was also found to reduce the expression level of Wee1 and PLK1 and the phosphorylation level of CCNB1, CDC25C and CDK1 in HeLa cells. Lamprey prohibitin 2 could arrest G2/M phase transition of HeLa cells through down-regulating expression and phosphorylation level of cell cycle proteins.

Different endothelin receptor affinities in dog tissues

International Nuclear Information System (INIS)

Loeffler, B.M.L.; Loehrer, W.

1991-01-01

Endothelin (ET) is a long-lasting potent vasoconstrictor-peptide. Here the authors report different binding affinities of endothelin-1 (ET-1) to ET-receptors of various dog tissues. Crude microsomal fractions were prepared after homogenisation of dog tissues in 50 mM Tris/HCl, 20 mM MnCl2, 1 mM EDTA, pH 7.4 by differential centrifugation. Aliquots of microsomal fractions (70 micrograms of protein) were incubated at 25 degrees C for 180 min in the presence of 20 pM 125I-ET-1 and various concentrations of cold ET-1. Four different ET-1 receptor binding affinities were found: adrenals, cerebrum, liver, heart, skeletal muscle and stomach microsomal membranes contained high affinity binding sites (Kd 50 - 80 pM, Bmax 60 - 250 fmol/mg). In cerebellum and spleen medium affinity ET-1 receptors (Kd 350 pM, Bmax 880 and 1200 fmol/mg respectively) were present. In comparison lung and kidney microsomes contained a low affinity ET-1 receptor (Kd 800 and 880 pM, Bmax 1600 and 350 fmol/mg). Receptors of even lower affinity were present in heart, intestine and liver microsomes with Kd values of 3 - 6 nM

Low affinity uniporter carrier proteins can increase net substrate uptake rate by reducing efflux

NARCIS (Netherlands)

Bosdriesz, Evert; Wortel, Meike T.; Haanstra, Jurgen R.; Wagner, Marijke J.; De La Torre Cortés, Pilar; Teusink, Bas

2018-01-01

Many organisms have several similar transporters with different affinities for the same substrate. Typically, high-affinity transporters are expressed when substrate is scarce and low-affinity ones when it is abundant. The benefit of using low instead of high-affinity transporters remains unclear,

Low affinity uniporter carrier proteins can increase net substrate uptake rate by reducing efflux

NARCIS (Netherlands)

Bosdriesz, Evert; Wortel, M.T.; Haanstra, Jurgen R.; Wagner, Marijke J.; De La Torre, P.; Teusink, Bas

2018-01-01

Many organisms have several similar transporters with different affinities for the same substrate. Typically, high-affinity transporters are expressed when substrate is scarce and low-affinity ones when it is abundant. The benefit of using low instead of high-affinity transporters remains

Lectin affinity electrophoresis.

Science.gov (United States)

Kobayashi, Yuka

2014-01-01

An interaction or a binding event typically changes the electrophoretic properties of a molecule. Affinity electrophoresis methods detect changes in the electrophoretic pattern of molecules (mainly macromolecules) that occur as a result of biospecific interactions or complex formation. Lectin affinity electrophoresis is a very effective method for the detection and analysis of trace amounts of glycobiological substances. It is particularly useful for isolating and separating the glycoisomers of target molecules. Here, we describe a sensitive technique for the detection of glycoproteins separated by agarose gel-lectin affinity electrophoresis that uses antibody-affinity blotting. The technique is tested using α-fetoprotein with lectin (Lens culinaris agglutinin and Phaseolus vulgaris agglutinin)-agarose gels.

High-Affinity Accumulation of Chloroquine by Mouse Erythrocytes Infected with Plasmodium berghei

Science.gov (United States)

Fitch, Coy D.; Yunis, Norman G.; Chevli, Rekha; Gonzalez, Yolanda

1974-01-01

Washed erythrocytes infected with chloroquine-susceptible (CS) or with chloroquine-resistant (CR) P. berghei were used in model systems in vitro to study the accumulation of chloroquine with high affinity. The CS model could achieve distribution ratios (chloroquine in cells: chloroquine in medium) of 100 in the absence of substrate. 200—300 in the presence of 10 mM pyruvate or lactate, and over 600 in the presence of 1 mM glucose or glycerol. In comparable studies of the CR model, the distribution ratios were 100 in the absence of substrate and 300 or less in the presence of glucose or glycerol. The presence of lactate stimulated chloroquine accumulation in the CR model, whereas the presence of pyruvate did not. Lactate production from glucose and glycerol was undiminished in the CR model, and ATP concentrations were higher than in the CS model. Cold, iodoacetate, 2,4-dinitrophenol, or decreasing pH inhibited chloroquine accumulation in both models. These findings demonstrate substrate involvement in the accumulation of chloroquine with high affinity. In studies of the CS model, certain compounds competitively inhibited chloroquine accumulation, while others did not. This finding is attributable to a specific receptor that imposes structural constraints on the process of accumulation. For chloroquine analogues, the position and length of the side chain, the terminal nitrogen atom of the side chain, and the nitrogen atom in the quinoline ring are important determinants of binding to this receptor. PMID:4600044

Na+-Dependent High-Affinity Nitrate, Phosphate and Amino Acids Transport in Leaf Cells of the Seagrass Posidonia oceanica (L. Delile

Directory of Open Access Journals (Sweden)

Lourdes Rubio

2018-05-01

Full Text Available Posidonia oceanica (L. Delile is a seagrass, the only group of vascular plants to colonize the marine environment. Seawater is an extreme yet stable environment characterized by high salinity, alkaline pH and low availability of essential nutrients, such as nitrate and phosphate. Classical depletion experiments, membrane potential and cytosolic sodium measurements were used to characterize the high-affinity NO3−, Pi and amino acids uptake mechanisms in this species. Net uptake rates of both NO3− and Pi were reduced by more than 70% in the absence of Na+. Micromolar concentrations of NO3− depolarized mesophyll leaf cells plasma membrane. Depolarizations showed saturation kinetics (Km = 8.7 ± 1 μM NO3−, which were not observed in the absence of Na+. NO3− induced depolarizations at increasing Na+ also showed saturation kinetics (Km = 7.2 ± 2 mM Na+. Cytosolic Na+ measured in P. oceanica leaf cells (17 ± 2 mM Na+ increased by 0.4 ± 0.2 mM Na+ upon the addition of 100 μM NO3−. Na+-dependence was also observed for high-affinity l-ala and l-cys uptake and high-affinity Pi transport. All together, these results strongly suggest that NO3−, amino acids and Pi uptake in P. oceanica leaf cells are mediated by high-affinity Na+-dependent transport systems. This mechanism seems to be a key step in the process of adaptation of seagrasses to the marine environment.

High Affinity IgE-Fc Receptor alpha and gamma Subunit Interactions

International Nuclear Information System (INIS)

Rashid, A.; Housden, J. E. M.; Sabban, S.; Helm, B.

2014-01-01

Objective: To explore the relationships between the subunits (alpha, beta and gamma) of the high affinity IgE receptor (Fc and RI) and its ability to mediate transmembrane signaling. Study Design: Experimental study. Place and Duration of Study: Department of Molecular Biology and Biotechnology, University of Sheffield, UK, from 2008 to 2009. Methodology: The approach employed was to create a chimera (human alpha-gamma-gamma) using the extracellular (EC) domain of the human high affinity IgE receptor. The alpha subunit (huFc and RIalpha) of IgE receptor was spliced onto the rodent gamma TM and cytoplasmic domain (CD). This was transfected into the Rat Basophilic Leukemia cell line in order to assess the possibility of selectively activating cells transfected with this single pass construct for antigen induced mediator release. Results: The RBLs cell lines transfected with the huFc and RIalpha/gamma/gamma cDNA constructs were assessed for the cell surface expression of the huFc and RIalpha subunit and the response to the antigenic stimulus by looking for degranulation and intracellular Ca2+ mobilisation. The results obtained showed the absence of huFc and RIalpha subunit expression on the surface of transfected cells as seen by flowcytometric studies, beta-hexosaminidase assays and intracellular calcium mobilisation studies. Conclusion: In the present study the grounds for non-expression of huFc and RIalpha/gamma/gamma cDNA remains elusive but may be due to the fact that the human-rodent chimeric receptors are assembled differently than the endogenous rodent receptors as seen in study in which COS 7 cells were transfected with human/rat chimeric complexes. (author)

Female sea lamprey shift orientation toward a conspecific chemical cue to escape a sensory trap

Science.gov (United States)

Brant, Cory O.; Johnson, Nicholas; Li, Ke; Buchinger, Tyler J.; Li, Weiming

2016-01-01

The sensory trap model of signal evolution hypothesizes that signalers adapt to exploit a cue used by the receiver in another context. Although exploitation of receiver biases can result in conflict between the sexes, deceptive signaling systems that are mutually beneficial drive the evolution of stable communication systems. However, female responses in the nonsexual and sexual contexts may become uncoupled if costs are associated with exhibiting a similar response to a trait in both contexts. Male sea lamprey (Petromyzon marinus) signal with a mating pheromone, 3-keto petromyzonol sulfate (3kPZS), which may be a match to a juvenile cue used by females during migration. Upstream movement of migratory lampreys is partially guided by 3kPZS, but females only move toward 3kPZS with proximal accuracy during spawning. Here, we use in-stream behavioral assays paired with gonad histology to document the transition of female preference for juvenile- and male-released 3kPZS that coincides with the functional shift of 3kPZS as a migratory cue to a mating pheromone. Females became increasingly biased toward the source of synthesized 3kPZS as their maturation progressed into the reproductive phase, at which point, a preference for juvenile odor (also containing 3kPZS naturally) ceased to exist. Uncoupling of female responses during migration and spawning makes the 3kPZS communication system a reliable means of synchronizing mate search. The present study offers a rare example of a transition in female responses to a chemical cue between nonsexual and sexual contexts, provides insights into the origins of stable communication signaling systems.

N-Oxide analogs of WAY-100635 : new high affinity 5-HT (1A) receptor antagonists

NARCIS (Netherlands)

Oberwinkler - Marchais, Sandrine; Nowicki, B; Pike, VW; Halldin, C; Sandell, J; Chou, YH; Gulyas, B; Brennum, LT; Farde, L; Wikstrom, H V

2005-01-01

WAY-100635 [N-(2-(1-(4-(2-methoxyphenyl)piperazinyl)ethyl))-N-(2-pyridinyl)cyclohexanecarboxamide] 1 and its O-des-methyl derivative DWAY 2 are well-known high affinity 5-HT1A receptor antagonists. which when labeled with carbon-II (beta(+): t(1/2) 20.4min) in the carbonyl group are effective

Affinity chromatography with pseudobiospecific ligands on high-performance supports for purification of proteins of biotechnological interest

Directory of Open Access Journals (Sweden)

N.B. Iannucci

2003-03-01

Full Text Available High-performance affinity matrices were obtained by attaching pseudobiospecific ligands to hollow-fibre membranes. The neutral protease contained in FlavourzymeTM was purified to homogeneity with Yellow 4R-HE affinity hollow-fibre membranes. Immobilisation of Red HE-3B allowed purification of a milk-clotting enzyme obtained by solid-state culture of Mucor bacilliformis. Copper immobilisation through iminodiacetic acid allowed fractionation of Biocon Bioconcentrated PlusTM to separate the pectinesterase-containing fraction. The productivity of the developed processes - 1900, 94 and 750 U/ml.min, respectively - was 10- to 15-fold higher than that achieved with the same ligands immobilised on agarose-based soft gels, mainly due to the shortening of the purification processes.

Fenobody: A Ferritin-Displayed Nanobody with High Apparent Affinity and Half-Life Extension.

Science.gov (United States)

Fan, Kelong; Jiang, Bing; Guan, Zhe; He, Jiuyang; Yang, Dongling; Xie, Ni; Nie, Guohui; Xie, Can; Yan, Xiyun

2018-04-10

Nanobodies consist of a single domain variable fragment of a camelid heavy-chain antibody. Nanobodies have potential applications in biomedical fields because of their simple production procedures and low cost. Occasionally, nanobody clones of interest exhibit low affinities for their target antigens, which, together with their short half-life limit bioanalytical or therapeutic applications. Here, we developed a novel platform we named fenobody, in which a nanobody developed against H5N1 virus is displayed on the surface of ferritin in the form of a 24mer. We constructed a fenobody by substituting the fifth helix of ferritin with the nanobody. TEM analysis showed that nanobodies were displayed on the surface of ferritin in the form of 6 × 4 bundles, and that these clustered nanobodies are flexible for antigen binding in spatial structure. Comparing fenobodies with conventional nanobodies currently used revealed that the antigen binding apparent affinity of anti-H5N1 fenobody was dramatically increased (∼360-fold). Crucially, their half-life extension in a murine model was 10-fold longer than anti-H5N1 nanobody. In addition, we found that our fenobodies are highly expressed in Escherichia coli, and are both soluble and thermo-stable nanocages that self-assemble as 24-polymers. In conclusion, our results demonstrate that fenobodies have unique advantages over currently available systems for apparent affinity enhancement and half-life extension of nanobodies. Our fenobody system presents a suitable platform for various large-scale biotechnological processes and should greatly facilitate the application of nanobody technology in these areas.

In Vivo Neutralization of α-Cobratoxin with High-Affinity Llama Single-Domain Antibodies (VHHs) and a VHH-Fc Antibody

Science.gov (United States)

Richard, Gabrielle; Meyers, Ashley J.; McLean, Michael D.; Arbabi-Ghahroudi, Mehdi; MacKenzie, Roger; Hall, J. Christopher

2013-01-01

Small recombinant antibody fragments (e.g. scFvs and VHHs), which are highly tissue permeable, are being investigated for antivenom production as conventional antivenoms consisting of IgG or F(ab’)2 antibody fragments do not effectively neutralize venom toxins located in deep tissues. However, antivenoms composed entirely of small antibody fragments may have poor therapeutic efficacy due to their short serum half-lives. To increase serum persistence and maintain tissue penetration, we prepared low and high molecular mass antivenom antibodies. Four llama VHHs were isolated from an immune VHH-displayed phage library and were shown to have high affinity, in the low nM range, for α-cobratoxin (α–Cbtx), the most lethal component of Naja kaouthia venom. Subsequently, our highest affinity VHH (C2) was fused to a human Fc fragment to create a VHH2-Fc antibody that would offer prolonged serum persistence. After in planta (Nicotiana benthamiana) expression and purification, we show that our VHH2-Fc antibody retained high affinity binding to α–Cbtx. Mouse α–Cbtx challenge studies showed that our highest affinity VHHs (C2 and C20) and the VHH2-Fc antibody effectively neutralized lethality induced by α–Cbtx at an antibody:toxin molar ratio as low as ca. 0.75×:1. Further research towards the development of an antivenom therapeutic involving these anti-α-Cbtx VHHs and VHH2-Fc antibody molecules should involve testing them as a combination, to determine whether they maintain tissue penetration capability and low immunogenicity, and whether they exhibit improved serum persistence and therapeutic efficacy. PMID:23894495

Lp-dual affine surface area

Science.gov (United States)

Wei, Wang; Binwu, He

2008-12-01

According to the notion of Lp-affine surface area by Lutwak, in this paper, we introduce the concept of Lp-dual affine surface area. Further, we establish the affine isoperimetric inequality and the Blaschke-Santaló inequality for Lp-dual affine surface area. Besides, the dual Brunn-Minkowski inequality for Lp-dual affine surface area is presented.

Lamprey immune protein-1 (LIP-1) from Lampetra japonica induces cell cycle arrest and cell death in HeLa cells.

Science.gov (United States)

Chi, Xiaoyuan; Su, Peng; Bi, Dan; Tai, Zhao; Li, Yingying; Pang, Yue; Li, Qingwei

2018-04-01

The lamprey (Lampetra japonica), a representative of the jawless vertebrates, is the oldest extant species in the world. LIP-1, which has a jacalin-like domain and an aerolysin pore-forming domain, has previously been identified in Lampetra japonica. However, the structure and function of the LIP-1 protein have not been described. In this study, the LIP-1 gene was overexpressed in HeLa cells and H293T cells. The results showed that the overexpression of LIP-1 in HeLa cells significantly elevated LDH release (P HeLa cells, while it had no effect on H293T cell organelles. Array data indicated that overexpression of LIP-1 primarily upregulated P53 signaling pathways in HeLa cells. Cell cycle assay results confirmed that LIP-1 caused arrest in the G 2 /M phase of the cell cycle in HeLa cells. In summary, our findings provide insights into the function and characterization of LIP-1 genes in vertebrates and establish the foundation for further research into the biological function of LIP-1. Our observations suggest that this lamprey protein has the potential for use in new applications in the medical field. Copyright © 2018. Published by Elsevier Ltd.

Specific recognition of the C-terminal end of A beta 42 by a high affinity monoclonal antibody

DEFF Research Database (Denmark)

Axelsen, Trine Veje; Holm, Arne; Birkelund, Svend

2009-01-01

The neurotoxic peptide A beta(42) is derived from the amyloid precursor protein by proteolytic cleavage and is deposited in the brain of patients suffering from Alzheimer's disease (AD). In this study we generate a high affinity monoclonal antibody that targets the C-terminal end of A beta(42......) with high specificity. By this is meant that the paratope of the antibody must enclose the C-terminal end of A beta(42) including the carboxy-group of amino acid 42, and not just recognize a linear epitope in the C-terminal part of A beta. This has been accomplished by using a unique antigen construct made...... by the Ligand Presenting Assembly technology (LPA technology). This strategy results in dimeric presentation of the free C-terminal end of A beta(42). The generated Mab A beta1.1 is indeed specific for the C-terminal end of A beta(42) to which it binds with high affinity. Mab A beta1.1 recognizes the epitope...

Selection of DNA aptamers against epidermal growth factor receptor with high affinity and specificity.

Science.gov (United States)

Wang, Deng-Liang; Song, Yan-Ling; Zhu, Zhi; Li, Xi-Lan; Zou, Yuan; Yang, Hai-Tao; Wang, Jiang-Jie; Yao, Pei-Sen; Pan, Ru-Jun; Yang, Chaoyong James; Kang, De-Zhi

2014-10-31

Epidermal growth factor receptor (EGFR/HER1/c-ErbB1), is overexpressed in many solid cancers, such as epidermoid carcinomas, malignant gliomas, etc. EGFR plays roles in proliferation, invasion, angiogenesis and metastasis of malignant cancer cells and is the ideal antigen for clinical applications in cancer detection, imaging and therapy. Aptamers, the output of the systematic evolution of ligands by exponential enrichment (SELEX), are DNA/RNA oligonucleotides which can bind protein and other substances with specificity. RNA aptamers are undesirable due to their instability and high cost of production. Conversely, DNA aptamers have aroused researcher's attention because they are easily synthesized, stable, selective, have high binding affinity and are cost-effective to produce. In this study, we have successfully identified DNA aptamers with high binding affinity and selectivity to EGFR. The aptamer named TuTu22 with Kd 56±7.3nM was chosen from the identified DNA aptamers for further study. Flow cytometry analysis results indicated that the TuTu22 aptamer was able to specifically recognize a variety of cancer cells expressing EGFR but did not bind to the EGFR-negative cells. With all of the aforementioned advantages, the DNA aptamers reported here against cancer biomarker EGFR will facilitate the development of novel targeted cancer detection, imaging and therapy. Copyright © 2014 Elsevier Inc. All rights reserved.

Peptides in headlock--a novel high-affinity and versatile peptide-binding nanobody for proteomics and microscopy.

Science.gov (United States)

Braun, Michael B; Traenkle, Bjoern; Koch, Philipp A; Emele, Felix; Weiss, Frederik; Poetz, Oliver; Stehle, Thilo; Rothbauer, Ulrich

2016-01-21

Nanobodies are highly valuable tools for numerous bioanalytical and biotechnical applications. Here, we report the characterization of a nanobody that binds a short peptide epitope with extraordinary affinity. Structural analysis reveals an unusual binding mode where the extended peptide becomes part of a β-sheet structure in the nanobody. This interaction relies on sequence-independent backbone interactions augmented by a small number of specificity-determining side chain contacts. Once bound, the peptide is fastened by two nanobody side chains that clamp it in a headlock fashion. Exploiting this unusual binding mode, we generated a novel nanobody-derived capture and detection system. Matrix-coupled nanobody enables the fast and efficient isolation of epitope-tagged proteins from prokaryotic and eukaryotic expression systems. Additionally, the fluorescently labeled nanobody visualizes subcellular structures in different cellular compartments. The high-affinity-binding and modifiable peptide tag of this system renders it a versatile and robust tool to combine biochemical analysis with microscopic studies.

N-Acetyl-2-Aminofluorene (AAF) Processing in Adult Rat Hepatocytes in Primary Culture Occurs by High-Affinity Low-Velocity and Low-Affinity High-Velocity AAF Metabolite-Forming Systems.

Science.gov (United States)

Koch, Katherine S; Moran, Tom; Shier, W Thomas; Leffert, Hyam L

2018-05-01

N-acetyl-2-aminofluorene (AAF) is a procarcinogen used widely in physiological investigations of chemical hepatocarcinogenesis. Its metabolic pathways have been described extensively, yet little is known about its biochemical processing, growth cycle expression, and pharmacological properties inside living hepatocytes-the principal cellular targets of this hepatocarcinogen. In this report, primary monolayer adult rat hepatocyte cultures and high specific-activity [ring G-3 H]-N-acetyl-2-aminofluorene were used to extend previous observations of metabolic activation of AAF by highly differentiated, proliferation-competent hepatocytes in long-term cultures. AAF metabolism proceeded by zero-order kinetics. Hepatocytes processed significant amounts of procarcinogen (≈12 μg AAF/106 cells/day). Five ring-hydroxylated and one deacetylated species of AAF were secreted into the culture media. Extracellular metabolite levels varied during the growth cycle (days 0-13), but their rank quantitative order was time invariant: 5-OH-AAF > 7-OH-AAF > 3-OH-AAF > N-OH-AAF > aminofluorene (AF) > 1-OH-AAF. Lineweaver-Burk analyses revealed two principal classes of metabolism: System I (high-affinity and low-velocity), Km[APPARENT] = 1.64 × 10-7  M and VMAX[APPARENT] = 0.1 nmol/106 cells/day and System II (low-affinity and high-velocity), Km[APPARENT] = 3.25 × 10-5  M and VMAX[APPARENT] = 1000 nmol/106 cells/day. A third system of metabolism of AAF to AF, with Km[APPARENT] and VMAX[APPARENT] constants of 9.6 × 10-5  M and 4.7 nmol/106 cells/day, was also observed. Evidence provided in this report and its companion paper suggests selective roles and intracellular locations for System I- and System II-mediated AAF metabolite formation during hepatocarcinogenesis, although some of the molecules and mechanisms responsible for multi-system processing remain to be fully defined.

Patterns of invasion and colonization of the sea lamprey (Petromyzon marinus) in North America as revealed by microsatellite genotypes.

Science.gov (United States)

Bryan, M B; Zalinski, D; Filcek, K B; Libants, S; Li, W; Scribner, K T

2005-10-01

Invasions by exotic organisms have had devastating affects on aquatic ecosystems, both ecologically and economically. One striking example of a successful invader that has dramatically affected fish community structure in freshwater lakes of North America is the sea lamprey (Petromyzon marinus). We used eight microsatellite loci and multiple analytical techniques to examine competing hypotheses concerning the origins and colonization history of sea lamprey (n = 741). Analyses were based on replicated invasive populations from Lakes Erie, Huron, Michigan, and Superior, populations of unknown origins from Lakes Ontario, Champlain, and Cayuga, and populations of anadromous putative progenitor populations in North America and Europe. Populations in recently colonized lakes were each established by few colonists through a series of genetic bottlenecks which resulted in lower allelic diversity in more recently established populations. The spatial genetic structure of invasive populations differed from that of native populations on the Atlantic coast, reflecting founder events and connectivity of invaded habitats. Anadromous populations were found to be panmictic (theta(P) = 0.002; 95% CI = -0.003-0.006; P > 0.05). In contrast, there was significant genetic differentiation between populations in the lower and upper Great Lakes (theta(P) = 0.007; P freshwater habitats were examined using coalescent-based analyses, and demonstrated that populations likely originated from natural migrations via the St Lawrence River.

Role of H2O2 on the kinetics of low-affinity high-capacity Na+-dependent alanine transport in SHR proximal tubular epithelial cells

International Nuclear Information System (INIS)

Pinto, Vanda; Pinho, Maria Joao; Jose, Pedro A.; Soares-da-Silva, Patricio

2010-01-01

Research highlights: → H 2 O 2 in excess is required for the presence of a low-affinity high-capacity component for the Na + -dependent [ 14 C]-L-alanine uptake in SHR PTE cells only. → It is suggested that Na + binding in renal ASCT2 may be regulated by ROS in SHR PTE cells. -- Abstract: The presence of high and low sodium affinity states for the Na + -dependent [ 14 C]-L-alanine uptake in immortalized renal proximal tubular epithelial (PTE) cells was previously reported (Am. J. Physiol. 293 (2007) R538-R547). This study evaluated the role of H 2 O 2 on the Na + -dependent [ 14 C]-L-alanine uptake of ASCT2 in immortalized renal PTE cells from Wistar Kyoto rat (WKY) and spontaneously hypertensive rat (SHR). Na + dependence of [ 14 C]-L-alanine uptake was investigated replacing NaCl with an equimolar concentration of choline chloride in vehicle- and apocynin-treated cells. Na + removal from the uptake solution abolished transport activity in both WKY and SHR PTE cells. Decreases in H 2 O 2 levels in the extracellular medium significantly reduced Na + -K m and V max values of the low-affinity high-capacity component in SHR PTE cells, with no effect on the high-affinity low-capacity state of the Na + -dependent [ 14 C]-L-alanine uptake. After removal of apocynin from the culture medium, H 2 O 2 levels returned to basal values within 1 to 3 h in both WKY and SHR PTE cells and these were found stable for the next 24 h. Under these experimental conditions, the Na + -K m and V max of the high-affinity low-capacity state were unaffected and the low-affinity high-capacity component remained significantly decreased 1 day but not 4 days after apocynin removal. In conclusion, H 2 O 2 in excess is required for the presence of a low-affinity high-capacity component for the Na + -dependent [ 14 C]-L-alanine uptake in SHR PTE cells only. It is suggested that Na + binding in renal ASCT2 may be regulated by ROS in SHR PTE cells.

Novel high-affinity and selective biaromatic 4-substituted ¿-hydroxybutyric acid (GHB) analogues as GHB ligands

DEFF Research Database (Denmark)

Høg, Signe; Wellendorph, Petrine; Nielsen, Birgitte

2008-01-01

Gamma-hydroxybutyrate (GHB) is a metabolite of gamma-aminobutyric acid (GABA) and has been proposed to function as a neurotransmitter or neuromodulator. GHB is used in the treatment of narcolepsy and is a drug of abuse. GHB binds to both GABA(B) receptors and specific high-affinity GHB sites...

Are basophil histamine release and high affinity IgE receptor expression involved in asymptomatic skin sensitization?

DEFF Research Database (Denmark)

Jensen, Bettina Margrethe; Assing, K; Jensen, Lone Hummelshøj

2006-01-01

Immunoglobulin (Ig)E-sensitized persons with positive skin prick test, but no allergy symptoms, are classified as being asymptomatic skin sensitized (AS). The allergic type 1 disease is dependant on IgE binding to the high affinity IgE-receptor (FcepsilonRI) expressed on basophils and mast cells....

Structure-guided development of a high-affinity human Programmed Cell Death-1: Implications for tumor immunotherapy

Energy Technology Data Exchange (ETDEWEB)

Lázár-Molnár, Eszter; Scandiuzzi, Lisa; Basu, Indranil; Quinn, Thomas; Sylvestre, Eliezer; Palmieri, Edith; Ramagopal, Udupi A.; Nathenson, Stanley G.; Guha, Chandan; Almo, Steven C.

2017-03-01

Programmed Cell Death-1 (PD-1) is an inhibitory immune receptor, which plays critical roles in T cell co-inhibition and exhaustion upon binding to its ligands PD-L1 and PD-L2. We report the crystal structure of the human PD-1 ectodomain and the mapping of the PD-1 binding interface. Mutagenesis studies confirmed the crystallographic interface, and resulted in mutant PD-1 receptors with altered affinity and ligand-specificity. In particular, a high-affinity mutant PD-1 (HA PD-1) exhibited 45 and 30-fold increase in binding to PD-L1 and PD-L2, respectively, due to slower dissociation rates. This mutant (A132L) was used to engineer a soluble chimeric Ig fusion protein for cell-based and in vivo studies. HA PD-1 Ig showed enhanced binding to human dendritic cells, and increased T cell proliferation and cytokine production in a mixed lymphocyte reaction (MLR) assay. Moreover, in an experimental model of murine Lewis lung carcinoma, HA PD-1 Ig treatment synergized with radiation therapy to decrease local and metastatic tumor burden, as well as in the establishment of immunological memory responses. Our studies highlight the value of structural considerations in guiding the design of a high-affinity chimeric PD-1 Ig fusion protein with robust immune modulatory properties, and underscore the power of combination therapies to selectively manipulate the PD-1 pathway for tumor immunotherapy.

Structure-guided development of a high-affinity human Programmed Cell Death-1: Implications for tumor immunotherapy

Directory of Open Access Journals (Sweden)

Eszter Lázár-Molnár

2017-03-01

Full Text Available Programmed Cell Death-1 (PD-1 is an inhibitory immune receptor, which plays critical roles in T cell co-inhibition and exhaustion upon binding to its ligands PD-L1 and PD-L2. We report the crystal structure of the human PD-1 ectodomain and the mapping of the PD-1 binding interface. Mutagenesis studies confirmed the crystallographic interface, and resulted in mutant PD-1 receptors with altered affinity and ligand-specificity. In particular, a high-affinity mutant PD-1 (HA PD-1 exhibited 45 and 30-fold increase in binding to PD-L1 and PD-L2, respectively, due to slower dissociation rates. This mutant (A132L was used to engineer a soluble chimeric Ig fusion protein for cell-based and in vivo studies. HA PD-1 Ig showed enhanced binding to human dendritic cells, and increased T cell proliferation and cytokine production in a mixed lymphocyte reaction (MLR assay. Moreover, in an experimental model of murine Lewis lung carcinoma, HA PD-1 Ig treatment synergized with radiation therapy to decrease local and metastatic tumor burden, as well as in the establishment of immunological memory responses. Our studies highlight the value of structural considerations in guiding the design of a high-affinity chimeric PD-1 Ig fusion protein with robust immune modulatory properties, and underscore the power of combination therapies to selectively manipulate the PD-1 pathway for tumor immunotherapy.

Mapping of barley alpha-amylases and outer subsite mutants reveals dynamic high-affinity subsites and barriers in the long substrate binding cleft

DEFF Research Database (Denmark)

Kandra, L.; Abou Hachem, Maher; Gyemant, G.

2006-01-01

Subsite affinity maps of long substrate binding clefts in barley alpha-amylases, obtained using a series of maltooligosaccharides of degree of polymerization of 3-12, revealed unfavorable binding energies at the internal subsites -3 and -5 and at subsites -8 and +3/+4 defining these subsites...... as binding barriers. Barley a-amylase I mutants Y105A and T212Y at subsite -6 and +4 resulted in release or anchoring of bound substrate, thus modifying the affinities of other high-affinity subsites (-2 and +2) and barriers. The double mutant Y105A-T212Y displayed a hybrid subsite affinity profile......, converting barriers to binding areas. These findings highlight the dynamic binding energy distribution and the versatility of long maltooligosaccharide derivatives in mapping extended binding clefts in a-amylases....

Characterization of glucagon-like peptide-1 receptor beta-arrestin 2 interaction: a high-affinity receptor phenotype

DEFF Research Database (Denmark)

Jorgensen, Rasmus; Martini, Lene; Schwartz, Thue W

2005-01-01

To dissect the interaction between beta-arrestin ((beta)arr) and family B G protein-coupled receptors, we constructed fusion proteins between the glucagon-like peptide 1 receptor and (beta)arr2. The fusion constructs had an increase in apparent affinity selectively for glucagon, suggesting...... that (beta)arr2 interaction locks the receptor in a high-affinity conformation, which can be explored by some, but not all, ligands. The fusion constructs adopted a signaling phenotype governed by the tethered (beta)arr2 with an attenuated G protein-mediated cAMP signal and a higher maximal internalization...... of that which has previously been characterized for family A G protein-coupled receptors, suggesting similarities in the effect of (beta)arr interaction between family A and B receptors also at the molecular level....

An in vitro-identified high-affinity nucleosome-positioning signal is capable of transiently positioning a nucleosome in vivo

Directory of Open Access Journals (Sweden)

Gracey Lia E

2010-07-01

Full Text Available Abstract Background The physiological function of eukaryotic DNA occurs in the context of nucleosomal arrays that can expose or obscure defined segments of the genome. Certain DNA sequences are capable of strongly positioning a nucleosome in vitro, suggesting the possibility that favorable intrinsic signals might reproducibly structure chromatin segments. As high-throughput sequencing analyses of nucleosome coverage in vitro and in vivo have become possible, a vigorous debate has arisen over the degree to which intrinsic DNA:nucleosome affinities orchestrate the in vivo positions of nucleosomes, thereby controlling physical accessibility of specific sequences in DNA. Results We describe here the in vivo consequences of placing a synthetic high-affinity nucleosome-positioning signal, the 601 sequence, into a DNA plasmid vector in mice. Strikingly, the 601 sequence was sufficient to position nucleosomes during an early phase after introduction of the DNA into the mice (when the plasmid vector transgene was active. This positioning capability was transient, with a loss of strong positioning at a later time point when the transgenes had become silent. Conclusions These results demonstrate an ability of DNA sequences selected solely for nucleosome affinity to organize chromatin in vivo, and the ability of other mechanisms to overcome these interactions in a dynamic nuclear environment.

High-affinity binding of two molecules of cysteine proteinases to low-molecular-weight kininogen.

Science.gov (United States)

Turk, B.; Stoka, V.; Björk, I.; Boudier, C.; Johansson, G.; Dolenc, I.; Colic, A.; Bieth, J. G.; Turk, V.

1995-01-01

Human low-molecular-weight kininogen (LK) was shown by fluorescence titration to bind two molecules of cathepsins L and S and papain with high affinity. By contrast, binding of a second molecule of cathepsin H was much weaker. The 2:1 binding stoichiometry was confirmed by titration monitored by loss of enzyme activity and by sedimentation velocity experiments. The kinetics of binding of cathepsins L and S and papain showed the two proteinase binding sites to have association rate constants kass,1 = 10.7-24.5 x 10(6) M-1 s-1 and kass,2 = 0.83-1.4 x 10(6) M-1 s-1. Comparison of these kinetic constants with previous data for intact LK and its separated domains indicate that the faster-binding site is also the tighter-binding site and is present on domain 3, whereas the slower-binding, lower-affinity site is on domain 2. These results also indicate that there is no appreciable steric hindrance for the binding of proteinases between the two binding sites or from the kininogen light chain. PMID:8528085

Continuous affine processes

DEFF Research Database (Denmark)

Buchardt, Kristian

2016-01-01

Affine processes possess the property that expectations of exponential affine transformations are given by a set of Riccati differential equations, which is the main feature of this popular class of processes. In this paper we generalise these results for expectations of more general transformati...

Affinity in electrophoresis.

Science.gov (United States)

Heegaard, Niels H H

2009-06-01

The journal Electrophoresis has greatly influenced my approaches to biomolecular affinity studies. The methods that I have chosen as my main tools to study interacting biomolecules--native gel and later capillary zone electrophoresis--have been the topic of numerous articles in Electrophoresis. Below, the role of the journal in the development and dissemination of these techniques and applications reviewed. Many exhaustive reviews on affinity electrophoresis and affinity CE have been published in the last few years and are not in any way replaced by the present deliberations that are focused on papers published by the journal.

Primary structure and conformational analysis of peptide methionine-tyrosine, a peptide related to neuropeptide Y and peptide YY isolated from lamprey intestine

DEFF Research Database (Denmark)

Conlon, J M; Bjørnholm, B; Jørgensen, Flemming Steen

1991-01-01

A peptide belonging to the pancreatic-polypeptide-fold family of regulatory peptides has been isolated from the intestine of an Agnathan, the sea lamprey (Petromyzon marinus). The primary structure of the peptide (termed peptide methionine-tyrosine) was established as Met-Pro-Pro-Lys-Pro-Asp-Asn-...... in a preferred structure in which the conformation of the beta-turn between the two helical domains (residues 9-14) is appreciably different....

Detection-Guided Fast Affine Projection Channel Estimator for Speech Applications

Directory of Open Access Journals (Sweden)

Yan Wu Jennifer

2007-04-01

Full Text Available In various adaptive estimation applications, such as acoustic echo cancellation within teleconferencing systems, the input signal is a highly correlated speech. This, in general, leads to extremely slow convergence of the NLMS adaptive FIR estimator. As a result, for such applications, the affine projection algorithm (APA or the low-complexity version, the fast affine projection (FAP algorithm, is commonly employed instead of the NLMS algorithm. In such applications, the signal propagation channel may have a relatively low-dimensional impulse response structure, that is, the number m of active or significant taps within the (discrete-time modelled channel impulse response is much less than the overall tap length n of the channel impulse response. For such cases, we investigate the inclusion of an active-parameter detection-guided concept within the fast affine projection FIR channel estimator. Simulation results indicate that the proposed detection-guided fast affine projection channel estimator has improved convergence speed and has lead to better steady-state performance than the standard fast affine projection channel estimator, especially in the important case of highly correlated speech input signals.

Peptides in headlock – a novel high-affinity and versatile peptide-binding nanobody for proteomics and microscopy

Science.gov (United States)

Braun, Michael B.; Traenkle, Bjoern; Koch, Philipp A.; Emele, Felix; Weiss, Frederik; Poetz, Oliver; Stehle, Thilo; Rothbauer, Ulrich

2016-01-01

Nanobodies are highly valuable tools for numerous bioanalytical and biotechnical applications. Here, we report the characterization of a nanobody that binds a short peptide epitope with extraordinary affinity. Structural analysis reveals an unusual binding mode where the extended peptide becomes part of a β-sheet structure in the nanobody. This interaction relies on sequence-independent backbone interactions augmented by a small number of specificity-determining side chain contacts. Once bound, the peptide is fastened by two nanobody side chains that clamp it in a headlock fashion. Exploiting this unusual binding mode, we generated a novel nanobody-derived capture and detection system. Matrix-coupled nanobody enables the fast and efficient isolation of epitope-tagged proteins from prokaryotic and eukaryotic expression systems. Additionally, the fluorescently labeled nanobody visualizes subcellular structures in different cellular compartments. The high-affinity-binding and modifiable peptide tag of this system renders it a versatile and robust tool to combine biochemical analysis with microscopic studies. PMID:26791954

The metric-affine gravitational theory as the gauge theory of the affine group

International Nuclear Information System (INIS)

Lord, E.A.

1978-01-01

The metric-affine gravitational theory is shown to be the gauge theory of the affine group, or equivalently, the gauge theory of the group GL(4,R) of tetrad deformations in a space-time with a locally Minkowskian metric. The identities of the metric-affine theory, and the relationship between them and those of general relativity and Sciama-Kibble theory, are derived. (Auth.)

Cyclic GMP-AMP Containing Mixed Phosphodiester Linkages Is An Endogenous High Affinity Ligand for STING

Science.gov (United States)

Zhang, Xu; Shi, Heping; Wu, Jiaxi; Zhang, Xuewu; Sun, Lijun; Chen, Chuo; Chen, Zhijian J.

2013-01-01

The presence of microbial or self DNA in the cytoplasm of mammalian cells is a danger signal detected by the DNA sensor cyclic-GMP-AMP (cGAMP) synthase (cGAS), which catalyzes the production of cGAMP that in turn serves as a second messenger to activate innate immune responses. Here we show that endogenous cGAMP in mammalian cells contains two distinct phosphodiester linkages, one between 2′-OH of GMP and 5′-phosphate of AMP, and the other between 3′-OH of AMP and 5′-phosphate of GMP. This molecule, termed 2′3′-cGAMP, is unique in that it binds to the adaptor protein STING with a much greater affinity than cGAMP molecules containing other combinations of phosphodiester linkages. The crystal structure of STING bound to 2′3′-cGAMP revealed the structural basis of this high-affinity binding and a ligand-induced conformational change in STING that may underlie its activation. PMID:23747010

Cyclic GMP-AMP containing mixed phosphodiester linkages is an endogenous high-affinity ligand for STING.

Science.gov (United States)

Zhang, Xu; Shi, Heping; Wu, Jiaxi; Zhang, Xuewu; Sun, Lijun; Chen, Chuo; Chen, Zhijian J

2013-07-25

The presence of microbial or self DNA in the cytoplasm of mammalian cells is a danger signal detected by the DNA sensor cyclic-GMP-AMP (cGAMP) synthase (cGAS), which catalyzes the production of cGAMP that in turn serves as a second messenger to activate innate immune responses. Here we show that endogenous cGAMP in mammalian cells contains two distinct phosphodiester linkages, one between 2'-OH of GMP and 5'-phosphate of AMP, and the other between 3'-OH of AMP and 5'-phosphate of GMP. This molecule, termed 2'3'-cGAMP, is unique in that it binds to the adaptor protein STING with a much greater affinity than cGAMP molecules containing other combinations of phosphodiester linkages. The crystal structure of STING bound to 2'3'-cGAMP revealed the structural basis of this high-affinity binding and a ligand-induced conformational change in STING that may underlie its activation. Copyright © 2013 Elsevier Inc. All rights reserved.

Solution structure of the Grb2 SH2 domain complexed with a high-affinity inhibitor

International Nuclear Information System (INIS)

Ogura, Kenji; Shiga, Takanori; Yokochi, Masashi; Yuzawa, Satoru; Burke, Terrence R.; Inagaki, Fuyuhiko

2008-01-01

The solution structure of the growth factor receptor-bound protein 2 (Grb2) SH2 domain complexed with a high-affinity inhibitor containing a non-phosphorus phosphate mimetic within a macrocyclic platform was determined by nuclear magnetic resonance (NMR) spectroscopy. Unambiguous assignments of the bound inhibitor and intermolecular NOEs between the Grb2 SH2 domain and the inhibitor was accomplished using perdeuterated Grb2 SH2 protein. The well-defined solution structure of the complex was obtained and compared to those by X-ray crystallography. Since the crystal structure of the Grb2 SH2 domain formed a domain-swapped dimer and several inhibitors were bound to a hinge region, there were appreciable differences between the solution and crystal structures. Based on the binding interactions between the inhibitor and the Grb2 SH2 domain in solution, we proposed a design of second-generation inhibitors that could be expected to have higher affinity

The ketamine analogue methoxetamine and 3- and 4-methoxy analogues of phencyclidine are high affinity and selective ligands for the glutamate NMDA receptor.

Directory of Open Access Journals (Sweden)

Bryan L Roth

Full Text Available In this paper we determined the pharmacological profiles of novel ketamine and phencyclidine analogues currently used as 'designer drugs' and compared them to the parent substances via the resources of the National Institute of Mental Health Psychoactive Drug Screening Program. The ketamine analogues methoxetamine ((RS-2-(ethylamino-2-(3-methoxyphenylcyclohexanone and 3-MeO-PCE (N-ethyl-1-(3-methoxyphenylcyclohexanamine and the 3- and 4-methoxy analogues of phencyclidine, (1-[1-(3-methoxyphenylcyclohexyl]piperidine and 1-[1-(4-methoxyphenylcyclohexyl]piperidine, were all high affinity ligands for the PCP-site on the glutamate NMDA receptor. In addition methoxetamine and PCP and its analogues displayed appreciable affinities for the serotonin transporter, whilst the PCP analogues exhibited high affinities for sigma receptors. Antagonism of the NMDA receptor is thought to be the key pharmacological feature underlying the actions of dissociative anaesthetics. The novel ketamine and PCP analogues had significant affinities for the NMDA receptor in radioligand binding assays, which may explain their psychotomimetic effects in human users. Additional actions on other targets could be important for delineating side-effects.

Molecular cloning of a second subunit of the receptor for human granulocyte - macrophage colony-stimulating factor (GM-CSF): Reconstitution of a high-affinity GM-CSF receptor

International Nuclear Information System (INIS)

Hayashida, Kazuhiro; Kitamura, Toshio; Gorman, D.M.; Miyajima, Atsushi; Arai, Kenichi; Yokota, Takashi

1990-01-01

Using the mouse interleukin 3 (IL-3) receptor cDNA as a probe, the authors obtained a monologous cDNA (KH97) from a cDNA library of a human hemopoietic cell line, TF-1. The protein encoded by the KH97 cDNA has 56% amino acid sequence identity with the mouse IL-3 receptor and retains features common to the family of cytokine receptors. Fibroblasts transfected with the KH97 cDNA expressed a protein of 120 kDa but did not bind any human cytokines, including IL-3 and granulocyte - macrophage colony-stimulating factor (GM-CSF). Interestingly, cotransfection of cDNAs for KH97 and the low-affinity human GM-CSF receptor in fibroblasts resulted in formation of a high-affinity receptor for GM-CSF. The dissociation rate of GM-CSF from the reconstituted high-affinity receptor was slower than that from the low-affinity site, whereas the association rate was unchanged. Cross-linking of 125 I-labeled GM-CSF to fibroblasts cotransfected with both cDNAs revealed the same cross-linking patterns as in TF-1 cells - i.e., two major proteins of 80 and 120 kDa which correspond to the low-affinity GM-CSF receptor and the KH97 protein, respectively. These results indicate that the high-affinity GM-CSF receptor is composed of at least two components in a manner analogous to the IL-2 receptor. They therefore propose to designate the low-affinity GM-CSF receptor and the KH97 protein as the α and β subunits of the GM-CSF receptor, respectively

A rhodamine-labeled citalopram analogue as a high-affinity fluorescent probe for the serotonin transporter

DEFF Research Database (Denmark)

Zhang, Peng; Jørgensen, Trine Nygaard; Løland, Claus Juul

2013-01-01

A novel fluorescent ligand was synthesized as a high-affinity, high specificity probe for visualizing the serotonin transporter (SERT). The rhodamine fluorophore was extended from an aniline substitution on the 5-position of the dihydroisobenzofuran ring of citalopram (2, 1-(3-(dimethylamino......)propyl)-1-(4-fluorophenyl)-1,3-dihydroisobenzofuran-5-carbonitrile), using an ethylamino linker. The resulting rhodamine-labeled ligand 8 inhibited [3H]5-HT uptake in COS-7 cells (Ki = 225 nM) with similar potency to the tropane-based JHC 1-064 (1), but with higher specificity towards the SERT relative...

Characterization of the neurohypophysial hormone gene loci in elephant shark and the Japanese lamprey: origin of the vertebrate neurohypophysial hormone genes

Directory of Open Access Journals (Sweden)

Brenner Sydney

2009-02-01

Full Text Available Abstract Background Vasopressin and oxytocin are mammalian neurohypophysial hormones with distinct functions. Vasopressin is involved mainly in osmoregulation and oxytocin is involved primarily in parturition and lactation. Jawed vertebrates contain at least one homolog each of vasopressin and oxytocin, whereas only a vasopressin-family hormone, vasotocin, has been identified in jawless vertebrates. The genes encoding vasopressin and oxytocin are closely linked tail-to-tail in eutherian mammals whereas their homologs in chicken, Xenopus and coelacanth (vasotocin and mesotocin are linked tail-to-head. In contrast, their pufferfish homologs, vasotocin and isotocin, are located on the same strand of DNA with isotocin located upstream of vasotocin and separated by five genes. These differences in the arrangement of the two genes in different bony vertebrate lineages raise questions about their origin and ancestral arrangement. To trace the origin of these genes, we have sequenced BAC clones from the neurohypophysial gene loci in a cartilaginous fish, the elephant shark (Callorhinchus milii, and in a jawless vertebrate, the Japanese lamprey (Lethenteron japonicum. We have also analyzed the neurohypophysial hormone gene locus in an invertebrate chordate, the amphioxus (Branchiostoma floridae. Results The elephant shark neurohypophysial hormone genes encode vasotocin and oxytocin, and are linked tail-to-head like their homologs in coelacanth and non-eutherian tetrapods. Besides the hypothalamus, the two genes are also expressed in the ovary. In addition, the vasotocin gene is expressed in the kidney, rectal gland and intestine. These expression profiles indicate a paracrine role for the two hormones. The lamprey locus contains a single neurohypophysial hormone gene, the vasotocin. The synteny of genes in the lamprey locus is conserved in elephant shark, coelacanth and tetrapods but disrupted in teleost fishes. The amphioxus locus encodes a single

Influence of a weak field of pulsed DC electricity on the behavior and incidence of injury in adult Steelhead and Pacific Lamprey

Science.gov (United States)

Mesa, Matthew G.; Copeland, Elizabeth S.

2009-01-01

Predation by pinnipeds, such as California sea lions Zalophus californianus, Pacific harbor seals Phoca vitulina, and Stellar sea lions Eumetopias jubatus on adult Pacific salmon Oncorhynchus spp in the lower Columbia River has become a serious concern for fishery managers trying to conserve and restore runs of threatened and endangered fish. As a result, Smith-Root, Incorporated (SRI; Vancouver, Washington), manufacturers of electrofishing and closely-related equipment, proposed a project to evaluate the potential of an electrical barrier to deter marine mammals and reduce the amount of predation on adult salmonids (SRI 2007). The objectives of their work were to develop, deploy, and evaluate a passive, integrated sonar and electric barrier that would selectively inhibit the upstream movements of marine mammals and reduce predation, but would not injure pinnipeds or impact anadromous fish migrations. However, before such a device could be deployed in the field, concerns by regional fishery managers about the potential effects of such a device on the migratory behavior of Pacific salmon, steelhead O. mykiss, Pacific lampreys Entoshpenus tridentata, and white sturgeon Acipenser transmontanus, needed to be addressed. In this report, we describe the results of laboratory research designed to evaluate the effects of prototype electric barriers on adult steelhead and Pacific lampreys.

Novel and high affinity fluorescent ligands for the serotonin transporter based on (s)-citalopram

DEFF Research Database (Denmark)

Kumar, Vivek; Rahbek-Clemmensen, Troels; Billesbølle, Christian B

2014-01-01

Novel rhodamine-labeled ligands, based on (S)-citalopram, were synthesized and evaluated for uptake inhibition at the human serotonin, dopamine, and norepinephrine transporters (hSERT, hDAT, and hNET, respectively) and for binding at SERT, in transiently transfected COS7 cells. Compound 14 demons...... demonstrated high affinity binding and selectivity for SERT (K i = 3 nM). Visualization of SERT, using confocal laser scanning microscopy, validated compound 14 as a novel tool for studying SERT expression and distribution in living cells....

A Generalized Affine Isoperimetric Inequality

OpenAIRE

Chen, Wenxiong; Howard, Ralph; Lutwak, Erwin; Yang, Deane; Zhang, Gaoyong

2004-01-01

A purely analytic proof is given for an inequality that has as a direct consequence the two most important affine isoperimetric inequalities of plane convex geometry: The Blaschke-Santalo inequality and the affine isoperimetric inequality of affine differential geometry.

Generation of high-affinity, internalizing anti-FGFR2 single-chain variable antibody fragment fused with Fc for targeting gastrointestinal cancers.

Science.gov (United States)

Borek, Aleksandra; Sokolowska-Wedzina, Aleksandra; Chodaczek, Grzegorz; Otlewski, Jacek

2018-01-01

Fibroblast growth factor receptors (FGFRs) are promising targets for antibody-based cancer therapies, as their substantial overexpression has been found in various tumor cells. Aberrant activation of FGF receptor 2 (FGFR2) signaling through overexpression of FGFR2 and/or its ligands, mutations, or receptor amplification has been reported in multiple cancer types, including gastric, colorectal, endometrial, ovarian, breast and lung cancer. In this paper, we describe application of the phage display technology to produce a panel of high affinity single chain variable antibody fragments (scFvs) against the extracellular ligand-binding domain of FGFR2 (ECD_FGFR2). The binders were selected from the human single chain variable fragment scFv phage display libraries Tomlinson I + J and showed high specificity and binding affinity towards human FGFR2 with nanomolar KD values. To improve the affinity of the best binder selected, scFvF7, we reformatted it to a bivalent diabody format, or fused it with the Fc region (scFvF7-Fc). The scFvF7-Fc antibody construct presented the highest affinity for FGFR2, with a KD of 0.76 nM, and was selectively internalized into cancer cells overexpressing FGFR2, Snu-16 and NCI-H716. Finally, we prepared a conjugate of scFvF7-Fc with the cytotoxic drug monomethyl-auristatin E (MMAE) and evaluated its cytotoxicity. The conjugate delivered MMAE selectively to FGFR2-positive tumor cells. These results indicate that scFvF7-Fc-vcMMAE is a highly potent molecule for the treatment of cancers with FGFR2 overexpression.

Compound immobilization and drug-affinity chromatography.

Science.gov (United States)

Rix, Uwe; Gridling, Manuela; Superti-Furga, Giulio

2012-01-01

Bioactive small molecules act through modulating a yet unpredictable number of targets. It is therefore of critical importance to define the cellular target proteins of a compound as an entry point to understanding its mechanism of action. Often, this can be achieved in a direct fashion by chemical proteomics. As with any affinity chromatography, immobilization of the bait to a solid support is one of the earliest and most crucial steps in the process. Interfering with structural features that are important for identification of a target protein will be detrimental to binding affinity. Also, many molecules are sensitive to heat or to certain chemicals, such as acid or base, and might be destroyed during the process of immobilization, which therefore needs to be not only efficient, but also mild. The subsequent affinity chromatography step needs to preserve molecular and conformational integrity of both bait compound and proteins in order to result in the desired specific enrichment while ensuring a high level of compatibility with downstream analysis by mass spectrometry. Thus, the right choice of detergent, buffer, and protease inhibitors is also essential. This chapter describes a widely applicable procedure for the immobilization of small molecule drugs and for drug-affinity chromatography with subsequent protein identification by mass spectrometry.

Determination of High-affinity Antibody-antigen Binding Kinetics Using Four Biosensor Platforms.

Science.gov (United States)

Yang, Danlin; Singh, Ajit; Wu, Helen; Kroe-Barrett, Rachel

2017-04-17

Label-free optical biosensors are powerful tools in drug discovery for the characterization of biomolecular interactions. In this study, we describe the use of four routinely used biosensor platforms in our laboratory to evaluate the binding affinity and kinetics of ten high-affinity monoclonal antibodies (mAbs) against human proprotein convertase subtilisin kexin type 9 (PCSK9). While both Biacore T100 and ProteOn XPR36 are derived from the well-established Surface Plasmon Resonance (SPR) technology, the former has four flow cells connected by serial flow configuration, whereas the latter presents 36 reaction spots in parallel through an improvised 6 x 6 crisscross microfluidic channel configuration. The IBIS MX96 also operates based on the SPR sensor technology, with an additional imaging feature that provides detection in spatial orientation. This detection technique coupled with the Continuous Flow Microspotter (CFM) expands the throughput significantly by enabling multiplex array printing and detection of 96 reaction sports simultaneously. In contrast, the Octet RED384 is based on the BioLayer Interferometry (BLI) optical principle, with fiber-optic probes acting as the biosensor to detect interference pattern changes upon binding interactions at the tip surface. Unlike the SPR-based platforms, the BLI system does not rely on continuous flow fluidics; instead, the sensor tips collect readings while they are immersed in analyte solutions of a 384-well microplate during orbital agitation. Each of these biosensor platforms has its own advantages and disadvantages. To provide a direct comparison of these instruments' ability to provide quality kinetic data, the described protocols illustrate experiments that use the same assay format and the same high-quality reagents to characterize antibody-antigen kinetics that fit the simple 1:1 molecular interaction model.

Lp-mixed affine surface area

Science.gov (United States)

Wang, Weidong; Leng, Gangsong

2007-11-01

According to the three notions of mixed affine surface area, Lp-affine surface area and Lp-mixed affine surface area proposed by Lutwak, in this article, we give the concept of ith Lp-mixed affine surface area such that the first and second notions of Lutwak are its special cases. Further, some Lutwak's results are extended associated with this concept. Besides, applying this concept, we establish an inequality for the volumes and dual quermassintegrals of a class of star bodies.

Content-Based High-Resolution Remote Sensing Image Retrieval via Unsupervised Feature Learning and Collaborative Affinity Metric Fusion

Directory of Open Access Journals (Sweden)

Yansheng Li

2016-08-01

Full Text Available With the urgent demand for automatic management of large numbers of high-resolution remote sensing images, content-based high-resolution remote sensing image retrieval (CB-HRRS-IR has attracted much research interest. Accordingly, this paper proposes a novel high-resolution remote sensing image retrieval approach via multiple feature representation and collaborative affinity metric fusion (IRMFRCAMF. In IRMFRCAMF, we design four unsupervised convolutional neural networks with different layers to generate four types of unsupervised features from the fine level to the coarse level. In addition to these four types of unsupervised features, we also implement four traditional feature descriptors, including local binary pattern (LBP, gray level co-occurrence (GLCM, maximal response 8 (MR8, and scale-invariant feature transform (SIFT. In order to fully incorporate the complementary information among multiple features of one image and the mutual information across auxiliary images in the image dataset, this paper advocates collaborative affinity metric fusion to measure the similarity between images. The performance evaluation of high-resolution remote sensing image retrieval is implemented on two public datasets, the UC Merced (UCM dataset and the Wuhan University (WH dataset. Large numbers of experiments show that our proposed IRMFRCAMF can significantly outperform the state-of-the-art approaches.

Quantitative relationship between antibody affinity and antibody avidity

International Nuclear Information System (INIS)

Griswold, W.R.

1987-01-01

The relationship between antibody avidity, measured by the dissociation of the antigen-antibody bond in antigen excess, and antibody affinity was studied. Complexes of radiolabelled antigen and antibody of known affinity were prepared in vitro and allowed to stand for seven days to reach equilibrium. Then nonlabelled antigen in one hundred fold excess was added to dissociate the complexes. After an appropriate incubation the fraction of antigen bound to antibody was measured by the ammonium sulfate precipitation method. The dissociation index was the fraction bound in the experimental sample divided by the fraction bound in the control. The correlation coefficient between the dissociation index and the antibody binding constant was 0.92 for early dissociation and 0.98 for late dissociation. The regression equation relating the binding constant to the dissociation index was K = 6.4(DI) + 6.25, where DI is the late dissociation index and K is the logarithm to the base 10 of the binding constant. There is a high correlation between avidity and affinity of antibody. Antibody affinity can be estimated from avidity data. The stability of antigen-antibody complexes can be predicted from antibody affinity

Novel trends in affinity biosensors: current challenges and perspectives

International Nuclear Information System (INIS)

Arugula, Mary A; Simonian, Aleksandr

2014-01-01

Molecular biorecognition processes facilitate physical and biochemical interactions between molecules in all crucial metabolic pathways. Perhaps the target analyte and the biorecognition element interactions have the most impactful use in biosensing applications. Traditional analytical sensing systems offer excellent biorecognition elements with the ability to detect and determine the presence of analytes. High affinity antibodies and DNA play an important role in the development of affinity biosensors based on electrochemical, optical and mass sensitive approaches. Advancements in this area routinely employ labels, label free, nanoparticles, multifunctional matrices, carbon nanotubes and other methods to meet the requirements of its own application. However, despite increasing affinity ceilings for conventional biosensors, the field draws back in meeting specifically important demands, such as long-term stability, ultrasensitivity, rapid detection, extreme selectivity, strong biological base, calibration, in vivo measurements, regeneration, satisfactory performance and ease of production. Nevertheless, recent efforts through this line have produced novel high-tech nanosensing systems such as ‘aptamers’ and ‘phages’ which exhibit high-throughput sensing. Aptamers and phages are powerful tools that excel over antibodies in sensibility, stability, multi-detection, in vivo measurements and regeneration. Phages are superior in stability, screening for affinity-based target molecules ranging from small to proteins and even cells, and easy production. In this review, we focus mainly on recent developments in affinity-based biosensors such as immunosensors, DNA sensors, emphasizing aptasensors and phage-based biosensors basing on novel electrochemical, optical and mass sensitive detection techniques. We also address enzyme inhibition-based biosensors and the current problems associated with the above sensors and their future perspectives. (topical review)

Specificity and affinity quantification of flexible recognition from underlying energy landscape topography.

Directory of Open Access Journals (Sweden)

Xiakun Chu

2014-08-01

Full Text Available Flexibility in biomolecular recognition is essential and critical for many cellular activities. Flexible recognition often leads to moderate affinity but high specificity, in contradiction with the conventional wisdom that high affinity and high specificity are coupled. Furthermore, quantitative understanding of the role of flexibility in biomolecular recognition is still challenging. Here, we meet the challenge by quantifying the intrinsic biomolecular recognition energy landscapes with and without flexibility through the underlying density of states. We quantified the thermodynamic intrinsic specificity by the topography of the intrinsic binding energy landscape and the kinetic specificity by association rate. We found that the thermodynamic and kinetic specificity are strongly correlated. Furthermore, we found that flexibility decreases binding affinity on one hand, but increases binding specificity on the other hand, and the decreasing or increasing proportion of affinity and specificity are strongly correlated with the degree of flexibility. This shows more (less flexibility leads to weaker (stronger coupling between affinity and specificity. Our work provides a theoretical foundation and quantitative explanation of the previous qualitative studies on the relationship among flexibility, affinity and specificity. In addition, we found that the folding energy landscapes are more funneled with binding, indicating that binding helps folding during the recognition. Finally, we demonstrated that the whole binding-folding energy landscapes can be integrated by the rigid binding and isolated folding energy landscapes under weak flexibility. Our results provide a novel way to quantify the affinity and specificity in flexible biomolecular recognition.

High-throughput and multiplexed regeneration buffer scouting for affinity-based interactions

NARCIS (Netherlands)

Geuijen, K.P.M.; Schasfoort, R.B.; Wijffels, R.H.; Eppink, M.H.M.

2014-01-01

Affinity-based analyses on biosensors depend partly on regeneration between measurements. Regeneration is performed with a buffer that efficiently breaks all interactions between ligand and analyte while maintaining the active binding site of the ligand. We demonstrated a regeneration buffer

A potential therapy for chordoma via antibody-dependent cell-mediated cytotoxicity employing NK or high-affinity NK cells in combination with cetuximab.

Science.gov (United States)

Fujii, Rika; Schlom, Jeffrey; Hodge, James W

2018-05-01

OBJECTIVE Chordoma is a rare bone tumor derived from the notochord and is resistant to conventional therapies such as chemotherapy, radiotherapy, and targeting therapeutics. Expression of epidermal growth factor receptor (EGFR) in a large proportion of chordoma specimens indicates a potential target for therapeutic intervention. In this study the authors investigated the potential role of the anti-EGFR antibody cetuximab in immunotherapy for chordoma. METHODS Since cetuximab is a monoclonal antibody of the IgG1 isotype, it has the potential to mediate antibody-dependent cell-mediated cytotoxicity (ADCC) employing natural killer (NK) cells as effectors. Polymorphisms in the CD16 allele expressed on NK cells have been shown to influence the degree of ADCC of tumor cells, with the high-affinity valine (V)/V allele being responsible for more lysis than the V/phenylalanine (F) or FF allele. Unfortunately, however, only approximately 10% of the population expresses the VV allele on NK cells. An NK cell line, NK-92, has now been engineered to endogenously express IL-2 and the high-affinity CD16 allele. These irradiated high-affinity (ha)NK cells were analyzed for lysis of chordoma cells with and without cetuximab, and the levels of lysis observed in ADCC were compared with those of NK cells from donors expressing the VV, VF, and FF alleles. RESULTS Here the authors demonstrate for the first time 1) that cetuximab in combination with NK cells can mediate ADCC of chordoma cells; 2) the influence of the NK CD16 polymorphism in cetuximab-mediated ADCC for chordoma cell lysis; 3) that engineered haNK cells-that is, cells transduced to express the CD16 V158 FcγRIIIa receptor-bind cetuximab with similar affinity to normal NK cells expressing the high-affinity VV allele; and 4) that irradiated haNK cells induce ADCC with cetuximab in chordoma cells. CONCLUSIONS These studies provide rationale for the use of cetuximab in combination with irradiated haNK cells for therapy for

High-affinity DNA-binding Domains of Replication Protein A (RPA) Direct SMARCAL1-dependent Replication Fork Remodeling*

Science.gov (United States)

Bhat, Kamakoti P.; Bétous, Rémy; Cortez, David

2015-01-01

SMARCAL1 catalyzes replication fork remodeling to maintain genome stability. It is recruited to replication forks via an interaction with replication protein A (RPA), the major ssDNA-binding protein in eukaryotic cells. In addition to directing its localization, RPA also activates SMARCAL1 on some fork substrates but inhibits it on others, thereby conferring substrate specificity to SMARCAL1 fork-remodeling reactions. We investigated the mechanism by which RPA regulates SMARCAL1. Our results indicate that although an interaction between SMARCAL1 and RPA is essential for SMARCAL1 activation, the location of the interacting surface on RPA is not. Counterintuitively, high-affinity DNA binding of RPA DNA-binding domain (DBD) A and DBD-B near the fork junction makes it easier for SMARCAL1 to remodel the fork, which requires removing RPA. We also found that RPA DBD-C and DBD-D are not required for SMARCAL1 regulation. Thus, the orientation of the high-affinity RPA DBDs at forks dictates SMARCAL1 substrate specificity. PMID:25552480

High-affinity DNA-binding domains of replication protein A (RPA) direct SMARCAL1-dependent replication fork remodeling.

Science.gov (United States)

Bhat, Kamakoti P; Bétous, Rémy; Cortez, David

2015-02-13

SMARCAL1 catalyzes replication fork remodeling to maintain genome stability. It is recruited to replication forks via an interaction with replication protein A (RPA), the major ssDNA-binding protein in eukaryotic cells. In addition to directing its localization, RPA also activates SMARCAL1 on some fork substrates but inhibits it on others, thereby conferring substrate specificity to SMARCAL1 fork-remodeling reactions. We investigated the mechanism by which RPA regulates SMARCAL1. Our results indicate that although an interaction between SMARCAL1 and RPA is essential for SMARCAL1 activation, the location of the interacting surface on RPA is not. Counterintuitively, high-affinity DNA binding of RPA DNA-binding domain (DBD) A and DBD-B near the fork junction makes it easier for SMARCAL1 to remodel the fork, which requires removing RPA. We also found that RPA DBD-C and DBD-D are not required for SMARCAL1 regulation. Thus, the orientation of the high-affinity RPA DBDs at forks dictates SMARCAL1 substrate specificity. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Deltorphins: a family of naturally occurring peptides with high affinity and selectivity for delta opioid binding sites.

Science.gov (United States)

Erspamer, V; Melchiorri, P; Falconieri-Erspamer, G; Negri, L; Corsi, R; Severini, C; Barra, D; Simmaco, M; Kreil, G

1989-07-01

Deltorphins are endogenous linear heptapeptides, isolated from skin extracts of frogs belonging to the genus Phyllomedusa, that have a higher affinity and selectivity for delta opioid binding sites than any other natural compound known. Two deltorphins with the sequence Tyr-Ala-Phe-Asp(or Glu)-Val-Val-Gly-NH2 have been isolated from skin extracts of Phyllomedusa bicolor. The alanine in position 2 is in the D configuration. These peptides, [D-Ala2]deltorphins I and II, show an even higher affinity for delta receptors than the previously characterized deltorphin, which contains D-methionine as the second amino acid. These peptides show some similarity to another constituent of Phyllomedusa skin, dermorphin, which is highly selective for mu-opioid receptors. These peptides all have the N-terminal sequence Tyr-D-Xaa-Phe, where D-Xaa is either D-alanine or D-methionine. While this structure seems to be capable of activating both mu and delta opioid receptors, differences in the C-terminal regions of these peptides are probably responsible for the observed high receptor selectivity of dermorphin and deltorphin.

New Synthesis and Tritium Labeling of a Selective Ligand for Studying High-Affinity γ-Hydroxybutyrate (GHB) Binding Sites

DEFF Research Database (Denmark)

Vogensen, Stine B.; Marek, Ales; Bay, Tina

2013-01-01

3-Hydroxycyclopent-1-enecarboxylic acid (HOCPCA, 1) is a potent ligand for the high-affinity GHB binding sites in the CNS. An improved synthesis of 1 together with a very efficient synthesis of [3H]-1 is described. The radiosynthesis employs in situ generated lithium trimethoxyborotritide. Screen...

Phosphopeptide enrichment by immobilized metal affinity chromatography

DEFF Research Database (Denmark)

Thingholm, Tine E.; Larsen, Martin R.

2016-01-01

Immobilized metal affinity chromatography (IMAC) has been the method of choice for phosphopeptide enrichment prior to mass spectrometric analysis for many years and it is still used extensively in many laboratories. Using the affinity of negatively charged phosphate groups towards positively...... charged metal ions such as Fe3+, Ga3+, Al3+, Zr4+, and Ti4+ has made it possible to enrich phosphorylated peptides from peptide samples. However, the selectivity of most of the metal ions is limited, when working with highly complex samples, e.g., whole-cell extracts, resulting in contamination from...

Mapping Affinities in Academic Organizations

Directory of Open Access Journals (Sweden)

Dario Rodighiero

2018-02-01

Full Text Available Scholarly affinities are one of the most fundamental hidden dynamics that drive scientific development. Some affinities are actual, and consequently can be measured through classical academic metrics such as co-authoring. Other affinities are potential, and therefore do not leave visible traces in information systems; for instance, some peers may share interests without actually knowing it. This article illustrates the development of a map of affinities for academic collectives, designed to be relevant to three audiences: the management, the scholars themselves, and the external public. Our case study involves the School of Architecture, Civil and Environmental Engineering of EPFL, hereinafter ENAC. The school consists of around 1,000 scholars, 70 laboratories, and 3 institutes. The actual affinities are modeled using the data available from the information systems reporting publications, teaching, and advising scholars, whereas the potential affinities are addressed through text mining of the publications. The major challenge for designing such a map is to represent the multi-dimensionality and multi-scale nature of the information. The affinities are not limited to the computation of heterogeneous sources of information; they also apply at different scales. The map, thus, shows local affinities inside a given laboratory, as well as global affinities among laboratories. This article presents a graphical grammar to represent affinities. Its effectiveness is illustrated by two actualizations of the design proposal: an interactive online system in which the map can be parameterized, and a large-scale carpet of 250 square meters. In both cases, we discuss how the materiality influences the representation of data, in particular the way key questions could be appropriately addressed considering the three target audiences: the insights gained by the management and their consequences in terms of governance, the understanding of the scholars’ own

Photoaffinity labeling of mammalian α1-adrenergic receptors: identification of the ligand binding subunit with a high affinity radioiodinated probe

International Nuclear Information System (INIS)

Leeb-Lundberg, L.M.F.; Dickinson, K.E.J.; Heald, S.L.

1984-01-01

A description is given of the synthesised and characterization of a novel high affinity radioiodinated α 1 -adrenergic receptor photoaffinity probe, 4-amino-6,7-dimethoxy-2-[4-[5-(4-azido-3-[ 125 I]iodophenyl)pentanoyl]-1-piperazinyl] quinazoline. In the absence of light, this ligand binds with high affinity (K/sub d/ = 130 pm) in a reverisble and saturable manner to sites in rat hepatic plasma membranes. The binding is stereoselective and competitively inhibited by adrenergic agonists and antagonists with an α 1 -adrenergic specificity. Upon photolysis, this ligand incorporates irreversibly into plasma membranes prepared from several mammalian tissues including rat liver, rat, guinea pig, and rabbit spleen, rabbit lung, and rabbit aorta vascular smooth muscle cells, also with typical α 1 -adrenergic specificity. Autoradiograms of such membrane samples subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis reveal a major specifically labeled polypeptide at M/sub 4/ = 78,000-85,000, depending on the tissue used, in addition to some lower molecular weight peptides. Protease inhibitors, in particular EDTA, a metalloprotease inhibitor, dramatically increases the predominance of the M/sub r/ = 78,000-85,000 polypeptide while attenuating the labeling of the lower molecular weight bands. This new high affinity radioiodinated photoaffinity probe should be of great value for the molecular characterization of the α 1 -adrenergic receptor

Immunostimulatory CpG-oligonucleotides induce functional high affinity IL-2 receptors on B-CLL cells: costimulation with IL-2 results in a highly immunogenic phenotype.

Science.gov (United States)

Decker, T; Schneller, F; Kronschnabl, M; Dechow, T; Lipford, G B; Wagner, H; Peschel, C

2000-05-01

CpG-oligodeoxynucleotides (CpG-ODN) have been shown to induce proliferation, cytokine production, and surface molecule regulation in normal and malignant human B cells. In the present study, we investigated the potential of CpG-ODN to induce functional high-affinity receptors in leukemic and normal B cells and the effects of costimulation with IL-2 on proliferation, cytokine secretion, and surface molecule regulation. Highly purified B cells from B-CLL patients and normal controls were stimulated with CpG-ODN with or without IL-2. Expression of CD25 was determined using FACS, and the presence of high-affinity IL-2 receptors was determined by scatchard analysis. Costimulatory effects of IL-2 and CpG-ODN were investigated using proliferation assays, ELISA (IL-6, TNF-alpha), and FACS analysis (CD80, CD86 expression). Reactivity of autologous and allogeneic T cells toward activated B-CLL cells was determined in mixed lymphocyte reactions and Interferon-gamma Elispot assays. The CpG-ODN DSP30 caused a significantly stronger induction of the IL-2 receptor alpha chain in malignant as compared with normal B cells (p = 0.03). This resulted in the expression of functional high-affinity IL-2 receptors in B-CLL cells, but fewer numbers of receptors with less affinity were expressed in normal B cells. Although addition of IL-2 to CpG-ODN-stimulated cells augmented proliferation in both normal B cells and B-CLL cells, no costimulatory effect on cytokine production or surface molecule expression could be observed in normal B cells. In contrast, TNF-alpha and IL-6 production was increased in B-CLL cells, and the expression of CD80 and CD86 was further enhanced when IL-2 was used as a costimulus. Autologous and allogeneic immune recognition of B-CLL cells stimulated with CpG-ODN and IL-2 was increased compared with B-CLL cells stimulated with CpG-ODN alone. Stimulation of B-CLL cells with CpG-ODN and IL-2 might be an attractive strategy for potential immunotherapies for B

Functional characterization of the high affinity IgG Receptor : making heads and tails of FcγRI

NARCIS (Netherlands)

van der Poel, C.E.

2011-01-01

This thesis focuses on human FcγRI, a high affinity receptor for antibodies of the IgG isotype. IgG is the most abundant antibody type in blood and all currently FDA approved therapeutic antibodies are of the IgG isotype. FcγRI, a member of the activating Fcγ receptors, exists as a complex of a

Affinity chromatographic purification of tetrodotoxin by use of tetrodotoxin-binding high molecular weight substances in the body fluid of shore crab (Hemigrapsus sanguineus) as ligands.

Science.gov (United States)

Shiomi, K; Yamaguchi, S; Shimakura, K; Nagashima, Y; Yamamori, K; Matsui, T

1993-12-01

A purification method for tetrodotoxin (TTX), based on affinity chromatography using the TTX-binding high mol. wt substances in the body fluid of shore crab (Hemigrapsus sanguineus) as ligands, was developed. This method was particularly useful for analysis of TTX in biological samples with low concentrations of TTX. The affinity gel prepared was highly specific for TTX, having no ability to bind 4-epi-TTX and anhydro-TTX as well as saxitoxin.

Endothelial targeting of high-affinity multivalent polymer nanocarriers directed to intercellular adhesion molecule 1.

Science.gov (United States)

Muro, Silvia; Dziubla, Thomas; Qiu, Weining; Leferovich, John; Cui, Xiumin; Berk, Erik; Muzykantov, Vladimir R

2006-06-01

Targeting of diagnostic and therapeutic agents to endothelial cells (ECs) provides an avenue to improve treatment of many maladies. For example, intercellular adhesion molecule 1 (ICAM-1), a constitutive endothelial cell adhesion molecule up-regulated in many diseases, is a good determinant for endothelial targeting of therapeutic enzymes and polymer nanocarriers (PNCs) conjugated with anti-ICAM (anti-ICAM/PNCs). However, intrinsic and extrinsic factors that control targeting of anti-ICAM/PNCs to ECs (e.g., anti-ICAM affinity and PNC valency and flow) have not been defined. In this study we tested in vitro and in vivo parameters of targeting to ECs of anti-ICAM/PNCs consisting of either prototype polystyrene or biodegradable poly(lactic-coglycolic) acid polymers (approximately 200 nm diameter spheres carrying approximately 200 anti-ICAM molecules). Anti-ICAM/PNCs, but not control IgG/PNCs 1) rapidly (t1/2 approximately 5 min) and specifically bound to tumor necrosis factor-activated ECs in a dose-dependent manner (Bmax approximately 350 PNC/cell) at both static and physiological shear stress conditions and 2) bound to ECs and accumulated in the pulmonary vasculature after i.v. injection in mice. Anti-ICAM/PNCs displayed markedly higher EC affinity versus naked anti-ICAM (Kd approximately 80 pM versus approximately 8 nM) in cell culture and, probably because of this factor, higher value (185.3 +/- 24.2 versus 50.5 +/- 1.5% injected dose/g) and selectivity (lung/blood ratio 81.0 +/- 10.9 versus 2.1 +/- 0.02, in part due to faster blood clearance) of pulmonary targeting. These results 1) show that reformatting monomolecular anti-ICAM into high-affinity multivalent PNCs boosts their vascular immuno-targeting, which withstands physiological hydrodynamics and 2) support potential anti-ICAM/PNCs utility for medical applications.

Fundamentals of affinity cell separations.

Science.gov (United States)

Zhang, Ye; Lyons, Veronica; Pappas, Dimitri

2018-03-01

Cell separations using affinity methods continue to be an enabling science for a wide variety of applications. In this review, we discuss the fundamental aspects of affinity separation, including the competing forces for cell capture and elution, cell-surface interactions, and models for cell adhesion. Factors affecting separation performance such as bond affinity, contact area, and temperature are presented. We also discuss and demonstrate the effects of nonspecific binding on separation performance. Metrics for evaluating cell separations are presented, along with methods of comparing separation techniques for cell isolation using affinity capture. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Comparison of high affinity binding of {sup 3}H-proadifen and {sup 3}H-(-)-cocaine t rat liver membranes

Energy Technology Data Exchange (ETDEWEB)

Ross, S.B. [Astra Arcus AB, Dept. of Neuropharmacology, Soedertaelje (Sweden)

1995-06-01

The characteristics of the binding of {sup 3}H-proadifen to rat liver membranes were studied and compared to those of {sup 3}H-cocaine. It was found that {sup 3}H-proadifen was bound reversibly with high affinity (K{sub D}=1.8{+-}0.5 nM) and large capacity (B{sub max}=2010{+-}340 pmol/g wet tissue) to liver membranes. The corresponding values for the {sup 3}H-cocaine binding were 3.5 nM and 1000 pmol/g wet tissue. The binding of {sup 3}H-proadifen was mainly localised to the microsomal fraction. The number of binding sites was not increased by treatment of rats with phenobarbitone. With 1 {mu}M CdCl{sub 2} in the incubation buffer it was possible to differentiate between two {sup 3}H-cocaine binding sites with K{sub d} values of 1.6 and 7.7 nM and B{sub max} values of 280 and 940 pmol/g wet liver tissue. S-(-)-Alaproclate inhibited the binding of {sup 3}H-proadifen and {sup 3}H-cocaine inhibited the binding of {sup 3}H-proadifen (IC{sub 50}=10 nM) and proadifen that of {sup 3}H-cocaine (IC{sub 50}=1 nM). There was a high correlation coefficient (r{sub r}=0.972; P<0.01; n=12) in the Spearman rank test between the inhibitory potencies of compounds examined in both systems. Beside some potent alaproclate analogues a couple of compounds had moderately high affinity (IC{sub 50}=100-500 nM): chloroquine, phenoxybenzamine, amitriptyline, ajmaline, remoxipride, imipramine and (-)-alaprenolol. CdCl{sub 2}, ZnCl{sub 2} and CuCl{sub 2} inhibited the binding of both ligands with low Hill coefficients, indicating heterogeneous binding sites. The inhibition curve of Cd{sup 2+} on the cocaine binding was biphasic with a high affinity part around 50 nM and a low affinity part at 15{mu}M. The similarity of the characteristics of the binding of these ligands with that of {sup 3}H-alaproclate is discussed. It is suggested that all three compounds bind to the same sites, although additional binding sites seem to exist for proadifen. (au) (9 refs.).

High-Affinity Low-Capacity and Low-Affinity High-Capacity N-Acetyl-2-Aminofluorene (AAF) Macromolecular Binding Sites Are Revealed During the Growth Cycle of Adult Rat Hepatocytes in Primary Culture.

Science.gov (United States)

Koch, Katherine S; Moran, Tom; Shier, W Thomas; Leffert, Hyam L

2018-05-01

Long-term cultures of primary adult rat hepatocytes were used to study the effects of N-acetyl-2-aminofluorene (AAF) on hepatocyte proliferation during the growth cycle; on the initiation of hepatocyte DNA synthesis in quiescent cultures; and, on hepatocyte DNA replication following the initiation of DNA synthesis. Scatchard analyses were used to identify the pharmacologic properties of radiolabeled AAF metabolite binding to hepatocyte macromolecules. Two classes of growth cycle-dependent AAF metabolite binding sites-a high-affinity low-capacity site (designated Site I) and a low-affinity high-capacity site (designated Site II)-associated with two spatially distinct classes of macromolecular targets, were revealed. Based upon radiolabeled AAF metabolite binding to purified hepatocyte genomic DNA or to DNA, RNA, proteins, and lipids from isolated nuclei, Site IDAY 4 targets (KD[APPARENT] ≈ 2-4×10-6 M and BMAX[APPARENT] ≈ 6 pmol/106 cells/24 h) were consistent with genomic DNA; and with AAF metabolized by a nuclear cytochrome P450. Based upon radiolabeled AAF binding to total cellular lysates, Site IIDAY 4 targets (KD[APPARENT] ≈ 1.5×10-3 M and BMAX[APPARENT] ≈ 350 pmol/106 cells/24 h) were consistent with cytoplasmic proteins; and with AAF metabolized by cytoplasmic cytochrome P450s. DNA synthesis was not inhibited by concentrations of AAF that saturated DNA binding in the neighborhood of the Site I KD. Instead, hepatocyte DNA synthesis inhibition required higher concentrations of AAF approaching the Site II KD. These observations raise the possibility that carcinogenic DNA adducts derived from AAF metabolites form below concentrations of AAF that inhibit replicative and repair DNA synthesis.

Effect of human chorionic gonadotropin on sexual maturation, sex steroids and thyroid hormone levels in Caspian lamprey (Caspiomyzon wagneri Kessler, 1870)

OpenAIRE

Abedi, M.; Mojazi Amiri, B.; Abdoli, A.; Javanshir, A.; Benam, S.; Namdarian, A.

2017-01-01

The objective of this study was to determine the effect of human chorionic gonadotropin (hCG) on sexual maturation, plasma sex steroids [17β-estradiol, (E2) and 17α-hydroxy progesterone (17α_OHP)] and thyroid hormones (triiodothyronine, T3 and thyroxin, T4) levels in upstream - migrating Caspian lamprey. During the experiment, 36 fish (24 females and 12 males) in spring 2013 and 36 fish (24 females and 12 males) in fall 2013 were collected from the Shirud River estuary in Mazandaran Province,...

Peptides in headlock ? a novel high-affinity and versatile peptide-binding nanobody for proteomics and microscopy

OpenAIRE

Braun, Michael B.; Traenkle, Bjoern; Koch, Philipp A.; Emele, Felix; Weiss, Frederik; Poetz, Oliver; Stehle, Thilo; Rothbauer, Ulrich

2016-01-01

Nanobodies are highly valuable tools for numerous bioanalytical and biotechnical applications. Here, we report the characterization of a nanobody that binds a short peptide epitope with extraordinary affinity. Structural analysis reveals an unusual binding mode where the extended peptide becomes part of a ?-sheet structure in the nanobody. This interaction relies on sequence-independent backbone interactions augmented by a small number of specificity-determining side chain contacts. Once boun...

High affinity antigen recognition of the dual specific variants of herceptin is entropy-driven in spite of structural plasticity.

Directory of Open Access Journals (Sweden)

Jenny Bostrom

Full Text Available The antigen-binding site of Herceptin, an anti-human Epidermal Growth Factor Receptor 2 (HER2 antibody, was engineered to add a second specificity toward Vascular Endothelial Growth Factor (VEGF to create a high affinity two-in-one antibody bH1. Crystal structures of bH1 in complex with either antigen showed that, in comparison to Herceptin, this antibody exhibited greater conformational variability, also called "structural plasticity". Here, we analyzed the biophysical and thermodynamic properties of the dual specific variants of Herceptin to understand how a single antibody binds two unrelated protein antigens. We showed that while bH1 and the affinity-improved bH1-44, in particular, maintained many properties of Herceptin including binding affinity, kinetics and the use of residues for antigen recognition, they differed in the binding thermodynamics. The interactions of bH1 and its variants with both antigens were characterized by large favorable entropy changes whereas the Herceptin/HER2 interaction involved a large favorable enthalpy change. By dissecting the total entropy change and the energy barrier for dual interaction, we determined that the significant structural plasticity of the bH1 antibodies demanded by the dual specificity did not translate into the expected increase of entropic penalty relative to Herceptin. Clearly, dual antigen recognition of the Herceptin variants involves divergent antibody conformations of nearly equivalent energetic states. Hence, increasing the structural plasticity of an antigen-binding site without increasing the entropic cost may play a role for antibodies to evolve multi-specificity. Our report represents the first comprehensive biophysical analysis of a high affinity dual specific antibody binding two unrelated protein antigens, furthering our understanding of the thermodynamics that drive the vast antigen recognition capacity of the antibody repertoire.

Two distinct affinity binding sites for IL-1 on human cell lines

International Nuclear Information System (INIS)

Bensimon, C.; Wakasugi, N.; Tagaya, Y.; Takakura, K.; Yodoi, J.; Tursz, T.; Wakasugi, H.

1989-01-01

We used two human cell lines, NK-like YT-C3 and an EBV-containing B cell line, 3B6, as models to study the receptor(s) for IL-1. Two distinct types of saturable binding sites were found on both cell lines at 37 degrees C. Between 1 pM and 100 pM of 125I-IL-1-alpha concentration, saturable binding sites were detected on the YT-C3 cells with a K of 4 x 10(-11) M. The K found for the IL-1-alpha binding sites on 3B6 cells was 7.5 x 10(-11) M. An additional binding curve was detected above 100 pM on YT-C3 cells with a K of 7 x 10(-9) M and on 3B6 cells with a K of 5 x 10(-9) M. Scatchard plot analysis revealed 600 sites/cell with high affinity binding and 7000 sites/cell with low affinity for YT-C3 cells and 300 sites/cell with high affinity binding and 6000 sites/cell with low affinity for 3B6 cells. At 37 degrees C, the internalization of 125I-labeled IL-1 occurred via both high and low affinity IL-1R on both YT-C3 and 3B6 cells, whereas the rates of internalization for high affinity binding sites on YT-C3 cells were predominant in comparison to that of low affinity binding sites. In chemical cross-linking studies of 125 I-IL-1-alpha to 3B6 and YT-C3 cells, two protein bands were immunoprecipitated with Mr around 85 to 90 kDa leading to an estimation of the Mr of the IL-1R around 68 to 72 kDa. In similar experiments, the Mr found for the IL-1R expressed on the murine T cell line EL4 was slightly higher (around 80 kDa). Whether these distinct affinity binding sites are shared by a single molecule or by various chains remains to be elucidated

Dopamine inhibition of anterior pituitary adenylate cyclase is mediated through the high-affinity state of the D2 receptor

International Nuclear Information System (INIS)

Borgundvaag, B.; George, S.R.

1985-01-01

The diterpinoid forskolin stimulated adenylate cyclase activity (measured by conversion of [ 3 H]-ATP to [ 3 H]-cAMP) in anterior pituitary from male and female rats. Inhibition of stimulated adenylate cyclase activity by potent dopaminergic agonists was demonstrable only in female anterior pituitary. The inhibition of adenylate cyclase activity displayed a typically dopaminergic rank order of agonist potencies and could be completely reversed by a specific dopamine receptor antagonist. The IC 50 values of dopamine agonist inhibition of adenylate cyclase activity correlated with equal molarity with the dissociation constant of the high-affinity dopamine agonist-detected receptor binding site and with the IC 50 values for inhibition of prolactin secretion. These findings support the hypothesis that it is the high-affinity form of the D 2 dopamine receptor in anterior pituitary which is responsible for mediating the dopaminergic function of attenuating adenylate cyclase activity. 12 references, 4 figures, 1 table

Generation and characterization of a human-mouse chimeric high-affinity antibody that detects the DYKDDDDK FLAG peptide.

Science.gov (United States)

Ikeda, Koki; Koga, Tomoaki; Sasaki, Fumiyuki; Ueno, Ayumi; Saeki, Kazuko; Okuno, Toshiaki; Yokomizo, Takehiko

2017-05-13

DYKDDDDK peptide (FLAG) is a useful tool for investigating the function and localization of proteins whose antibodies (Abs) are not available. We recently established a high-affinity monoclonal antibody (mAb) for FLAG (clone 2H8). The 2H8 Ab is highly sensitive for detecting FLAG-tagged proteins by flowcytometry and immunoprecipitation, but it can yield nonspecific signals in immunohistochemistry of mouse tissues because it is of mouse origin. In this study, we reduced nonspecific signals by generating a chimeric 2H8 Ab with Fc fragments derived from human immunoglobulin. We fused a 5' terminal cDNA fragments for the Fab region of 2H8 mAb with 3' terminal cDNA fragments for Fc region of human IgG1. We transfected both chimeric plasmids and purified the resulting human-mouse chimeric 2H8. The chimeric 2H8 Ab successfully detected FLAG-tagged proteins in flowcytometry with anti-human IgG secondary Ab with comparable sensitivity to 2H8 mAb. Importantly, chimeric 2H8 detected specific FLAG peptide signals without nonspecific signals in immunohistochemical analysis with mouse tissues. This human-mouse chimeric high-affinity anti-FLAG Ab will prove useful for future immunohistochemical analysis of mouse tissues. Copyright © 2017 Elsevier Inc. All rights reserved.

A High Affinity Adenosine Kinase from Anopheles gambiae

Science.gov (United States)

Cassera, María B.; Ho, Meng-Chiao; Merino, Emilio F.; Burgos, Emmanuel S.; Rinaldo-Matthis, Agnes; Almo, Steven C.; Schramm, Vern L.

2011-01-01

Genome analysis revealed a mosquito orthologue of adenosine kinase in Anopheles gambiae (AgAK; the most important vector for the transmission of Plasmodium falciparum in Africa). P. falciparum are purine auxotrophs and do not express an adenosine kinase but rely on their hosts for purines. AgAK was kinetically characterized and found to have the highest affinity for adenosine (Km 8.1 nM) of any known adenosine kinase. AgAK is specific for adenosine at the nucleoside site but several nucleotide triphosphate phosphoryl donors are tolerated. The AgAK crystal structure with a bound bisubstrate analogue Ap4A (2.0 Å resolution) reveals interactions for adenosine, ATP and the geometry for phosphoryl transfer. The polyphosphate charge is partly neutralized by a bound Mg2+ ion and an ion pair to a catalytic site Arg. The AgAK structure consists of a large catalytic core in a three-layered α/β/α sandwich, and a small cap domain in contact with adenosine. The specificity and tight-binding for adenosine arises from hydrogen bond interactions of Asn14, Leu16, Leu40, Leu133, Leu168, Phe168 and Thr171 and the backbone of Ile39 and Phe168 with the adenine ring as well as through hydrogen bond interactions between Asp18, Gly64 and Asn68 and the ribosyl 2′- and 3′-hydroxyl groups. The structure is more similar to human adenosine kinase (48% identity) than to AK from Toxoplasma gondii (31% identity). With this extraordinary affinity for AgAK, adenosine is efficiently captured and converted to AMP at near the diffusion limit, suggesting an important role of this enzyme to maintain the adenine nucleotide pool. mRNA analysis verifies that AgAK transcripts are produced in the adult insects. PMID:21247194

Affinity chromatography: A versatile technique for antibody purification.

Science.gov (United States)

Arora, Sushrut; Saxena, Vikas; Ayyar, B Vijayalakshmi

2017-03-01

Antibodies continue to be extremely utilized entities in myriad applications including basic research, imaging, targeted delivery, chromatography, diagnostics, and therapeutics. At production stage, antibodies are generally present in complex matrices and most of their intended applications necessitate purification. Antibody purification has always been a major bottleneck in downstream processing of antibodies, due to the need of high quality products and associated high costs. Over the years, extensive research has focused on finding better purification methodologies to overcome this holdup. Among a plethora of different techniques, affinity chromatography is one of the most selective, rapid and easy method for antibody purification. This review aims to provide a detailed overview on affinity chromatography and the components involved in purification. An array of support matrices along with various classes of affinity ligands detailing their underlying working principles, together with the advantages and limitations of each system in purifying different types of antibodies, accompanying recent developments and important practical methodological considerations to optimize purification procedure are discussed. Copyright © 2016 Elsevier Inc. All rights reserved.

Affinity purification using recombinant PXR as a tool to characterize environmental ligands.

Science.gov (United States)

Dagnino, Sonia; Bellet, Virginie; Grimaldi, Marina; Riu, Anne; Aït-Aïssa, Sélim; Cavaillès, Vincent; Fenet, Hélène; Balaguer, Patrick

2014-02-01

Many environmental endocrine disrupting compounds act as ligands for nuclear receptors. The human pregnane X receptor (hPXR), for instance, is activated by a variety of environmental ligands such as steroids, pharmaceutical drugs, pesticides, alkylphenols, polychlorinated biphenyls and polybromo diethylethers. Some of us have previously reported the occurrence of hPXR ligands in environmental samples but failed to identify them. The aim of this study was to test whether a PXR-affinity column, in which recombinant hPXR was immobilized on solid support, could help the purification of these chemicals. Using PXR ligands of different affinity (10 nM < EC50 < 10 μM), we demonstrated that the PXR-affinity preferentially column captured ligands with medium to high affinities (EC50 < 1 μM). Furthermore, by using the PXR-affinity column to analyze an environmental sample containing ERα, AhR, AR, and PXR activities, we show that (i) half of the PXR activity of the sample was due to compounds with medium to high affinity for PXR and (ii) PXR shared ligands with ERα, AR, and AhR. These findings demonstrate that the newly developed PXR-affinity column coupled to reporter cell lines represents a valuable tool for the characterization of the nature of PXR active compounds and should therefore guide and facilitate their further analysis. Copyright © 2012 Wiley Periodicals, Inc., a Wiley company.

Affine field theories

International Nuclear Information System (INIS)

Cadavid, A.C.

1989-01-01

The author constructs a non-Abelian field theory by gauging a Kac-Moody algebra, obtaining an infinite tower of interacting vector fields and associated ghosts, that obey slightly modified Feynman rules. She discusses the spontaneous symmetry breaking of such theory via the Higgs mechanism. If the Higgs particle lies in the Cartan subalgebra of the Kac-Moody algebra, the previously massless vectors acquire a mass spectrum that is linear in the Kac-Moody index and has additional fine structure depending on the associated Lie algebra. She proceeds to show that there is no obstacle in implementing the affine extension of supersymmetric Yang-Mills theories. The result is valid in four, six and ten space-time dimensions. Then the affine extension of supergravity is investigated. She discusses only the loop algebra since the affine extension of the super-Poincare algebra appears inconsistent. The construction of the affine supergravity theory is carried out by the group manifold method and leads to an action describing infinite towers of spin 2 and spin 3/2 fields that interact subject to the symmetries of the loop algebra. The equations of motion satisfy the usual consistency check. Finally, she postulates a theory in which both the vector and scalar fields lie in the loop algebra of SO(3). This theory has an expanded soliton sector, and corresponding to the original 't Hooft-Polyakov solitonic solutions she now finds an infinite family of exact, special solutions of the new equations. She also proposes a perturbation method for obtaining an arbitrary solution of those equations for each level of the affine index

Structural basis for high substrate-binding affinity and enantioselectivity of 3-quinuclidinone reductase AtQR

International Nuclear Information System (INIS)

Hou, Feng; Miyakawa, Takuya; Kataoka, Michihiko; Takeshita, Daijiro; Kumashiro, Shoko; Uzura, Atsuko; Urano, Nobuyuki; Nagata, Koji; Shimizu, Sakayu; Tanokura, Masaru

2014-01-01

Highlights: • Crystal structure of AtQR has been determined at 1.72 Å. • NADH binding induces the formation of substrate binding site. • AtQR possesses a conserved hydrophobic wall for stereospecific binding of substrate. • Additional Glu197 residue is critical to the high binding affinity. - Abstract: (R)-3-Quinuclidinol, a useful compound for the synthesis of various pharmaceuticals, can be enantioselectively produced from 3-quinuclidinone by 3-quinuclidinone reductase. Recently, a novel NADH-dependent 3-quinuclidionone reductase (AtQR) was isolated from Agrobacterium tumefaciens, and showed much higher substrate-binding affinity (>100 fold) than the reported 3-quinuclidionone reductase (RrQR) from Rhodotorula rubra. Here, we report the crystal structure of AtQR at 1.72 Å. Three NADH-bound protomers and one NADH-free protomer form a tetrameric structure in an asymmetric unit of crystals. NADH not only acts as a proton donor, but also contributes to the stability of the α7 helix. This helix is a unique and functionally significant part of AtQR and is related to form a deep catalytic cavity. AtQR has all three catalytic residues of the short-chain dehydrogenases/reductases family and the hydrophobic wall for the enantioselective reduction of 3-quinuclidinone as well as RrQR. An additional residue on the α7 helix, Glu197, exists near the active site of AtQR. This acidic residue is considered to form a direct interaction with the amine part of 3-quinuclidinone, which contributes to substrate orientation and enhancement of substrate-binding affinity. Mutational analyses also support that Glu197 is an indispensable residue for the activity

Structural basis for high substrate-binding affinity and enantioselectivity of 3-quinuclidinone reductase AtQR

Energy Technology Data Exchange (ETDEWEB)

Hou, Feng; Miyakawa, Takuya [Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657 (Japan); Kataoka, Michihiko [Division of Applied Life Sciences, Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen-cho, Naka-ku, Sakai 559-8531 (Japan); Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-Oiwakecho, Sakyo-ku, Kyoto 606-8502 (Japan); Takeshita, Daijiro [Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657 (Japan); Kumashiro, Shoko [Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-Oiwakecho, Sakyo-ku, Kyoto 606-8502 (Japan); Uzura, Atsuko [Research and Development Center, Nagase and Co., Ltd., 2-2-3 Muratani, Nishi-ku, Kobe 651-2241 (Japan); Urano, Nobuyuki [Division of Applied Life Sciences, Graduate School of Life and Environmental Sciences, Osaka Prefecture University, 1-1 Gakuen-cho, Naka-ku, Sakai 559-8531 (Japan); Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-Oiwakecho, Sakyo-ku, Kyoto 606-8502 (Japan); Nagata, Koji [Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657 (Japan); Shimizu, Sakayu [Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-Oiwakecho, Sakyo-ku, Kyoto 606-8502 (Japan); Faculty of Bioenvironmental Science, Kyoto Gakuen University, Sogabe-cho, Kameoka 621-8555 (Japan); Tanokura, Masaru, E-mail: amtanok@mail.ecc.u-tokyo.ac.jp [Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657 (Japan)

2014-04-18

Highlights: • Crystal structure of AtQR has been determined at 1.72 Å. • NADH binding induces the formation of substrate binding site. • AtQR possesses a conserved hydrophobic wall for stereospecific binding of substrate. • Additional Glu197 residue is critical to the high binding affinity. - Abstract: (R)-3-Quinuclidinol, a useful compound for the synthesis of various pharmaceuticals, can be enantioselectively produced from 3-quinuclidinone by 3-quinuclidinone reductase. Recently, a novel NADH-dependent 3-quinuclidionone reductase (AtQR) was isolated from Agrobacterium tumefaciens, and showed much higher substrate-binding affinity (>100 fold) than the reported 3-quinuclidionone reductase (RrQR) from Rhodotorula rubra. Here, we report the crystal structure of AtQR at 1.72 Å. Three NADH-bound protomers and one NADH-free protomer form a tetrameric structure in an asymmetric unit of crystals. NADH not only acts as a proton donor, but also contributes to the stability of the α7 helix. This helix is a unique and functionally significant part of AtQR and is related to form a deep catalytic cavity. AtQR has all three catalytic residues of the short-chain dehydrogenases/reductases family and the hydrophobic wall for the enantioselective reduction of 3-quinuclidinone as well as RrQR. An additional residue on the α7 helix, Glu197, exists near the active site of AtQR. This acidic residue is considered to form a direct interaction with the amine part of 3-quinuclidinone, which contributes to substrate orientation and enhancement of substrate-binding affinity. Mutational analyses also support that Glu197 is an indispensable residue for the activity.

Enhanced membrane pore formation through high-affinity targeted antimicrobial peptides.

Directory of Open Access Journals (Sweden)

Christopher J Arnusch

Full Text Available Many cationic antimicrobial peptides (AMPs target the unique lipid composition of the prokaryotic cell membrane. However, the micromolar activities common for these peptides are considered weak in comparison to nisin, which follows a targeted, pore-forming mode of action. Here we show that AMPs can be modified with a high-affinity targeting module, which enables membrane permeabilization at low concentration. Magainin 2 and a truncated peptide analog were conjugated to vancomycin using click chemistry, and could be directed towards specific membrane embedded receptors both in model membrane systems and whole cells. Compared with untargeted vesicles, a gain in permeabilization efficacy of two orders of magnitude was reached with large unilamellar vesicles that included lipid II, the target of vancomycin. The truncated vancomycin-peptide conjugate showed an increased activity against vancomycin resistant Enterococci, whereas the full-length conjugate was more active against a targeted eukaryotic cell model: lipid II containing erythrocytes. This study highlights that AMPs can be made more selective and more potent against biological membranes that contain structures that can be targeted.

Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging

Energy Technology Data Exchange (ETDEWEB)

Orcutt, Kelly Davis; Slusarczyk, Adrian L. [Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Cieslewicz, Maryelise [Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Ruiz-Yi, Benjamin [Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Bhushan, Kumar R. [Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, Boston, MA 02215 (United States); Frangioni, John V. [Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, Boston, MA 02215 (United States); Department of Radiology, Beth Israel Deaconess Medical Center, Boston, MA 02215 (United States); Wittrup, K. Dane, E-mail: wittrup@mit.ed [Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States)

2011-02-15

Introduction: In pretargeted radioimmunotherapy (PRIT), a bifunctional antibody is administered and allowed to pre-localize to tumor cells. Subsequently, a chelated radionuclide is administered and captured by cell-bound antibody while unbound hapten clears rapidly from the body. We aim to engineer high-affinity binders to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelates for use in PRIT applications. Methods: We mathematically modeled antibody and hapten pharmacokinetics to analyze hapten tumor retention as a function of hapten binding affinity. Motivated by model predictions, we used directed evolution and yeast surface display to affinity mature the 2D12.5 antibody to DOTA, reformatted as a single chain variable fragment (scFv). Results: Modeling predicts that for high antigen density and saturating bsAb dose, a hapten-binding affinity of 100 pM is needed for near-maximal hapten retention. We affinity matured 2D12.5 with an initial binding constant of about 10 nM to DOTA-yttrium chelates. Affinity maturation resulted in a 1000-fold affinity improvement to biotinylated DOTA-yttrium, yielding an 8.2{+-}1.9 picomolar binder. The high-affinity scFv binds DOTA complexes of lutetium and gadolinium with similar picomolar affinity and indium chelates with low nanomolar affinity. When engineered into a bispecific antibody construct targeting carcinoembryonic antigen, pretargeted high-affinity scFv results in significantly higher tumor retention of a {sup 111}In-DOTA hapten compared to pretargeted wild-type scFv in a xenograft mouse model. Conclusions: We have engineered a versatile, high-affinity, DOTA-chelate-binding scFv. We anticipate it will prove useful in developing pretargeted imaging and therapy protocols to exploit the potential of a variety of radiometals.

Electron affinities: theoretical

International Nuclear Information System (INIS)

Kaufman, J.J.

1976-01-01

A brief description is given of the conceptual background and formalism of the various ab-initio and semi-ab-initio quantum computational techniques for calculating atomic and molecular electron affinities: Hartree--Fock--Roothaan SCF, configuration interaction (CI), multiconfiguration SCF (MC-SCF), Bethe--Goldstone, superposition of configurations (SOC), ab-initio effective core model potentials, Xα-MS, plus other less common methods. Illustrative and comparative examples of electron affinities calculated by these various methods are presented

Supramolecular Affinity Chromatography for Methylation-Targeted Proteomics.

Science.gov (United States)

Garnett, Graham A E; Starke, Melissa J; Shaurya, Alok; Li, Janessa; Hof, Fraser

2016-04-05

Proteome-wide studies of post-translationally methylated species using mass spectrometry are complicated by high sample diversity, competition for ionization among peptides, and mass redundancies. Antibody-based enrichment has powered methylation proteomics until now, but the reliability, pan-specificity, polyclonal nature, and stability of the available pan-specific antibodies are problematic and do not provide a standard, reliable platform for investigators. We have invented an anionic supramolecular host that can form host-guest complexes selectively with methyllysine-containing peptides and used it to create a methylysine-affinity column. The column resolves peptides on the basis of methylation-a feat impossible with a comparable commercial cation-exchange column. A proteolyzed nuclear extract was separated on the methyl-affinity column prior to standard proteomics analysis. This experiment demonstrates that such chemical methyl-affinity columns are capable of enriching and improving the analysis of methyllysine residues from complex protein mixtures. We discuss the importance of this advance in the context of biomolecule-driven enrichment methods.

Kinetics and autoradiography of high affinity uptake of serotonin by primary astrocyte cultures

International Nuclear Information System (INIS)

Katz, D.M.; Kimelberg, H.K.

1985-01-01

Primary astrocyte cultures prepared from the cerebral cortices of neonatal rats showed significant accumulation of serotonin (5-hydroxytryptamine; [ 3 H]-5-HT). At concentrations in the range of 0.01 to 0.7 microM [ 3 H]-5-HT, this uptake was 50 to 85% Na+ dependent and gave a Km of 0.40 +/- 0.11 microM [ 3 H]-5-HT and a Vmax of 6.42 +/- 0.85 (+/- SEM) pmol of [ 3 H]-5-HT/mg of protein/4 min for the Na+-dependent component. In the absence of Na+ the uptake was nonsaturable. Omission of the monoamine oxidase inhibitor pargyline markedly reduced the Na+-dependent component of [ 3 H]-5-HT uptake but had a negligible effect on the Na+-independent component. This suggest significant oxidative deamination of serotonin after it has been taken up by the high affinity system, followed by release of its metabolite. The authors estimated that this system enabled the cells to concentrate [ 3 H]-5-HT up to 44-fold at an external [ 3 H]-5-HT concentration of 10(-7) M. Inhibition of [ 3 H]-5-HT uptake by a number of clinically effective antidepressants was also consistent with a specific high affinity uptake mechanism for 5-HT, the order of effectiveness of inhibition being chlorimipramine greater than fluoxetine greater than imipramine = amitriptyline greater than desmethylimipramine greater than iprindole greater than mianserin. Uptake of [ 3 H]-5-HT was dependent on the presence of Cl- as well as Na+ in the medium, and the effect of omission of both ions was nonadditive. Varying the concentration of K+ in the media from 1 to 50 mM had a limited effect on [ 3 H]-5-HT uptake

Transformations Based on Continuous Piecewise-Affine Velocity Fields

DEFF Research Database (Denmark)

Freifeld, Oren; Hauberg, Søren; Batmanghelich, Kayhan

2017-01-01

We propose novel finite-dimensional spaces of well-behaved transformations. The latter are obtained by (fast and highly-accurate) integration of continuous piecewise-affine velocity fields. The proposed method is simple yet highly expressive, effortlessly handles optional constraints (e.g., volum...

Effect of quinolinic acid in the nucleus basalis magnocellularis on cortical high-affinity choline uptake

Energy Technology Data Exchange (ETDEWEB)

Metcalf, R.H.; Boegman, R.J.; Quirion, R.; Riopelle, R.J.; Ludwin, S.K.

1987-08-01

A transient 45% increase in cortical high-affinity choline uptake (HACU) was observed after an injection of quinolinic acid (QUIN) into the nucleus basalis magnocellularis (nbM) of the rat. This was followed by a steady decline in choline uptake, which resulted in a 46% decrease by day 7. Specific (/sup 3/H)hemicholinium-3 binding to coronal brain sections showed a similar pattern following injections of QUIN into the nbM. The increase in cortical HACU elicited by QUIN appeared to be dose dependent.

Comparison of high affinity binding of 3H-proadifen and 3H-(-)-cocaine t rat liver membranes

International Nuclear Information System (INIS)

Ross, S.B.

1995-01-01

The characteristics of the binding of 3 H-proadifen to rat liver membranes were studied and compared to those of 3 H-cocaine. It was found that 3 H-proadifen was bound reversibly with high affinity (K D =1.8±0.5 nM) and large capacity (B max =2010±340 pmol/g wet tissue) to liver membranes. The corresponding values for the 3 H-cocaine binding were 3.5 nM and 1000 pmol/g wet tissue. The binding of 3 H-proadifen was mainly localised to the microsomal fraction. The number of binding sites was not increased by treatment of rats with phenobarbitone. With 1 μM CdCl 2 in the incubation buffer it was possible to differentiate between two 3 H-cocaine binding sites with K d values of 1.6 and 7.7 nM and B max values of 280 and 940 pmol/g wet liver tissue. S-(-)-Alaproclate inhibited the binding of 3 H-proadifen and 3 H-cocaine inhibited the binding of 3 H-proadifen (IC 50 =10 nM) and proadifen that of 3 H-cocaine (IC 50 =1 nM). There was a high correlation coefficient (r r =0.972; P 50 =100-500 nM): chloroquine, phenoxybenzamine, amitriptyline, ajmaline, remoxipride, imipramine and (-)-alaprenolol. CdCl 2 , ZnCl 2 and CuCl 2 inhibited the binding of both ligands with low Hill coefficients, indicating heterogeneous binding sites. The inhibition curve of Cd 2+ on the cocaine binding was biphasic with a high affinity part around 50 nM and a low affinity part at 15μM. The similarity of the characteristics of the binding of these ligands with that of 3 H-alaproclate is discussed. It is suggested that all three compounds bind to the same sites, although additional binding sites seem to exist for proadifen. (au) (9 refs.)

Affinities and densities of high-affinity [3H]muscimol (GABA-A) binding sites and of central benzodiazepine receptors are unchanged in autopsied brain tissue from cirrhotic patients with hepatic encephalopathy

International Nuclear Information System (INIS)

Butterworth, R.F.; Lavoie, J.; Giguere, J.F.; Pomier-Layrargues, G.

1988-01-01

The integrity of GABA-A receptors and of central benzodiazepine receptors was evaluated in membrane preparations from prefrontal cortex and caudate nuclei obtained at autopsy from nine cirrhotic patients who died in hepatic coma and an equal number of age-matched control subjects. Histopathological studies revealed Alzheimer Type II astrocytosis in all cases in the cirrhotic group; controls were free from neurological, psychiatric or hepatic diseases. Binding to GABA-A receptors was studied using [ 3 H]muscimol as radioligand. The integrity of central benzodiazepine receptors was evaluated using [ 3 H]flunitrazepam and [ 3 H]Ro15-1788. Data from saturation binding assays was analyzed by Scatchard plot. No modifications of either affinities (Kd) or densities (Bmax) of [ 3 H]muscimol of central benzodiazepine binding sites were observed. These findings do not support recent suggestions that alterations of either high-affinity GABA or benzodiazepine receptors play a significant role in the pathogenesis of hepatic encephalopathy

Affine-Invariant Geometric Constraints-Based High Accuracy Simultaneous Localization and Mapping

Directory of Open Access Journals (Sweden)

Gangchen Hua

2017-01-01

Full Text Available In this study we describe a new appearance-based loop-closure detection method for online incremental simultaneous localization and mapping (SLAM using affine-invariant-based geometric constraints. Unlike other pure bag-of-words-based approaches, our proposed method uses geometric constraints as a supplement to improve accuracy. By establishing an affine-invariant hypothesis, the proposed method excludes incorrect visual words and calculates the dispersion of correctly matched visual words to improve the accuracy of the likelihood calculation. In addition, camera’s intrinsic parameters and distortion coefficients are adequate for this method. 3D measuring is not necessary. We use the mechanism of Long-Term Memory and Working Memory (WM to manage the memory. Only a limited size of the WM is used for loop-closure detection; therefore the proposed method is suitable for large-scale real-time SLAM. We tested our method using the CityCenter and Lip6Indoor datasets. Our proposed method results can effectively correct the typical false-positive localization of previous methods, thus gaining better recall ratios and better precision.

Isolation and characterization of high affinity aptamers against DNA polymerase iota.

Science.gov (United States)

Lakhin, Andrei V; Kazakov, Andrei A; Makarova, Alena V; Pavlov, Yuri I; Efremova, Anna S; Shram, Stanislav I; Tarantul, Viacheslav Z; Gening, Leonid V

2012-02-01

Human DNA-polymerase iota (Pol ι) is an extremely error-prone enzyme and the fidelity depends on the sequence context of the template. Using the in vitro systematic evolution of ligands by exponential enrichment (SELEX) procedure, we obtained an oligoribonucleotide with a high affinity to human Pol ι, named aptamer IKL5. We determined its dissociation constant with homogenous preparation of Pol ι and predicted its putative secondary structure. The aptamer IKL5 specifically inhibits DNA-polymerase activity of the purified enzyme Pol ι, but did not inhibit the DNA-polymerase activities of human DNA polymerases beta and kappa. IKL5 suppressed the error-prone DNA-polymerase activity of Pol ι also in cellular extracts of the tumor cell line SKOV-3. The aptamer IKL5 is useful for studies of the biological role of Pol ι and as a potential drug to suppress the increase of the activity of this enzyme in malignant cells.

High-affinity cannabinoid binding site in brain: A possible marijuana receptor

International Nuclear Information System (INIS)

Nye, J.S.

1988-01-01

The mechanism by which delta 9 tetrahydrocannabinol (delta 9 THC), the major psychoactive component of marijuana or hashish, produces its potent psychological and physiological effects is unknown. To find receptor binding sites for THC, we designed a water-soluble analog for use as a radioligand. 5'-Trimethylammonium-delta 8 THC (TMA) is a positively charged analog of delta- 8 THC modified on the 5' carbon, a portion of the molecule not important for its psychoactivity. We have studied the binding of [ 3 H]-5'-trimethylammonium-delta- 8 THC ([ 3 H]TMA) to rat neuronal membranes. [ 3 H]TMA binds saturably and reversibly to brain membranes with high affinity to apparently one class of sites. Highest binding site density occurs in brain, but several peripheral organs also display specific binding. Detergent solubilizes the sites without affecting their pharmacologial properties. Molecular sieve chromatography reveals a bimodal peak of [ 3 H]TMA binding activity of approximately 60,000 daltons apparent molecular weight

Benzodiazepines have high-affinity binding sites and induce melanogenesis in B16/C3 melanoma cells.

OpenAIRE

Matthew, E; Laskin, J D; Zimmerman, E A; Weinstein, I B; Hsu, K C; Engelhardt, D L

1981-01-01

We found that two markers of differentiation, tyrosinase (monophenol, dihydroxyphenylalanine:oxygen oxidoreductase, EC 1.14.18.1) activity and melanin synthesis, are induced by diazepam in B16/C3 mouse melanoma cells. We also demonstrated high-affinity binding sites for [3H]diazepam in these cells by radioreceptor assay, and we visualized binding to the cell surface by fluorescence microscopy with a benzodiazepine analog conjugated to a fluorescein-labeled protein. Our studies also showed tha...

Analysis of Biological Interactions by Affinity Chromatography: Clinical and Pharmaceutical Applications.

Science.gov (United States)

Hage, David S

2017-06-01

The interactions between biochemical and chemical agents in the body are important in many clinical processes. Affinity chromatography and high-performance affinity chromatography (HPAC), in which a column contains an immobilized biologically related binding agent, are 2 methods that can be used to study these interactions. This review presents various approaches that can be used in affinity chromatography and HPAC to characterize the strength or rate of a biological interaction, the number and types of sites that are involved in this process, and the interactions between multiple solutes for the same binding agent. A number of applications for these methods are examined, with an emphasis on recent developments and high-performance affinity methods. These applications include the use of these techniques for fundamental studies of biological interactions, high-throughput screening of drugs, work with modified proteins, tools for personalized medicine, and studies of drug-drug competition for a common binding agent. The wide range of formats and detection methods that can be used with affinity chromatography and HPAC for examining biological interactions makes these tools attractive for various clinical and pharmaceutical applications. Future directions in the development of small-scale columns and the coupling of these methods with other techniques, such as mass spectrometry or other separation methods, should continue to increase the flexibility and ease with which these approaches can be used in work involving clinical or pharmaceutical samples. © 2016 American Association for Clinical Chemistry.

The role of antibody affinity and titre in immunity to Schistosoma mansoni following vaccination with highly irradiated cercariae

International Nuclear Information System (INIS)

Vignali, D.A.A.; Devey, M.E.; Bickle, Q.D.; Taylor, M.G.

1990-01-01

Sera from rabbits and rats vaccinated with highly irradiated cercariae of Schistosoma mansoni (VRabS, VRatS) were found to be of substantially higher affinity than sera from CBA mice vaccinated four times (4 x CVMS), single sex sera (SSS) or chronic infection sera (CIS). In contrast, immunoprecipitation studies demonstrated that sera from vaccinated LA mice (LVMS) recognized 125 I-labelled schistosomular surface antigens more intensely than sera from vaccinated HA mice (HVMS). However, peritoneal macrophages from HA and LA mice in the presence of HVMS, LVMS or 4 x CVMS, and naive macrophages activated in vitro with interferon-gamma (IFN-γ)/lipopolysaccharide (LPS) mediated comparable levels of schistosomula killing in vitro. The experiments described here provide evidence that the titre of antibody rather than its affinity may be a more critical factor in the development of optimal immunity to S. mansoni. (author)

N- and C-terminally truncated forms of glucose-dependent insulinotropic polypeptide are high-affinity competitive antagonists of the human GIP receptor

DEFF Research Database (Denmark)

Hansen, L S; Sparre-Ulrich, A H; Christensen, M.

2016-01-01

functions and pharmacological potential. GIP(1-30)NH2 is a naturally occurring truncation of GIP(1-42). Here we characterize eight N-terminal trrncations of human GIP(1-30)NH2 : GIP(2- to 9-30)NH2 . EXPERIMENTAL APPROACH: COS-7 cells were transiently transfected with the human GIP receptor and assessed...... displayed lower affinities (Ki 2.3-347 nM) with highest affinities of GIP(3-30)NH2 and (5-30)NH2 . Agonism was only observed for GIP(1-30)NH2 with an Emax on 100% of GIP(1-42) and GIP(2-30)NH2 (Emax 20%). GIP(2- to 9-30)NH2 displayed antagonism (IC50 12-450 nM) and right-shifts of the GIP(1-42)-response......, but superior antagonist GIP(3-30)NH2 , that together with GIP(5-30)NH2 were high-affinity competitive antagonist and thus may be suitable tool compounds for basic GIP research and future pharmacological interventions....

Covalent labeling of the beta-adrenergic ligand-binding site with para-(bromoacetamidyl)benzylcarazolol. A highly potent beta-adrenergic affinity label

International Nuclear Information System (INIS)

Dickinson, K.E.; Heald, S.L.; Jeffs, P.W.; Lefkowitz, R.J.; Caron, M.G.

1985-01-01

Para-(Bromoacetamidyl)benzylcarazolol (pBABC) was synthesized and found to be an extremely potent affinity label for beta-adrenergic receptors. Its interaction with mammalian (rabbit and hamster lung) and nonmammalian (turkey and frog erythrocyte) beta-adrenergic receptors was similar, displaying EC 50 values of 400-900 pM for inhibiting 125 I-cyanopindolol binding to these receptors. pBABC reduced the number of beta-adrenergic receptors in frog erythrocyte membranes, without any change in the affinity of the remaining sites for [ 125 I]iodocyanopindolol. pBABC has been radioiodinated. As assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, this affinity probe specifically labeled the beta-adrenergic peptide of a purified preparation of hamster lung, with high efficiency (approximately 40%) and with a pharmacological specificity characteristic of an interaction at the beta 2-adrenergic receptor ligand-binding site. Comparison of the proteolyzed products derived from purified receptor labeled with [ 125 I]pBABC and with the photoaffinity agent [ 125 I]p-azidobenzylcarazolol suggested that covalent labeling of the beta-adrenergic receptor by these probes occurs at similar domains of the beta-adrenergic receptor

APPLICATION OF IMMUNOGLOBULIN-BINDING PROTEINS A, G, L IN THE AFFINITY CHROMATOGRAPHY

Directory of Open Access Journals (Sweden)

О. V. Sviatenko

2014-04-01

Full Text Available Proteins A, G and L are native or recombinant proteins of microbial origin that bind to mammalian immunoglobulins. Preferably recombinant variants of proteins A, G, L are used in biotechnology for affinity sorbents production. Сomparative characteristics of proteins A, G, L and affinity sorbents on the basis of them, advantages and disadvantages of these proteins application as ligands in the affinity chromatography are done. Analysis of proteins A, G, L properties is presented. Binding specificities and affinities of these proteins differ between species and antibody subclass. Protein А has high affinity to human IgG1, IgG2, IgG4, mouse IgG2a, IgG2b, IgG3, goat and sheep IgG2, dog, cat, guinea pig, rabbit IgG. Protein G binds strongly to human, mouse, cow, goat, sheep and rabbit IgG. Protein L has ability of strong binding to immunoglobulin kappa-chains of human, mouse, rat and pig. Expediency of application of affinity chromatography with usage of sorbents on the basis of immobilized proteins A, G, L are shown for isolation and purification of antibodies different classes. Previously mentioned method is used as an alternative to conventional methods of protein purification, such as ion-exchange, hydrophobic interactions, metal affinity chromatography, ethanol precipitation due to simplicity in usage, possibility of one-step purification process, obtaining of proteins high level purity, multiuse at maintenance of proper storage and usage conditions. Affinity sorbents on the basis of immobilized proteins A, G, L are used not only for antibodies purification, but also for extraction of different antibodies fractions from blood serum.

In Vitro and In Vivo Evaluations of A High Affinity and Specificity Photoacoustic Nanoparticle Targeting to Cancer

DEFF Research Database (Denmark)

Ma, Lixin; Xu, Hang; Lee, Li Ean

oxide (SIO) nanoparticle as a potent cancer cell selective PA contrast agent, with a high binding affinity and selectivity to the gastrin releasing peptide receptor (GRPR) which is overexpressed in many human cancers including prostate cancer, breast cancer and small cell lung cancer etc.......Photoacoustic (PA) imaging uses a short-pulsed laser to create ultrasound signals for the high resolution acoustic imaging. The development of targeting PA contrast agents offers a unique opportunity to improve the early detection of cancer cells. This work aims to develop a silica coated iron...

Human metabolites of synthetic cannabinoids JWH-018 and JWH-073 bind with high affinity and act as potent agonists at cannabinoid type-2 receptors

International Nuclear Information System (INIS)

Rajasekaran, Maheswari; Brents, Lisa K.; Franks, Lirit N.; Moran, Jeffery H.; Prather, Paul L.

2013-01-01

K2 or Spice is an emerging drug of abuse that contains synthetic cannabinoids, including JWH-018 and JWH-073. Recent reports indicate that monohydroxylated metabolites of JWH-018 and JWH-073 retain high affinity and activity at cannabinoid type-1 receptors (CB 1 Rs), potentially contributing to the enhanced toxicity of K2 compared to marijuana. Since the parent compounds also bind to cannabinoid type-2 receptors (CB 2 Rs), this study investigated the affinity and intrinsic activity of JWH-018, JWH-073 and several monohydroxylated metabolites at human CB 2 Rs (hCB 2 Rs). The affinity of cannabinoids for hCB 2 Rs was determined by competition binding studies employing CHO-hCB 2 membranes. Intrinsic activity of compounds was assessed by G-protein activation and adenylyl cyclase (AC)-inhibition in CHO-hCB 2 cells. JWH-073, JWH-018 and several of their human metabolites exhibit nanomolar affinity and act as potent agonists at hCB 2 Rs. Furthermore, a major omega hydroxyl metabolite of JWH-073 (JWH-073-M5) binds to CB 2 Rs with 10-fold less affinity than the parent molecule, but unexpectedly, is equipotent in regulating AC-activity when compared to the parent molecule. Finally, when compared to CP-55,940 and Δ 9 -tetrahydrocannabinol (Δ 9 -THC), JWH-018, JWH-018-M5 and JWH-073-M5 require significantly less CB 2 R occupancy to produce similar levels of AC-inhibition, indicating that these compounds may more efficiently couple CB 2 Rs to AC than the well characterized cannabinoid agonists examined. These results indicate that JWH-018, JWH-073 and several major human metabolites of these compounds exhibit high affinity and demonstrate distinctive signaling properties at CB 2 Rs. Therefore, future studies examining pharmacological and toxicological properties of synthetic cannabinoids present in K2 products should consider potential actions of these drugs at both CB 1 and CB 2 Rs. - Highlights: • JWH-018 and JWH-073 are synthetic cannabinoids present in abused K2

Autoradiographic imaging and quantification of the high-affinity GHB binding sites in rodent brain using (3)H-HOCPCA

DEFF Research Database (Denmark)

Klein, A B; Bay, T; Villumsen, I S

2016-01-01

analogue, 3-hydroxycyclopent-1-enecarboxylic acid (HOCPCA) as a tritiated version ((3)H-HOCPCA) to radioactively label the specific GHB high-affinity binding site and gain further insight into the density, distribution and developmental profile of this protein. We show that, in low nanomolar concentrations...... brain development. Due to the high sensitivity of this radioligand, we were able to detect low levels of specific binding already at E15 in mouse brain, which increased progressively until adulthood. Collectively, we show that (3)H-HOCPCA is a highly sensitive radioligand, offering advantages over...

2017 Guralp Affinity Digitizer Evaluation.

Energy Technology Data Exchange (ETDEWEB)

Merchant, Bion J.

2018-03-01

Sandia National Laboratories has tested and evaluated two Guralp Affinity digitizers. The Affinity digitizers are intended to record sensor output for seismic and infrasound monitoring applications. The purpose of this digitizer evaluation is to measure the performance characteristics in such areas as power consumption, input impedance, sensitivity, full scale, self- noise, dynamic range, system noise, response, passband, and timing. The Affinity digitizers are being evaluated for potential use in the International Monitoring System (IMS) of the Comprehensive Nuclear Test-Ban-Treaty Organization (CTBTO).

Synthetic Receptors for the High-Affinity Recognition of O-GlcNAc Derivatives

NARCIS (Netherlands)

Rios, Pablo; Carter, Tom S; Mooibroek, Tiddo J; Crump, Matthew P; Lisbjerg, Micke; Pittelkow, Michael; Supekar, Nitin T; Boons, Geert-Jan|info:eu-repo/dai/nl/088245489; Davis, Anthony P

2016-01-01

The combination of a pyrenyl tetraamine with an isophthaloyl spacer has led to two new water-soluble carbohydrate receptors ("synthetic lectins"). Both systems show outstanding affinities for derivatives of N-acetylglucosamine (GlcNAc) in aqueous solution. One receptor binds the methyl glycoside

Biphasic regulation of development of the high-affinity saxitoxin receptor by innervation in rat skeletal muscle

International Nuclear Information System (INIS)

Sherman, S.J.; Catterall, W.A.

1982-01-01

Specific binding of 3 H-saxitoxin (STX) was used to quantitate the density of voltage-sensitive sodium channels in developing rat skeletal muscle. In adult triceps surae, a single class of sites with a KD . 2.9 nM and a density of 21 fmol/mg wet wt was detected. The density of these high-affinity sites increased from 2.0 fmol/mg wet wt to the adult value in linear fashion during days 2-25 after birth. Denervation of the triceps surae at day 11 or 17 reduced final saxitoxin receptor site density to 10.4 or 9.2 fmol/mg wet wt, respectively, without changing KD. Denervation of the triceps surae at day 5 did not alter the subsequent development of saxitoxin receptor sites during days 5-9 and accelerated the increase of saxitoxin receptor sites during days 9-13. After day 13, saxitoxin receptor development abruptly ceased and the density of saxitoxin receptor sites declined to 11 fmol/wg wet wt. These results show that the regulation of high-affinity saxitoxin receptor site density by innervation is biphasic. During the first phase, which is independent of continuing innervation, the saxitoxin receptor density increases to 47-57% of the adult level. After day 11, the second phase of development, which is dependent on continuing innervation, gives rise to the adult density of saxitoxin receptors

Two-parameter quantum affine algebra Ur,s(sln-circumflex), Drinfeld realization and quantum affine Lyndon basis

International Nuclear Information System (INIS)

Hu Naihong; Rosso, M.; Zhang Honglian

2006-12-01

We further find the defining structure of a two-parameter quantum affine algebra U r,s (sl n -circumflex) (n > 2) in the sense of Benkart-Witherspoon [BW1] after the work of [BGH1], [HS] and [BH], which turns out to be a Drinfeld double. Of more importance for the 'affine' cases is that we work out the compatible two-parameter version of the Drinfeld realization as a quantum affinization of U r,s (sl n ) and establish the Drinfeld isomorphism Theorem in the two-parameter setting via developing a new remarkable combinatorial approach - quantum 'affine' Lyndon basis with an explicit valid algorithm, based on the Drinfeld realization. (author)

Susceptibility to endotoxin induced uveitis is not reduced in mice deficient in BLT1, the high affinity leukotriene B4 receptor

OpenAIRE

Smith, J R; Subbarao, K; Franc, D T; Haribabu, B; Rosenbaum, J T

2004-01-01

Aim: To investigate the role of arachidonic acid derived chemotactic factor, LTB4, in the development of endotoxin induced uveitis (EIU), using mice deficient in the BLT1 gene which encodes the high affinity LTB4 receptor.

High-Affinity Sites Form an Interaction Network to Facilitate Spreading of the MSL Complex across the X Chromosome in Drosophila

NARCIS (Netherlands)

Ramírez, Fidel; Lingg, Thomas; Toscano, Sarah; Lam, Kin Chung; Georgiev, Plamen; Chung, Ho-Ryun; Lajoie, Bryan R; de Wit, Elzo; Zhan, Ye; de Laat, Wouter; Dekker, Job; Manke, Thomas; Akhtar, Asifa

2015-01-01

Dosage compensation mechanisms provide a paradigm to study the contribution of chromosomal conformation toward targeting and spreading of epigenetic regulators over a specific chromosome. By using Hi-C and 4C analyses, we show that high-affinity sites (HAS), landing platforms of the male-specific

IA-2 autoantibody affinity in children at risk for type 1 diabetes.

Science.gov (United States)

Krause, Stephanie; Chmiel, Ruth; Bonifacio, Ezio; Scholz, Marlon; Powell, Michael; Furmaniak, Jadwiga; Rees Smith, Bernard; Ziegler, Anette-G; Achenbach, Peter

2012-12-01

Autoantibodies to insulinoma-associated protein 2 (IA-2A) are associated with increased risk for type 1 diabetes. Here we examined IA-2A affinity and epitope specificity to assess heterogeneity in response intensity in relation to pathogenesis and diabetes risk in 50 children who were prospectively followed from birth. At first IA-2A appearance, affinity ranged from 10(7) to 10(11)L/mol and was high (>1.0×10(9)L/mol) in 41 (82%) children. IA-2A affinity was not associated with epitope specificity or HLA class II haplotype. On follow-up, affinity increased or remained high, and IA-2A were commonly against epitopes within the protein tyrosine phosphatase-like IA-2 domain and the homologue protein IA-2β. IA-2A were preceded or accompanied by other islet autoantibodies in 49 (98%) children, of which 34 progressed to diabetes. IA-2A affinity did not stratify diabetes risk. In conclusion, the IA-2A response in children is intense with rapid maturation against immunogenic epitopes and a strong association with diabetes development. Copyright © 2012 Elsevier Inc. All rights reserved.

Rational design of peptide affinity ligands for the purification of therapeutic enzymes.

Science.gov (United States)

Trasatti, John P; Woo, James; Ladiwala, Asif; Cramer, Steven; Karande, Pankaj

2018-04-25

Non-mAb biologics represent a growing class of therapeutics under clinical development. Although affinity chromatography is a potentially attractive approach for purification, the development of platform technologies, such as Protein A for mAbs, has been challenging due to the inherent chemical and structural diversity of these molecules. Here, we present our studies on the rapid development of peptide affinity ligands for the purification of biologics using a prototypical enzyme therapeutic in clinical use. Employing a suite of de novo rational and combinatorial design strategies we designed and screened a library of peptides on microarray platforms for their ability to bind to the target with high affinity and selectivity in cell culture fluid. Lead peptides were evaluated on resin in batch conditions and compared with a commercially available resin to evaluate their efficacy. Two lead candidates identified from microarray studies provided high binding capacity to the target while demonstrating high selectivity against culture contaminants and product variants compared to a commercial resin system. These findings provide a proof-of-concept for developing affinity peptide-based bioseparations processes for a target biologic. Peptide affinity ligand design and screening approaches presented in this work can also be easily translated to other biologics of interest. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 2018. © 2018 American Institute of Chemical Engineers.

Representations of affine Hecke algebras

CERN Document Server

Xi, Nanhua

1994-01-01

Kazhdan and Lusztig classified the simple modules of an affine Hecke algebra Hq (q E C*) provided that q is not a root of 1 (Invent. Math. 1987). Ginzburg had some very interesting work on affine Hecke algebras. Combining these results simple Hq-modules can be classified provided that the order of q is not too small. These Lecture Notes of N. Xi show that the classification of simple Hq-modules is essentially different from general cases when q is a root of 1 of certain orders. In addition the based rings of affine Weyl groups are shown to be of interest in understanding irreducible representations of affine Hecke algebras. Basic knowledge of abstract algebra is enough to read one third of the book. Some knowledge of K-theory, algebraic group, and Kazhdan-Lusztig cell of Cexeter group is useful for the rest

Inhibition of high affinity choline uptake by N-allyl-3-quinuclidinol

International Nuclear Information System (INIS)

Asermely, K.E.; O'Neill, J.J.

1986-01-01

The peripheral actions of N-allyl-3-quinuclidinol (N-Al-3-OHQ) on high affinity choline uptake (HAChU) on rat phrenic nerve diaphragm are described. Endplate regions (EPA) identified by the Koelle histochemical techniques for acetylcholinesterase, were dissected from adult rat hemidiaphragms and placed in cold Krebs solution (pH-7.35). All measurements of HAChU were at 37 0 C in buffers containing tritium choline (5 μM 0.124 μC/mmole) at intervals of 1, 2, 4, 8, 15 and 30 min. Tissues were washed 3x, digested in 1N NaOH and counted for tritium in Chaikoff's solution. All data are expressed as pmole Ch/g wet weight. Comparison between EPA and non-EPA tissue demonstrate HAChU and slow choline diffusion, respectively. Steady state is observed in 15 min. N-Al-3-OHQ produces 15% inhibition at 5 x 10 -5 M compared with 50% inhibition on brain synaptosomes. At 5 x 10 -4 M N-Al-3-OHQ, 30% inhibition is observed. Attempts to deplete ACh by pre-stimulation with high K + -ion (25 mM) were unsuccessful; tissue 3 H-choline uptake appeared to oscillate over a 30 min period

Inhibition of high affinity choline uptake by N-allyl-3-quinuclidinol

Energy Technology Data Exchange (ETDEWEB)

Asermely, K.E.; O' Neill, J.J.

1986-03-01

The peripheral actions of N-allyl-3-quinuclidinol (N-Al-3-OHQ) on high affinity choline uptake (HAChU) on rat phrenic nerve diaphragm are described. Endplate regions (EPA) identified by the Koelle histochemical techniques for acetylcholinesterase, were dissected from adult rat hemidiaphragms and placed in cold Krebs solution (pH-7.35). All measurements of HAChU were at 37/sup 0/C in buffers containing tritium choline (5 ..mu..M 0.124 ..mu..C/mmole) at intervals of 1, 2, 4, 8, 15 and 30 min. Tissues were washed 3x, digested in 1N NaOH and counted for tritium in Chaikoff's solution. All data are expressed as pmole Ch/g wet weight. Comparison between EPA and non-EPA tissue demonstrate HAChU and slow choline diffusion, respectively. Steady state is observed in 15 min. N-Al-3-OHQ produces 15% inhibition at 5 x 10/sup -5/ M compared with 50% inhibition on brain synaptosomes. At 5 x 10/sup -4/ M N-Al-3-OHQ, 30% inhibition is observed. Attempts to deplete ACh by pre-stimulation with high K/sup +/-ion (25 mM) were unsuccessful; tissue /sup 3/H-choline uptake appeared to oscillate over a 30 min period.

The role of antibody affinity and titre in immunity to Schistosoma mansoni following vaccination with highly irradiated cercariae

Energy Technology Data Exchange (ETDEWEB)

Vignali, D.A.A.; Devey, M.E.; Bickle, Q.D.; Taylor, M.G. (London School of Hygiene and Tropical Medicine (UK))

1990-02-01

Sera from rabbits and rats vaccinated with highly irradiated cercariae of Schistosoma mansoni (VRabS, VRatS) were found to be of substantially higher affinity than sera from CBA mice vaccinated four times (4 x CVMS), single sex sera (SSS) or chronic infection sera (CIS). In contrast, immunoprecipitation studies demonstrated that sera from vaccinated LA mice (LVMS) recognized {sup 125}I-labelled schistosomular surface antigens more intensely than sera from vaccinated HA mice (HVMS). However, peritoneal macrophages from HA and LA mice in the presence of HVMS, LVMS or 4 x CVMS, and naive macrophages activated in vitro with interferon-gamma (IFN-{gamma})/lipopolysaccharide (LPS) mediated comparable levels of schistosomula killing in vitro. The experiments described here provide evidence that the titre of antibody rather than its affinity may be a more critical factor in the development of optimal immunity to S. mansoni. (author).

High-affinity small molecule-phospholipid complex formation: binding of siramesine to phosphatidicacid

DEFF Research Database (Denmark)

Khandelia, Himanshu

2008-01-01

, comparable to the affinities for the binding of small molecule ligands to proteins, was measured for phosphatidic acid (PA, mole fraction of XPA ) 0.2 in phosphatidylcholine vesicles), yielding a molecular partition coefficient of 240 ( 80 × 106. An MD simulation on the siramesine:PA interaction...

Antisymmetric tensor generalizations of affine vector fields.

Science.gov (United States)

Houri, Tsuyoshi; Morisawa, Yoshiyuki; Tomoda, Kentaro

2016-02-01

Tensor generalizations of affine vector fields called symmetric and antisymmetric affine tensor fields are discussed as symmetry of spacetimes. We review the properties of the symmetric ones, which have been studied in earlier works, and investigate the properties of the antisymmetric ones, which are the main theme in this paper. It is shown that antisymmetric affine tensor fields are closely related to one-lower-rank antisymmetric tensor fields which are parallelly transported along geodesics. It is also shown that the number of linear independent rank- p antisymmetric affine tensor fields in n -dimensions is bounded by ( n + 1)!/ p !( n - p )!. We also derive the integrability conditions for antisymmetric affine tensor fields. Using the integrability conditions, we discuss the existence of antisymmetric affine tensor fields on various spacetimes.

Insights from the Fungus Fusarium oxysporum Point to High Affinity Glucose Transporters as Targets for Enhancing Ethanol Production from Lignocellulose

Science.gov (United States)

Ali, Shahin S.; Nugent, Brian; Mullins, Ewen; Doohan, Fiona M.

2013-01-01

Ethanol is the most-widely used biofuel in the world today. Lignocellulosic plant biomass derived from agricultural residue can be converted to ethanol via microbial bioprocessing. Fungi such as Fusarium oxysporum can simultaneously saccharify straw to sugars and ferment sugars to ethanol. But there are many bottlenecks that need to be overcome to increase the efficacy of microbial production of ethanol from straw, not least enhancement of the rate of fermentation of both hexose and pentose sugars. This research tested the hypothesis that the rate of sugar uptake by F. oxysporum would enhance the ethanol yields from lignocellulosic straw and that high affinity glucose transporters can enhance ethanol yields from this substrate. We characterized a novel hexose transporter (Hxt) from this fungus. The F. oxysporum Hxt represents a novel transporter with homology to yeast glucose signaling/transporter proteins Rgt2 and Snf3, but it lacks their C-terminal domain which is necessary for glucose signalling. Its expression level decreased with increasing glucose concentration in the medium and in a glucose uptake study the Km(glucose) was 0.9 mM, which indicated that the protein is a high affinity glucose transporter. Post-translational gene silencing or over expression of the Hxt in F. oxysporum directly affected the glucose and xylose transport capacity and ethanol yielded by F. oxysporum from straw, glucose and xylose. Thus we conclude that this Hxt has the capacity to transport both C5 and C6 sugars and to enhance ethanol yields from lignocellulosic material. This study has confirmed that high affinity glucose transporters are ideal candidates for improving ethanol yields from lignocellulose because their activity and level of expression is high in low glucose concentrations, which is very common during the process of consolidated processing. PMID:23382943

Insights from the fungus Fusarium oxysporum point to high affinity glucose transporters as targets for enhancing ethanol production from lignocellulose.

Directory of Open Access Journals (Sweden)

Shahin S Ali

Full Text Available Ethanol is the most-widely used biofuel in the world today. Lignocellulosic plant biomass derived from agricultural residue can be converted to ethanol via microbial bioprocessing. Fungi such as Fusarium oxysporum can simultaneously saccharify straw to sugars and ferment sugars to ethanol. But there are many bottlenecks that need to be overcome to increase the efficacy of microbial production of ethanol from straw, not least enhancement of the rate of fermentation of both hexose and pentose sugars. This research tested the hypothesis that the rate of sugar uptake by F. oxysporum would enhance the ethanol yields from lignocellulosic straw and that high affinity glucose transporters can enhance ethanol yields from this substrate. We characterized a novel hexose transporter (Hxt from this fungus. The F. oxysporum Hxt represents a novel transporter with homology to yeast glucose signaling/transporter proteins Rgt2 and Snf3, but it lacks their C-terminal domain which is necessary for glucose signalling. Its expression level decreased with increasing glucose concentration in the medium and in a glucose uptake study the Km((glucose was 0.9 mM, which indicated that the protein is a high affinity glucose transporter. Post-translational gene silencing or over expression of the Hxt in F. oxysporum directly affected the glucose and xylose transport capacity and ethanol yielded by F. oxysporum from straw, glucose and xylose. Thus we conclude that this Hxt has the capacity to transport both C5 and C6 sugars and to enhance ethanol yields from lignocellulosic material. This study has confirmed that high affinity glucose transporters are ideal candidates for improving ethanol yields from lignocellulose because their activity and level of expression is high in low glucose concentrations, which is very common during the process of consolidated processing.

Manifolds with integrable affine shape operator

Directory of Open Access Journals (Sweden)

Daniel A. Joaquín

2005-05-01

Full Text Available This work establishes the conditions for the existence of vector fields with the property that theirs covariant derivative, with respect to the affine normal connection, be the affine shape operatorS in hypersurfaces. Some results are obtained from this property and, in particular, for some kind of affine decomposable hypersurfaces we explicitely get the actual vector fields.

Thermodynamic basis for engineering high-affinity, high-specificity binding-induced DNA clamp nanoswitches.

Science.gov (United States)

Idili, Andrea; Plaxco, Kevin W; Vallée-Bélisle, Alexis; Ricci, Francesco

2013-12-23

Naturally occurring chemoreceptors almost invariably employ structure-switching mechanisms, an observation that has inspired the use of biomolecular switches in a wide range of artificial technologies in the areas of diagnostics, imaging, and synthetic biology. In one mechanism for generating such behavior, clamp-based switching, binding occurs via the clamplike embrace of two recognition elements onto a single target molecule. In addition to coupling recognition with a large conformational change, this mechanism offers a second advantage: it improves both affinity and specificity simultaneously. To explore the physics of such switches we have dissected here the thermodynamics of a clamp-switch that recognizes a target DNA sequence through both Watson-Crick base pairing and triplex-forming Hoogsteen interactions. When compared to the equivalent linear DNA probe (which relies solely on Watson-Crick interactions), the extra Hoogsteen interactions in the DNA clamp-switch increase the probe's affinity for its target by ∼0.29 ± 0.02 kcal/mol/base. The Hoogsteen interactions of the clamp-switch likewise provide an additional specificity check that increases the discrimination efficiency toward a single-base mismatch by 1.2 ± 0.2 kcal/mol. This, in turn, leads to a 10-fold improvement in the width of the "specificity window" of this probe relative to that of the equivalent linear probe. Given these attributes, clamp-switches should be of utility not only for sensing applications but also, in the specific field of DNA nanotechnology, for applications calling for a better control over the building of nanostructures and nanomachines.

Constraints on the affinity term for modeling long-term glass dissolution rates

International Nuclear Information System (INIS)

Bourcier, W.L.; Carroll, S.A.; Phillips, B.L.

1993-11-01

Predictions of long-term glass dissolution rates are highly dependent on the form of the affinity term in the rate expression. Analysis of the quantitative effect of saturation state on glass dissolution rate for CSG glass (a simple analog of SRL-165 glass), shows that a simple (1-Q/K) affinity term does not match experimental results. Our data at 100 degree C show that the data is better fit by an affinity term having the form (1 - (Q/K) 1 /σ) where σ = 10

Identification of an adeno-associated virus binding epitope for AVB sepharose affinity resin

Directory of Open Access Journals (Sweden)

Qiang Wang

Full Text Available Recent successes of adeno-associated virus (AAV–based gene therapy have created a demand for large-scale AAV vector manufacturing and purification techniques for use in clinical trials and beyond. During the development of purification protocols for rh.10, hu.37, AAV8, rh.64R1, AAV3B, and AAV9 vectors, based on a widely used affinity resin, AVB sepharose (GE, we found that, under the same conditions, different serotypes have different affinities to the resin, with AAV3B binding the best and AAV9 the poorest. Further analysis revealed a surface-exposed residue (amino acid number 665 in AAV8 VP1 numbering differs between the high-affinity AAV serotypes (serine in AAV3B, rh.10, and hu.37 and the low-affinity ones (asparagine in AAV8, rh.64R1, and AAV9. The residue locates within a surface-exposed, variable epitope flanked by highly conserved residues. The substitution of the epitope in AAV8, rh.64R1, and AAV9 with the corresponding epitope of AAV3B (SPAKFA resulted in greatly increased affinity to AVB sepharose with no reduction in the vectors’ in vitro potency. The presence of the newly identified AVB-binding epitope will be useful for affinity resin selection for the purification of novel AAV serotypes. It also suggests the possibility of vector engineering to yield a universal affinity chromatography purification method for multiple AAV serotypes.

Light adaptation and the evolution of vertebrate photoreceptors.

Science.gov (United States)

Morshedian, Ala; Fain, Gordon L

2017-07-15

Lamprey are cyclostomes, a group of vertebrates that diverged from lines leading to jawed vertebrates (including mammals) in the late Cambrian, 500 million years ago. It may therefore be possible to infer properties of photoreceptors in early vertebrate progenitors by comparing lamprey to other vertebrates. We show that lamprey rods and cones respond to light much like rods and cones in amphibians and mammals. They operate over a similar range of light intensities and adapt to backgrounds and bleaches nearly identically. These correspondences are pervasive and detailed; they argue for the presence of rods and cones very early in the evolution of vertebrates with properties much like those of rods and cones in existing vertebrate species. The earliest vertebrates were agnathans - fish-like organisms without jaws, which first appeared near the end of the Cambrian radiation. One group of agnathans became cyclostomes, which include lamprey and hagfish. Other agnathans gave rise to jawed vertebrates or gnathostomes, the group including all other existing vertebrate species. Because cyclostomes diverged from other vertebrates 500 million years ago, it may be possible to infer some of the properties of the retina of early vertebrate progenitors by comparing lamprey to other vertebrates. We have previously shown that rods and cones in lamprey respond to light much like photoreceptors in other vertebrates and have a similar sensitivity. We now show that these affinities are even closer. Both rods and cones adapt to background light and to bleaches in a manner almost identical to other vertebrate photoreceptors. The operating range in darkness is nearly the same in lamprey and in amphibian or mammalian rods and cones; moreover background light shifts response-intensity curves downward and to the right over a similar range of ambient intensities. Rods show increment saturation at about the same intensity as mammalian rods, and cones never saturate. Bleaches decrease

Affine LIBOR Models with Multiple Curves

DEFF Research Database (Denmark)

Grbac, Zorana; Papapantoleon, Antonis; Schoenmakers, John

2015-01-01

are specified following the methodology of the affine LIBOR models and are driven by the wide and flexible class of affine processes. The affine property is preserved under forward measures, which allows us to derive Fourier pricing formulas for caps, swaptions, and basis swaptions. A model specification...... with dependent LIBOR rates is developed that allows for an efficient and accurate calibration to a system of caplet prices....

A Novel Fast and Robust Binary Affine Invariant Descriptor for Image Matching

Directory of Open Access Journals (Sweden)

Xiujie Qu

2014-01-01

Full Text Available As the current binary descriptors have disadvantages of high computational complexity, no affine invariance, and the high false matching rate with viewpoint changes, a new binary affine invariant descriptor, called BAND, is proposed. Different from other descriptors, BAND has an irregular pattern, which is based on local affine invariant region surrounding a feature point, and it has five orientations, which are obtained by LBP effectively. Ultimately, a 256 bits binary string is computed by simple random sampling pattern. Experimental results demonstrate that BAND has a good matching result in the conditions of rotating, image zooming, noising, lighting, and small-scale perspective transformation. It has better matching performance compared with current mainstream descriptors, while it costs less time.

Affinity Programs and the Real Estate Brokerage Industry

OpenAIRE

G Stacy Sirmans; David A. Macpherson

2001-01-01

This study surveys active real estate brokers obtaining information on involvement in affinity programs and referral/relocation networks. Some results regarding affinity involvement are: (a) 13% of respondents reported affinity affilliations, 75% reported no affiliations, and 12% indicated plans to become involved within the next year; (b) about half having affinity affiliations were involved with 2-4 groups; (c) affinity relationships were most often with membership organizations, corporatio...

Human metabolites of synthetic cannabinoids JWH-018 and JWH-073 bind with high affinity and act as potent agonists at cannabinoid type-2 receptors

Energy Technology Data Exchange (ETDEWEB)

Rajasekaran, Maheswari; Brents, Lisa K.; Franks, Lirit N. [Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock, AR 72205 (United States); Moran, Jeffery H. [Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock, AR 72205 (United States); Arkansas Department of Public Health, Public Health Laboratory, Little Rock, AR 72205 (United States); Prather, Paul L., E-mail: pratherpaull@uams.edu [Department of Pharmacology and Toxicology, University of Arkansas for Medical Sciences, Little Rock, AR 72205 (United States)

2013-06-01

K2 or Spice is an emerging drug of abuse that contains synthetic cannabinoids, including JWH-018 and JWH-073. Recent reports indicate that monohydroxylated metabolites of JWH-018 and JWH-073 retain high affinity and activity at cannabinoid type-1 receptors (CB{sub 1}Rs), potentially contributing to the enhanced toxicity of K2 compared to marijuana. Since the parent compounds also bind to cannabinoid type-2 receptors (CB{sub 2}Rs), this study investigated the affinity and intrinsic activity of JWH-018, JWH-073 and several monohydroxylated metabolites at human CB{sub 2}Rs (hCB{sub 2}Rs). The affinity of cannabinoids for hCB{sub 2}Rs was determined by competition binding studies employing CHO-hCB{sub 2} membranes. Intrinsic activity of compounds was assessed by G-protein activation and adenylyl cyclase (AC)-inhibition in CHO-hCB{sub 2} cells. JWH-073, JWH-018 and several of their human metabolites exhibit nanomolar affinity and act as potent agonists at hCB{sub 2}Rs. Furthermore, a major omega hydroxyl metabolite of JWH-073 (JWH-073-M5) binds to CB{sub 2}Rs with 10-fold less affinity than the parent molecule, but unexpectedly, is equipotent in regulating AC-activity when compared to the parent molecule. Finally, when compared to CP-55,940 and Δ{sup 9}-tetrahydrocannabinol (Δ{sup 9}-THC), JWH-018, JWH-018-M5 and JWH-073-M5 require significantly less CB{sub 2}R occupancy to produce similar levels of AC-inhibition, indicating that these compounds may more efficiently couple CB{sub 2}Rs to AC than the well characterized cannabinoid agonists examined. These results indicate that JWH-018, JWH-073 and several major human metabolites of these compounds exhibit high affinity and demonstrate distinctive signaling properties at CB{sub 2}Rs. Therefore, future studies examining pharmacological and toxicological properties of synthetic cannabinoids present in K2 products should consider potential actions of these drugs at both CB{sub 1} and CB{sub 2}Rs. - Highlights: • JWH-018

Reconstitution of high affinity α2 adrenergic agonist binding by fusion with a pertussis toxin substrate

International Nuclear Information System (INIS)

Kim, M.H.; Neubig, R.R.

1986-01-01

High affinity α 2 adrenergic agonist binding is thought to occur via a coupling of the α 2 receptor with N/sub i/, the inhibitory guanyl nucleotide binding protein. Human platelet membranes pretreated at pH 11.5 exhibit a selective inactivation of agonist binding and N/sub i/. To further study the mechanism of agonist binding, alkali treated membranes (ATM) were mixed with membranes pretreated with 10 μM phenoxybenzamine to block α 2 receptors (POB-M). The combined membrane pellet was incubated in 50% polyethylene glycol (PEG) to promote membrane-membrane fusion and assayed for binding to the α 2 agonist [ 3 H]UK 14,304 (UK) and the antagonist [ 3 H] yohimbine. PEG treatment resulted in a 2-4 fold enhancement of UK binding whereas yohimbine binding was unchanged. No enhancement of UK binding was observed in the absence of PEG treatment. The reconstitution was dependent on the addition of POB-M. They found that a 1:1 ratio of POB-M:ATM was optimal. Reconstituted binding was inhibited by GppNHp. Fusion of rat C6 glioma cell membranes, which do not contain α 2 receptors, also enhanced agonist binding to ATM. Fusion of C6 membranes from cells treated with pertussis toxin did not enhance [ 3 H] UK binding. These data show that a pertussis toxin sensitive membrane component, possibly N/sub i/, can reconstitute high affinity α 2 agonist binding

Development of 99mTc labelled somatostatin analogues with high affinity for somatostatin receptors

International Nuclear Information System (INIS)

Maina, T.; Nock, B.; Nicolopoulou, A.; Tsipra, C.; Poppe, M.; Chiotellis, E.

2001-01-01

A recent development in oncology involves the use of metabolically stabilized peptide hormone analogues labelled with metallic radionuclides for the diagnosis or therapy of malignant disease. This approach was successfully applied for the first time in the visualization of somatostatin positive tumours and their metastases with 111 In DTPA-octreotide. In an effort to obtain a 99m Tc somatostatin receptor affine radioligand we describe herein the synthesis, radiochemistry and preliminary biological evaluation of two novel 99m Tc labelled somatostatin analogues, N 4 -TOC and N 4 -RC-160. In these compounds a tetraamine bifunctional unit was covalently attached to the N-terminal (D)Phe 1 of the peptide chain using Boc-protection strategies. The peptide conjugates were purified by high performance liquid chromatography (HPLC) and characterized by UV/Vis and ES-MS spectroscopies. As revealed by HPLC, 99m Tc labelling was quantitative under mild conditions, leading to a single 99m Tc species in high specific activities. Affinity of 99m Tc N 4 -TOC for the somatostatin receptor, as determined by in vitro binding assays in rat brain cortex membranes, was found unaffected by the presence of the bulky metal chelate. The binding properties of 99m Tc N 4 -RC-160 could not be determined by this assay due to an extremely high non-specific binding of this radioligand, and will be shortly investigated by other methods. Tissue distribution in healthy mice revealed that 99m Tc N 4 -TOC is clearing mainly through the kidneys and the urinary tract whereas 99m Tc N 4 -RC-160 shows a high accumulation in the liver as a result of its lipophilicity. Analysis of urine samples by HPLC showed that 99m Tc N 4 -TOC is excreted integer from the body of mice, while 99m Tc N 4 -RC-160 is totally transformed to an unidentified hydrophilic metabolite in vivo. The location of this metabolism is currently investigated. In vivo blocking experiments using animals pre-treated with 50 μg octreotide

Experimental and theoretical binding affinity between polyvinylpolypyrrolidone and selected phenolic compounds from food matrices.

Science.gov (United States)

Durán-Lara, Esteban F; López-Cortés, Xaviera A; Castro, Ricardo I; Avila-Salas, Fabián; González-Nilo, Fernando D; Laurie, V Felipe; Santos, Leonardo S

2015-02-01

Polyvinylpolypyrrolidone (PVPP) is a fining agent, widely used in winemaking and brewing, whose mode of action in removing phenolic compounds has not been fully characterised. The aim of this study was to evaluate the experimental and theoretical binding affinity of PVPP towards six phenolic compounds representing different types of phenolic species. The interaction between PVPP and phenolics was evaluated in model solutions, where hydroxyl groups, hydrophobic bonding and steric hindrance were characterised. The results of the study indicated that PVPP exhibits high affinity for quercetin and catechin, moderate affinity for epicatechin, gallic acid and lower affinity for 4-methylcatechol and caffeic acid. The affinity has a direct correlation with the hydroxylation degree of each compound. The results show that the affinity of PVPP towards phenols is related with frontier orbitals. This work demonstrates a direct correlation between the experimental affinity and the interaction energy calculations obtained through computational chemistry methods. Copyright © 2014 Elsevier Ltd. All rights reserved.

Using Affinity Diagrams to Evaluate Interactive Prototypes

DEFF Research Database (Denmark)

Lucero, Andrés

2015-01-01

our particular use of affinity diagramming in prototype evaluations. We reflect on a decade’s experience using affinity diagramming across a number of projects, both in industry and academia. Our affinity diagramming process in interaction design has been tailored and consists of four stages: creating...

Fundamental and practical studies on high-performance liquid affinity chromatography of biopolymers with novel stationary phases

Energy Technology Data Exchange (ETDEWEB)

Bacolod, M.D.

1992-01-01

Rigid microparticulate stationary phases having surface-bound metal chelating functions were developed and evaluated in high performance metal chelate affinity chromatography of proteins. Silica- and polystyrene-divinylbenzene-based metal chelate sorbents were produced in wide pore and in non-porous type of column packings. A major effort has been placed on development of non-porous highly crosslinked polystyrene-divinylbenzene (PSDVB). These PSDVB microparticles were produced by a two-step swelling polymerization, and exhibited excellent mechanical strength over a wide range of flow-rates and composition used in high performance liquid chromatography (HPLC). Simple and reproducible hydrophilic coatings were developed for the surface modification of hydrophobic PSDVB supports. A tetradentate metal chelating ligand, ethylenediamine-N, N[prime]-diacetic acid (EDDA), was covalently bound to the surface of the various supports. Sorbents having iminodiacetic acid (IDA) metal chelating functions were also evaluated. The hydrophilic character and surface coverage of various stationary phases were assessed chromatographically. Studies concerning the effects of eluent pH as well as the nature and concentration of salts on retention and selectivity with different metal chelate stationary phases having various immobilized metal ions were carried out. Elution schemes were developed for rapid separation of proteins in metal chelate affinity chromatography. EDDA stationary phases in metal forms can be viewed as complementary to IDA stationary phases since they afforded different selectivity and retentivity toward proteins. Hydrophilic PSDVB could be functionalized with IDA or EDDA metal chelating ligands or lectins. The non-porous metal chelate stationary phases afforded rapid separation of proteins by the development of multiple gradient systems, which permitted higher column peak capacity, enabling the separation of a greater number of proteins in a single chromatographic run.

Inhibition of Enterococcus faecium adherence to collagen by antibodies against high-affinity binding subdomains of Acm.

Science.gov (United States)

Nallapareddy, Sreedhar R; Sillanpää, Jouko; Ganesh, Vannakambadi K; Höök, Magnus; Murray, Barbara E

2007-06-01

Strains of Enterococcus faecium express a cell wall-anchored protein, Acm, which mediates adherence to collagen. Here, we (i) identify the minimal and high-affinity binding subsegments of Acm and (ii) show that anti-Acm immunoglobulin Gs (IgGs) purified against these subsegments reduced E. faecium TX2535 strain collagen adherence up to 73 and 50%, respectively, significantly more than the total IgGs against the full-length Acm A domain (28%) (P Acm adherence with functional subsegment-specific antibodies raises the possibility of their use as therapeutic or prophylactic agents.

The Cutting Edge of Affinity Electrophoresis Technology.

Science.gov (United States)

Kinoshita, Eiji; Kinoshita-Kikuta, Emiko; Koike, Tohru

2015-03-18

Affinity electrophoresis is an important technique that is widely used to separate and analyze biomolecules in the fields of biology and medicine. Both quantitative and qualitative information can be gained through affinity electrophoresis. Affinity electrophoresis can be applied through a variety of strategies, such as mobility shift electrophoresis, charge shift electrophoresis or capillary affinity electrophoresis. These strategies are based on changes in the electrophoretic patterns of biological macromolecules that result from interactions or complex-formation processes that induce changes in the size or total charge of the molecules. Nucleic acid fragments can be characterized through their affinity to other molecules, for example transcriptional factor proteins. Hydrophobic membrane proteins can be identified by means of a shift in the mobility induced by a charged detergent. The various strategies have also been used in the estimation of association/disassociation constants. Some of these strategies have similarities to affinity chromatography, in that they use a probe or ligand immobilized on a supported matrix for electrophoresis. Such methods have recently contributed to profiling of major posttranslational modifications of proteins, such as glycosylation or phosphorylation. Here, we describe advances in analytical techniques involving affinity electrophoresis that have appeared during the last five years.

Affinity-based, biophysical methods to detect and analyze ligand binding to recombinant proteins: matching high information content with high throughput.

Science.gov (United States)

Holdgate, Geoff A; Anderson, Malcolm; Edfeldt, Fredrik; Geschwindner, Stefan

2010-10-01

Affinity-based technologies have become impactful tools to detect, monitor and characterize molecular interactions using recombinant target proteins. This can aid the understanding of biological function by revealing mechanistic details, and even more importantly, enables the identification of new improved ligands that can modulate the biological activity of those targets in a desired fashion. The selection of the appropriate technology is a key step in that process, as each one of the currently available technologies offers a characteristic type of biophysical information about the ligand-binding event. Alongside the indisputable advantages of each of those technologies they naturally display diverse restrictions that are quite frequently related to the target system to be studied but also to the affinity, solubility and molecular size of the ligands. This paper discusses some of the theoretical and experimental aspects of the most common affinity-based methods, what type of information can be gained from each one of those approaches, and what requirements as well as limitations are expected from working with recombinant proteins on those platforms and how those can be optimally addressed.

Single-step affinity purification for fungal proteomics.

Science.gov (United States)

Liu, Hui-Lin; Osmani, Aysha H; Ukil, Leena; Son, Sunghun; Markossian, Sarine; Shen, Kuo-Fang; Govindaraghavan, Meera; Varadaraj, Archana; Hashmi, Shahr B; De Souza, Colin P; Osmani, Stephen A

2010-05-01

A single-step protein affinity purification protocol using Aspergillus nidulans is described. Detailed protocols for cell breakage, affinity purification, and depending on the application, methods for protein release from affinity beads are provided. Examples defining the utility of the approaches, which should be widely applicable, are included.

The production of high affinity monoclonal antibodies to human growth hormone

International Nuclear Information System (INIS)

Stuart, M.C.; Walichnowski, C.M.; Hussain, S.; Underwood, P.A.; Harman, D.F.; Rathjen, D.A.; Sturmer, S.R. von

1983-01-01

The primary aim of this work was to produce specific monoclonal antibodies to human growth hormone (hGH) for use in a diagnostic RIA of hGH levels in serum. Three different schedules were used for immunization of BALB/c mice and the splenocytes from each mouse were fused with myeloma cells Sp 2/0 Ag 14. Each fusion resulted in the production of hundreds of hybridomas secreting hGH-directed antibodies. Six antibodies have been fully characterized and have been grouped into pairs which recognize 3 different epitopes on the hGH molecule. One pair exhibits no cross reaction with the structurally related placental hormone, human placental lactogen (hPL), a second pair has low cross reaction with hPL (1.6-3%) and a third pair reacts equally well with hGH and hPL indicating binding to a common epitope in the 2 molecules. The highest affinity antibody, 74/6, which has an affinity constant of 4.4x10 10 l/mol and 3% cross-reactivity with hPL, has been used to establish a RIA for serum hGH measurements. Evidence is provided that hGH levels measured in this assay correlate well with those obtained in a conventional rabbit antiserum assay. (Auth.)

A pharmacological profile of the high-affinity GluK5 kainate receptor

DEFF Research Database (Denmark)

Møllerud, Stine; Kastrup, Jette Sandholm Jensen; Pickering, Darryl S

2016-01-01

-hydroxyisoxazol-4-yl)propionate (ATPA), dihydrokainate and (2 S,4 R)−4-methyl-glutamate (SYM2081) have higher affinity at GluK3 compared to GluK5. Since some studies have indicated that GluK5 is associated with various diseases in the central nervous system (e.g. schizophrenia, temporal lobe epilepsy, bipolar...

Knock-down of a tonoplast localized low-affinity nitrate transporter OsNPF7.2 affects rice growth under high nitrate ssupply

Directory of Open Access Journals (Sweden)

Rui Hu

2016-10-01

Full Text Available The large nitrate transporter 1/peptide transporter family (NPF has been shown to transport diverse substrates, including nitrate, amino acids, peptides, phytohormones, and glucosinolates. However, the rice (Oryza sativa root-specific expressed member OsNPF7.2 has not been characterized. Here, our data show that OsNPF7.2 is a tonoplast localized low-affinity nitrate transporter, and affects rice growth under high nitrate supply. The expression analysis showed that OsNPF7.2 was mainly expressed in the elongation and maturation zones of roots, especially in the root sclerenchyma, cortex and stele. It was also induced by high concentrations of nitrate. Subcellular localization analysis showed that OsNPF7.2 was localized on the tonoplast of large and small vacuoles. Heterogenous expression in Xenopus laevis oocytes suggested that OsNPF7.2 was a low-affinity nitrate transporter. Knock-down of OsNPF7.2 retarded rice growth under high concentrations of nitrate. Therefore, we deduce that OsNPF7.2 plays a role in intracellular allocation of nitrate in roots, and thus influences rice growth under high nitrate supply.

Plasma-polymerized films providing selective affinity to the polarity of vaporized organic solvents

International Nuclear Information System (INIS)

Akimoto, Takuo; Ikeshita, Yusuke; Terashima, Ryo; Karube, Isao

2009-01-01

Plasma-polymerized films (PPFs) were fabricated as recognition membranes for a vapor-sensing device, and their affinity to vaporized organic solvents was evaluated with surface plasmon resonance. The affinity we intended to create is the selective sorption of the vaporized organic solvents depending on their polarity. For this purpose, acetonitrile, ethylenediamine (EDA), styrene, hexamethyldisiloxane (HMDSO), and hexamethyldisilazane were used to fabricate PPFs. Vaporized methanol, ethanol, and 1-propanol were used as high-polar solvents to be analyzed. Hexane, toluene, and p-xylene were used as low-polar solvents. As a result, the HMDSO-PPF with 97.3 o of contact angle was found to provide affinity to the low-polar solvents. In contrast, the EDA-PPF with 7.1 o of contact angle provided affinity to the high-polar solvents. Observations of the surface morphology of the HMDSO- and EDA-PPFs with a scanning electron microscope revealed that they are composed of nano-scale islands.

Locked nucleic acid (LNA): High affinity targeting of RNA for diagnostics and therapeutics

DEFF Research Database (Denmark)

Kauppinen, S.; Vester, Birte; Wengel, Jesper

2005-01-01

Locked nucleic acid (LNA) is a nucleic acid analogue containing one or more LNA nucleotide monomers with a bicyclic furanose unit locked in an RNA mimicking sugar conformation. This conformational restriction results in unprecedented hybridization affinity towards complementary single stranded RN...

Connections between quantized affine algebras and superalgebras

International Nuclear Information System (INIS)

Zhang, R.B.

1992-08-01

Every affine superalgebra with a symmetrizable Cartan matrix is closely related to an ordinary affine algebra with the same Cartan matrix. It is shown that the quantum supergroup associated with the former is essentially isomorphic to the quantum group associated with the latter in an appropriate class of representations. At the classical level, each integrable irreducible highest weight representation of the affine superalgebra has a corresponding irreducible representation of the affine algebra, which has the same weight space decomposition. (author). 5 refs, 3 tabs

High affinity soluble ILT2 receptor: a potent inhibitor of CD8(+) T cell activation.

Science.gov (United States)

Moysey, Ruth K; Li, Yi; Paston, Samantha J; Baston, Emma E; Sami, Malkit S; Cameron, Brian J; Gavarret, Jessie; Todorov, Penio; Vuidepot, Annelise; Dunn, Steven M; Pumphrey, Nicholas J; Adams, Katherine J; Yuan, Fang; Dennis, Rebecca E; Sutton, Deborah H; Johnson, Andy D; Brewer, Joanna E; Ashfield, Rebecca; Lissin, Nikolai M; Jakobsen, Bent K

2010-12-01

Using directed mutagenesis and phage display on a soluble fragment of the human immunoglobulin super-family receptor ILT2 (synonyms: LIR1, MIR7, CD85j), we have selected a range of mutants with binding affinities enhanced by up to 168,000-fold towards the conserved region of major histocompatibility complex (MHC) class I molecules. Produced in a dimeric form, either by chemical cross-linking with bivalent polyethylene glycol (PEG) derivatives or as a genetic fusion with human IgG Fc-fragment, the mutants exhibited a further increase in ligand-binding strength due to the avidity effect, with resident half-times (t(1/2)) on the surface of MHC I-positive cells of many hours. The novel compounds antagonized the interaction of CD8 co-receptor with MHC I in vitro without affecting the peptide-specific binding of T-cell receptors (TCRs). In both cytokine-release assays and cell-killing experiments the engineered receptors inhibited the activation of CD8(+) cytotoxic T lymphocytes (CTLs) in the presence of their target cells, with subnanomolar potency and in a dose-dependent manner. As a selective inhibitor of CD8(+) CTL responses, the engineered high affinity ILT2 receptor presents a new tool for studying the activation mechanism of different subsets of CTLs and could have potential for the development of novel autoimmunity therapies.

Screening for Natural Inhibitors of Topoisomerases I from Rhamnus davurica by Affinity Ultrafiltration and High-Performance Liquid Chromatography–Mass Spectrometry

Science.gov (United States)

Chen, Guilin; Guo, Mingquan

2017-01-01

Topoisomerase I (Topo I) catalyzes topological interconversion of duplex DNA during DNA replication and transcription, and has been deemed as important antineoplastic targets. In this study, the fraction R.d-60 from ethyl acetate extracts of Rhamnus davurica showed higher inhibitory rates against SGC-7901 and HT-29 compared with the R.d-30 fraction in vitro. However, the specific active components of R.d-60 fraction remain elusive. To this end, a method based on bio-affinity ultrafiltration and high performance liquid chromatography/electrospray mass spectrometry (HPLC- ESI-MS/MS) was developed to rapidly screen and identify the Topo I inhibitors in this fraction. The enrichment factors (EFs) were calculated to evaluate the binding affinities between the bioactive constituents and Topo I. As a result, eight ligands were identified and six of which with higher EFs showed more potential antitumor activity. Furthermore, antiproliferative assays in vitro (IC50 values) with two representative candidates (apigenin, quercetin) against SGC-7901, HT-29 and Hep G2 cells were conducted and further validated. Finally, the structure-activity relationships revealed that flavones contain a C2-C3 double bond of C ring exhibited higher bio-affinities to Topo I than those without it. This integrated method combining Topo I ultrafiltration with HPLC-MS/MS proved to be very efficient in rapid screening and identification of potential Topo I inhibitors from the complex extracts of medicinal plants, and could be further explored as a valuable high-throughput screening platform in the early drug discovery stage. PMID:28919906

The Cutting Edge of Affinity Electrophoresis Technology

Science.gov (United States)

Kinoshita, Eiji; Kinoshita-Kikuta, Emiko; Koike, Tohru

2015-01-01

Affinity electrophoresis is an important technique that is widely used to separate and analyze biomolecules in the fields of biology and medicine. Both quantitative and qualitative information can be gained through affinity electrophoresis. Affinity electrophoresis can be applied through a variety of strategies, such as mobility shift electrophoresis, charge shift electrophoresis or capillary affinity electrophoresis. These strategies are based on changes in the electrophoretic patterns of biological macromolecules that result from interactions or complex-formation processes that induce changes in the size or total charge of the molecules. Nucleic acid fragments can be characterized through their affinity to other molecules, for example transcriptional factor proteins. Hydrophobic membrane proteins can be identified by means of a shift in the mobility induced by a charged detergent. The various strategies have also been used in the estimation of association/disassociation constants. Some of these strategies have similarities to affinity chromatography, in that they use a probe or ligand immobilized on a supported matrix for electrophoresis. Such methods have recently contributed to profiling of major posttranslational modifications of proteins, such as glycosylation or phosphorylation. Here, we describe advances in analytical techniques involving affinity electrophoresis that have appeared during the last five years. PMID:28248262

Affine Fullerene C60 in a GS-Quasigroup

Directory of Open Access Journals (Sweden)

Vladimir Volenec

2014-01-01

Full Text Available It will be shown that the affine fullerene C60, which is defined as an affine image of buckminsterfullerene C60, can be obtained only by means of the golden section. The concept of the affine fullerene C60 will be constructed in a general GS-quasigroup using the statements about the relationships between affine regular pentagons and affine regular hexagons. The geometrical interpretation of all discovered relations in a general GS-quasigroup will be given in the GS-quasigroup C(1/2(1+5.

High- and low-affinity binding of S-citalopram to the human serotonin transporter mutated at 20 putatively important amino acid positions

DEFF Research Database (Denmark)

Plenge, Per; Wiborg, Ove

2005-01-01

of presumed importance. Binding of S-citalopram, both to the high-affinity-binding site and to the allosteric binding site, was measured in these mutants with the purpose of investigating the connection between the two binding sites. The amino acid substitutions did not introduce large changes in the two...

Affinity Spaces and 21st Century Learning

Science.gov (United States)

Gee, James Paul

2017-01-01

This article discusses video games as "attractors" to "affinity spaces." It argues that affinity spaces are key sites today where people teach and learn 21st Century skills. While affinity spaces are proliferating on the Internet as interest-and-passion-driven sites devoted to a common set of endeavors, they are not new, just…

New immunogenic form for vasopressin: production of high-affinity antiserum and RIA for plasmatic AVP

International Nuclear Information System (INIS)

Rougon-Rappuzi, G.; Delaage, M.A.; Conte-Devolx, B.; Millet, Y.

1977-01-01

A highly sensitive and specific radioimmunoassay (RIA) for arginine-vasopressin (AVP) was developped and applied to the measurement of AVP in human plasma. High-affinity antivasopressin antibodies with limited association constant heterogeneity have been induced by immunizing rabbits with Lysine-vasopressine (LVP) coupled to a human immunoglobulin (IgA). Replacing air drying of acetone-petroleum ether extracts by lyophilisation increased significantly the yields of AVP. Equilibrium dialysis was used for separating bound and free antigen, thus reducing the total time required for the assay to 48 hours. Only 1 ml of plasma was required for routine determinations due to a sensitivity threshold better than 0.5 pg/ml. Plasma AVP levels of normal subjects and of patients with inappropriate ADH secretion (SIADH) were determined during different hydratation states and following nicotin of ethanol infusions. (orig.) [de

On the structure of self-affine convex bodies

Energy Technology Data Exchange (ETDEWEB)

Voynov, A S [M. V. Lomonosov Moscow State University, Faculty of Mechanics and Mathematics, Moscow (Russian Federation)

2013-08-31

We study the structure of convex bodies in R{sup d} that can be represented as a union of their affine images with no common interior points. Such bodies are called self-affine. Vallet's conjecture on the structure of self-affine bodies was proved for d = 2 by Richter in 2011. In the present paper we disprove the conjecture for all d≥3 and derive a detailed description of self-affine bodies in R{sup 3}. Also we consider the relation between properties of self-affine bodies and functional equations with a contraction of an argument. Bibliography: 10 titles.

Affinity membranes for hormone removal from aqueous solutions

NARCIS (Netherlands)

Urmenyi, A.M.; Poot, Andreas A.; Wessling, Matthias; Mulder, M.H.V.

2005-01-01

A novel affinity membrane was prepared by covalent binding of antibodies (against 17--estradiol) to a micro-porous poly(ethylene vinyl alcohol) (EVAL) membrane, taking benefit from the high surface area of EVAL membranes and the large number of reactive groups available for further surface

ZipA binds to FtsZ with high affinity and enhances the stability of FtsZ protofilaments.

Directory of Open Access Journals (Sweden)

Anuradha Kuchibhatla

Full Text Available A bacterial membrane protein ZipA that tethers FtsZ to the membrane is known to promote FtsZ assembly. In this study, the binding of ZipA to FtsZ was monitored using fluorescence spectroscopy. ZipA was found to bind to FtsZ with high affinities at three different (6.0, 6.8 and 8.0 pHs, albeit the binding affinity decreased with increasing pH. Further, thick bundles of FtsZ protofilaments were observed in the presence of ZipA under the pH conditions used in this study indicating that ZipA can promote FtsZ assembly and stabilize FtsZ polymers under unfavorable conditions. Bis-ANS, a hydrophobic probe, decreased the interaction of FtsZ and ZipA indicating that the interaction between FtsZ and ZipA is hydrophobic in nature. ZipA prevented the dilution induced disassembly of FtsZ polymers suggesting that it stabilizes FtsZ protofilaments. Fluorescein isothiocyanate-labeled ZipA was found to be uniformly distributed along the length of the FtsZ protofilaments indicating that ZipA stabilizes FtsZ protofilaments by cross-linking them.

DOTA-derivatives of octreotide dicarba-analogues with high affinity for somatostatin sst2,5 receptors

Science.gov (United States)

Pratesi, Alessandro; Ginanneschi, Mauro; Lumini, Marco; Papini, Anna M.; Novellino, Ettore; Brancaccio, Diego; Carotenuto, Alfonso

2017-02-01

In vivo somatostatin receptor scintigraphy is a valuable method for the visualization of human endocrine tumours and their metastases. In fact, peptide ligands of somatostatin receptors (sst’s) conjugated with chelating agents are in clinical use. We have recently developed octreotide dicarba-analogues, which show interesting binding profiles at sst’s. In this context, it was mandatory to explore the possibility that our analogues could maintain their activity also upon conjugation with DOTA. In this paper, we report and discuss the synthesis, binding affinity and conformational preferences of three DOTA-conjugated dicarba-analogues of octreotide. Interestingly, two conjugated analogues exhibited nanomolar affinities on sst2 and sst5 somatostatin receptor subtypes.

Design, Synthesis, and Biological Evaluation of Small, High-Affinity Siglec-7 Ligands: Toward Novel Inhibitors of Cancer Immune Evasion.

Science.gov (United States)

Prescher, Horst; Frank, Martin; Gütgemann, Stephan; Kuhfeldt, Elena; Schweizer, Astrid; Nitschke, Lars; Watzl, Carsten; Brossmer, Reinhard

2017-02-09

Natural killer cells are able to directly lyse tumor cells, thereby participating in the immune surveillance against cancer. Unfortunately, many cancer cells use immune evasion strategies to avoid their eradication by the immune system. A prominent escape strategy of malignant cells is to camouflage themselves with Siglec-7 ligands, thereby recruiting the inhibitory receptor Siglec-7 expressed on the NK cell surface which subsequently inhibits NK-cell-mediated lysis. Here we describe the synthesis and evaluation of the first, high-affinity low molecular weight Siglec-7 ligands to interfere with cancer cell immune evasion. The compounds are Sialic acid derivatives and bind with low micromolar K d values to Siglec-7. They display up to a 5000-fold enhanced affinity over the unmodified sialic acid scaffold αMe Neu5Ac, the smallest known natural Siglec-7 ligand. Our results provide a novel immuno-oncology strategy employing natural immunity in the fight against cancers, in particular blocking Siglec-7 with low molecular weight compounds.

Polynomials associated with equilibria of affine Toda-Sutherland systems

International Nuclear Information System (INIS)

Odake, S; Sasaki, R

2004-01-01

An affine Toda-Sutherland system is a quasi-exactly solvable multi-particle dynamics based on an affine simple root system. It is a 'cross' between two well-known integrable multi-particle dynamics, an affine Toda molecule (exponential potential, periodic nearest-neighbour interaction) and a Sutherland system (inverse sine-square interaction). Polynomials describing the equilibrium positions of affine Toda-Sutherland systems are determined for all affine simple root systems

Duals of Affine Grassmann Codes and Their Relatives

DEFF Research Database (Denmark)

Beelen, P.; Ghorpade, S. R.; Hoholdt, T.

2012-01-01

Affine Grassmann codes are a variant of generalized Reed-Muller codes and are closely related to Grassmann codes. These codes were introduced in a recent work by Beelen Here, we consider, more generally, affine Grassmann codes of a given level. We explicitly determine the dual of an affine...... Grassmann code of any level and compute its minimum distance. Further, we ameliorate the results by Beelen concerning the automorphism group of affine Grassmann codes. Finally, we prove that affine Grassmann codes and their duals have the property that they are linear codes generated by their minimum......-weight codewords. This provides a clean analogue of a corresponding result for generalized Reed-Muller codes....

Cartilage Acidic Protein 2 a hyperthermostable, high affinity calcium-binding protein.

Science.gov (United States)

Anjos, Liliana; Gomes, Ana S; Melo, Eduardo P; Canário, Adelino V; Power, Deborah M

2013-03-01

Cartilage Acidic Protein 2 (CRTAC2) is a novel protein present from prokaryotes to vertebrates with abundant expression in the teleost fish pituitary gland and an isoform of CRTAC1, a chondrocyte marker in humans. The two proteins are non-integrins containing N-terminal integrin-like Ca(2+)-binding motifs and their structure and function remain to be assigned. Structural studies of recombinant sea bream (sb)CRTAC2 revealed it is composed of 8.8% α-helix, 33.4% β-sheet and 57.8% unordered protein. sbCRTAC2 bound Ca(2+) with high affinity (K(d)=1.46nM) and favourable Gibbs free energy (∆G=-12.4kcal/mol). The stoichiometry for Ca(2+) bound to sbCRTAC2 at saturation indicated six Ca(2+) ligand-binding sites exist per protein molecule. No conformational change in sbCRTAC2 occurred in the presence of Ca(2+). Fluorescence emission revealed that the tertiary structure of the protein is hyperthermostable between 25°C and 95°C and the fully unfolded state is only induced by chemical denaturing (4M GndCl). sbCRTAC has a widespread tissue distribution and is present as high molecular weight aggregates, although strong reducing conditions promote formation of the monomer. sbCRTAC2 promotes epithelial cell outgrowth in vitro suggesting it may share functional homology with mammalian CRTAC1, recently implicated in cell-cell and cell-matrix interactions. Copyright © 2012 Elsevier B.V. All rights reserved.

A Novel Vertex Affinity for Community Detection

Energy Technology Data Exchange (ETDEWEB)

Yoo, Andy [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Sanders, Geoffrey [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Henson, Van [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Vassilevski, Panayot [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

2015-10-05

We propose a novel vertex affinity measure in this paper. The new vertex affinity quantifies the proximity between two vertices in terms of their clustering strength and is ideal for such graph analytics applications as community detection. We also developed a framework that combines simple graph searches and resistance circuit formulas to compute the vertex affinity efficiently. We study the properties of the new affinity measure empirically in comparison to those of other popular vertex proximity metrics. Our results show that the existing metrics are ill-suited for community detection due to their lack of fundamental properties that are essential for correctly capturing inter- and intra-cluster vertex proximity.

Characterization of SynCAM surface trafficking using a SynCAM derived ligand with high homophilic binding affinity

International Nuclear Information System (INIS)

Breillat, Christelle; Thoumine, Olivier; Choquet, Daniel

2007-01-01

In order to better probe SynCAM function in neurons, we produced a fusion protein between the extracellular domain of SynCAM1 and the constant fragment of human IgG (SynCAM-Fc). Whether in soluble form or immobilized on latex microspheres, the chimera bound specifically to the surface of hippocampal neurons and recruited endogenous SynCAM molecules. SynCAM-Fc was also used in combination with Quantum Dots to follow the mobility of transfected SynCAM receptors at the neuronal surface. Both immobile and highly mobile SynCAM were found. Thus, SynCAM-Fc behaves as a high affinity ligand that can be used to study the function of SynCAM at the neuronal membrane

Complementary three-dimensional quantitative structure-activity relationship modeling of binding affinity and functional potency

DEFF Research Database (Denmark)

Tosco, Paolo; Ahring, Philip K; Dyhring, Tino

2009-01-01

Complementary 3D-QSAR modeling of binding affinity and functional potency is proposed as a tool to pinpoint the molecular features of the ligands, and the corresponding amino acids in the receptor, responsible for high affinity binding vs those driving agonist behavior and receptor activation. Th...

On affine non-negative matrix factorization

DEFF Research Database (Denmark)

Laurberg, Hans; Hansen, Lars Kai

2007-01-01

We generalize the non-negative matrix factorization (NMF) generative model to incorporate an explicit offset. Multiplicative estimation algorithms are provided for the resulting sparse affine NMF model. We show that the affine model has improved uniqueness properties and leads to more accurate id...

Identification of a High Affinity Nucleocapsid Protein Binding Element from The Bovine Leukemia Virus Genome

Science.gov (United States)

Yildiz, F. Zehra; Babalola, Kathleen; Summers, Michael F.

2012-01-01

Retroviral genome recognition is mediated by interactions between the nucleocapsid (NC) domain of the virally encoded Gag polyprotein and cognate RNA packaging elements that, for most retroviruses, appear to reside primarily within the 5′-untranslated region (5′-UTR) of the genome. Recent studies suggest that a major packaging determinant of Bovine Leukemia Virus (BLV), a member of the human T-cell leukemia virus (HTLV)/BLV family and a non-primate animal model for HTLV-induced leukemogenesis, resides within the gag open reading frame. We have prepared and purified the recombinant BLV NC protein and conducted electrophoretic mobility shift and isothermal titration calorimetry studies with RNA fragments corresponding to these proposed packaging elements. The gag-derived RNAs did not exhibit significant affinity for NC, suggesting an alternate role in packaging. However, an 83-nucleotide fragment of the 5′-UTR that resides just upstream of the gag start codon binds NC stoichiometrically and with high affinity (Kd = 136 ± 21 nM). These nucleotides were predicted to form tandem hairpin structures, and studies with smaller fragments indicate that the NC binding site resides exclusively within the distal hairpin (residues G369- U399, Kd = 67 ± 8 nM at physiological ionic strength). Unlike all other structurally characterized retroviral NC binding RNAs, this fragment is not expected to contain exposed guanosines, suggesting that RNA binding may be mediated by a previously uncharacterized mechanism. PMID:22846919

Peptide Binding to HLA Class I Molecules: Homogenous, High-Throughput Screening, and Affinity Assays

DEFF Research Database (Denmark)

Harndahl, Mikkel; Justesen, Sune Frederik Lamdahl; Lamberth, Kasper

2009-01-01

, better signal-to-background ratios, and a higher capacity. They also describe an efficient approach to screen peptides for binding to HLA molecules. For the occasional user, this will serve as a robust, simple peptide-HLA binding assay. For the more dedicated user, it can easily be performed in a high-throughput...... the luminescent oxygen channeling immunoassay technology (abbreviated LOCI and commercialized as AlphaScreen (TM)). Compared with an enzyme-linked immunosorbent assay-based peptide-HLA class I binding assay, the LOCI assay yields virtually identical affinity measurements, although having a broader dynamic range...... screening mode using standard liquid handling robotics and 384-well plates. We have successfully applied this assay to more than 60 different HLA molecules, leading to more than 2 million measurements. (Journal of Biomolecular Screening 2009: 173-180)...

Improved methodology for the affinity isolation of human protein complexes expressed at near endogenous levels

DEFF Research Database (Denmark)

Domanski, Michal; Molloy, Kelly; Jiang, Hua

2012-01-01

An efficient and reliable procedure for the capture of affinity-tagged proteins and associated complexes from human cell lines is reported. Through multiple optimizations, high yield and low background affinity-purifications are achieved from modest quantities of human cells expressing endogenous...

Generation of a novel high-affinity monoclonal antibody with conformational recognition epitope on human IgM.

Science.gov (United States)

Sarikhani, Sina; Mirshahi, Manouchehr; Gharaati, Mohammad Reza; Mirshahi, Tooran

2010-11-01

As IgM is the first isotype of antibody which appears in blood after initial exposure to a foreign antigen in the pattern of primary response, detection, and quantification of this molecule in blood seems invaluable. To approach these goals, generation, and characterization of a highly specific mAb (monoclonal antibody) against human IgM were investigated. Human IgM immunoglobulins were used to immunize Balb/c mice. Spleen cells taken from the immunized animals were fused with SP2/O myeloma cells using PEG (polyethylene glycol, MW 1450) as fusogen. The hybridomas were cultured in HAT containing medium and supernatants from the growing hybrids were screened by enzyme-linked immunosorbent assay (ELISA) using plates coated with pure human IgM and the positive wells were then cloned at limiting dilutions. The best clone designated as MAN-1, was injected intraperitoneally to some Pristane-injected mice. Anti-IgM mAb was purified from the animals' ascitic fluid by protein-G sepharose followed by DEAE-cellulose ion exchange chromatography. MAN-1 interacted with human IgM with a very high specificity and affinity. The purity of the sample was tested by SDS-PAGE and the affinity constant was measured (K(a) = 3.5 x 10(9)M(-1). Immunoblotting and competitive ELISA were done and the results showed that the harvested antibody recognizes a conformational epitope on the mu chain of human IgM and there was no cross-reactivity with other subclasses of immunoglobulins. Furthermore, isotyping test was done and the results showed the subclass of the obtained mAb which was IgG(1)kappa.

Specific capture and detection of Staphylococcus aureus with high-affinity modified aptamers to cell surface components.

Science.gov (United States)

Baumstummler, A; Lehmann, D; Janjic, N; Ochsner, U A

2014-10-01

Slow off-rate modified aptamer (SOMAmer) reagents were generated to several Staphylococcus aureus cell surface-associated proteins via SELEX with multiple modified DNA libraries using purified recombinant or native proteins. High-affinity binding agents with sub-nanomolar Kd 's were obtained for staphylococcal protein A (SpA), clumping factors (ClfA, ClfB), fibronectin-binding proteins (FnbA, FnbB) and iron-regulated surface determinants (Isd). Further screening revealed several SOMAmers that specifically bound to Staph. aureus cells from all strains that were tested, but not to other staphylococci or other bacteria. SpA and ClfA SOMAmers proved useful for the selective capture and enrichment of Staph. aureus cells, as shown by culture and PCR, leading to improved limits of detection and efficient removal of PCR inhibitors. Detection of Staph. aureus cells was enhanced by several orders of magnitude when the bacterial cell surface was coated with SOMAmers followed by qPCR of the SOMAmers. Furthermore, fluorescence-labelled SpA SOMAmers demonstrated their utility as direct detection agents in flow cytometry. Significance and impact of the study: Monitoring for microbial contamination of food, water, nonsterile products or the environment is typically based on culture, PCR or antibodies. Aptamers that bind with high specificity and affinity to well-conserved cell surface epitopes represent a promising novel type of reagents to detect bacterial cells without the need for culture or cell lysis, including for the capture and enrichment of bacteria present at low cell densities and for the direct detection via qPCR or fluorescent staining. © 2014 Soma Logic, Inc. published by John Wiley & Sons Ltd On behalf of the society for Applied Microbiology.

Septide and neurokinin A are high-affinity ligands on the NK-1 receptor: evidence from homologous versus heterologous binding analysis.

Science.gov (United States)

Hastrup, H; Schwartz, T W

1996-12-16

The three main tachykinins, substance P, neurokinin A (NKA), and neurokinin B, are believed to be selective ligands for respectively the NK-1, NK-2 and NK-3 receptors. However, NKA also has actions which cannot be mediated through its normal NK-2 receptor and the synthetic peptide [pGlu6,Pro9]-Substance P9-11--called septide--is known to have tachykinin-like actions despite its apparent lack of binding to any known tachykinin receptor. In the cloned NK-1 receptor expressed in COS-7 cells NKA and septide as expected were poor competitors for radiolabeled substance P. However, by using radiolabeled NKA and septide directly, it was found that both peptides in homologous binding assays as well as in competition against each other in fact bound to the NK-1 receptor with high affinity: Kd values of 0.51 +/- 0.15 nM (NKA) and 0.55 +/- 0.03 nM (septide). It is concluded that NKA and septide are high-affinity ligands for the NK-1 receptor but that they are poor competitors for substance P, which in contrast competes very well for binding with both NKA and septide.

Contractions of affine spherical varieties

International Nuclear Information System (INIS)

Arzhantsev, I V

1999-01-01

The language of filtrations and contractions is used to describe the class of G-varieties obtainable as the total spaces of the construction of contraction applied to affine spherical varieties, which is well-known in invariant theory. These varieties are local models for arbitrary affine G-varieties of complexity 1 with a one-dimensional categorical quotient. As examples, reductive algebraic semigroups and three-dimensional SL 2 -varieties are considered

Characterization of an AtCCX5 gene from Arabidopsis thaliana that involves in high-affinity K+ uptake and Na+ transport in yeast

International Nuclear Information System (INIS)

Zhang, Xinxin; Zhang, Min; Takano, Tetsuo; Liu, Shenkui

2011-01-01

Highlights: → The AtCCX5 protein coding a putative cation calcium exchanger was characterized. → AtCCX5 expressed in yeast was localized in the plasma membrane and nuclear periphery. → AtCCX5 protein did not show the same transport properties as the CAXs. → AtCCX5 protein involves in mediating high-affinity K + uptake in yeast. → AtCCX5 protein also involves in Na + transport in yeast. -- Abstract: The gene for a putative cation calcium exchanger (CCX) from Arabidopsis thaliana, AtCCX5, was cloned and its function was analyzed in yeast. Green fluorescent protein-tagged AtCCX5 expressed in yeast was localized in the plasma membrane and nuclear periphery. The yeast transformants expressing AtCCX5 were created and their growth in the presence of various cations (K + , Na + , Ca 2+ , Mg 2+ , Fe 2+ , Cu 2+ , Co 2+ , Cd 2+ , Mn 2+ , Ba 2+ , Ni 2+ , Zn 2+ , and Li + ) were analyzed. AtCCX5 expression was found to affect the response to K + and Na + in yeast. The AtCCX5 transformant also showed a little better growth to Zn 2+ . The yeast mutant 9.3 expressing AtCCX5 restored growth of the mutant on medium with low K + (0.5 mM), and also suppressed its Na + sensitivity. Ion uptake experiments showed that AtCCX5 mediated relatively high-affinity K + uptake and was also involved in Na + transport in yeast. Taken together, these findings suggest that the AtCCX5 is a novel transport protein involves in mediating high-affinity K + uptake and Na + transport in yeast.

Amine-functionalized PVA-co-PE nanofibrous membrane as affinity membrane with high adsorption capacity for bilirubin.

Science.gov (United States)

Wang, Wenwen; Zhang, Hao; Zhang, Zhifeng; Luo, Mengying; Wang, Yuedan; Liu, Qiongzhen; Chen, Yuanli; Li, Mufang; Wang, Dong

2017-02-01

In this study, poly(vinyl alcohol-co-ethylene) (PVA-co-PE) nanofibrous membrane was activated by sodium hydroxide and cyanuric chloride, and then the activated membranes were functionalized by 1,3-propanediamine, hexamethylenediamine and diethylenetriamine to be affinity membranes for bilirubin removal, respectively. The chemical structures and morphologies of membranes were investigated by SEM, FTIR and XPS. And the adsorption ability of different amine-functionalized nanofibrous membranes for bilirubin was characterized. Furthermore, the effects of temperature, initial concentration of bilirubin, NaCl concentration and BSA concentration on the adsorption capacity for bilirubin of diethylenetriamine-functionalized nanofibrous membrane were studied. Results indicated that the adsorption capacity for bilirubin of diethylenetriamine-functionalized nanofibrous membrane could reach 85mg/g membrane when the initial bilirubin concentration was 200mg/L while the adsorption capacity could be increased to 110mg/g membrane if the initial bilirubin concentration was more than 400mg/L. The dynamic adsorption of diethylenetriamine-functionalized nanofibrous membrane showed that the ligands of amine groups on the membrane surface could be used as far as possible by recirculating the plasma with certain flow rates. Therefore, the diethylenetriamine-functionalized PVA-co-PE nanofibrous membrane possessed high adsorption capacity for bilirubin and it can be candidate as affinity membrane for bilirubin removal. Copyright © 2016. Published by Elsevier B.V.

A high-affinity, dimeric inhibitor of PSD-95 bivalently interacts with PDZ1-2 and protects against ischemic brain damage

DEFF Research Database (Denmark)

Bach, Anders*; Clausen, Bettina H; Møller, Magda

2012-01-01

Inhibition of the ternary protein complex of the synaptic scaffolding protein postsynaptic density protein-95 (PSD-95), neuronal nitric oxide synthase (nNOS), and the N-methyl-d-aspartate (NMDA) receptor is a potential strategy for treating ischemic brain damage, but high-affinity inhibitors are ...... of Tat-N-dimer (3 nmol/g) to mice subjected to focal cerebral ischemia reduces infarct volume with 40% and restores motor functions. Thus, Tat-N-dimer is a highly efficacious neuroprotective agent with therapeutic potential in stroke....

hPEPT1 Affinity and Translocation of Selected Gln-Sar and Glu-Sar Dipeptide Derivatives

DEFF Research Database (Denmark)

Eriksson, A. H.; Elm, Peter L.; Begtrup, Mikael

2005-01-01

using 14C-labeled Gly-Sar. Translocation was measured as fluorescence ratios induced by the substrates using the fluorescent probe BCECF and an epifluorescence microscope setup. All compounds showed high affinity to hPEPT1, but only the amides l-Gln(N,N-dimethyl)-Sar and l-Gln(N-piperidinyl)-Sar were...... been suggested. However, these are not necessarily predictive of compounds that are actually translocated by hPEPT1. More information on affinity to and translocation via hPEPT1 of side-chain-modified dipeptides may be gained by conducting a study of selected dipeptide derivatives with variety in size...... translocated by hPEPT1. hPEPT1 is very susceptible to modifications of the N-terminal amino acid side chain of dipeptidomimetic substrates, in terms of achieving compounds with high affinity for the transporter. However, as affinity is not predictive of translocation, derivatization in this position must...

Electrochemical affinity biosensors for detection of mycotoxins: A review.

Science.gov (United States)

Vidal, Juan C; Bonel, Laura; Ezquerra, Alba; Hernández, Susana; Bertolín, Juan R; Cubel, Carlota; Castillo, Juan R

2013-11-15

This review discusses the current state of electrochemical biosensors in the determination of mycotoxins in foods. Mycotoxins are highly toxic secondary metabolites produced by molds. The acute toxicity of these results in serious human and animal health problems, although it has been only since early 1960s when the first studied aflatoxins were found to be carcinogenic. Mycotoxins affect a broad range of agricultural products, most important cereals and cereal-based foods. A majority of countries, mentioning especially the European Union, have established preventive programs to control contamination and strict laws of the permitted levels in foods. Official methods of analysis of mycotoxins normally requires sophisticated instrumentation, e.g. liquid chromatography with fluorescence or mass detectors, combined with extraction procedures for sample preparation. For about sixteen years, the use of simpler and faster analytical procedures based on affinity biosensors has emerged in scientific literature as a very promising alternative, particularly electrochemical (i.e., amperometric, impedance, potentiometric or conductimetric) affinity biosensors due to their simplicity and sensitivity. Typically, electrochemical biosensors for mycotoxins use specific antibodies or aptamers as affinity ligands, although recombinant antibodies, artificial receptors and molecular imprinted polymers show potential utility. This article deals with recent advances in electrochemical affinity biosensors for mycotoxins and covers complete literature from the first reports about sixteen years ago. Copyright © 2013 Elsevier B.V. All rights reserved.

Paper-based immune-affinity arrays for detection of multiple mycotoxins in cereals.

Science.gov (United States)

Li, Li; Chen, Hongpu; Lv, Xiaolan; Wang, Min; Jiang, Xizhi; Jiang, Yifei; Wang, Heye; Zhao, Yongfu; Xia, Liru

2018-03-01

Mycotoxins produced by different species of fungi may coexist in cereals and feedstuffs, and could be highly toxic for humans and animals. For quantification of multiple mycotoxins in cereals, we developed a paper-based mycotoxin immune-affinity array. First, paper-based microzone arrays were fabricated by photolithography. Then, monoclonal mycotoxin antibodies were added in a copolymerization reaction with a cross-linker to form an immune-affinity monolith on the paper-based microzone array. With use of a competitive immune-response format, paper-based mycotoxin immune-affinity arrays were successfully applied to detect mycotoxins in samples. The detection limits for deoxynivalenol, zearalenone, T-2 toxin, and HT-2 toxin were 62.7, 10.8, 0.36, and 0.23 μg·kg -1 , respectively, which meet relevant requirements for these compounds in food. The recovery rates were 81-86% for deoxynivalenol, 89-117% for zearalenone, 79-86% for T-2 toxin, and 78-83% for HT-2 toxin, and showed the paper-based immune-affinity arrays had good reproducibility. In summary, the paper-based mycotoxin immune-affinity array provides a sensitive, rapid, accurate, stable, and convenient platform for detection of multiple mycotoxins in agro-foods. Graphical abstract Paper-based immune-affinity monolithic array. DON deoxynivalenol, HT-2 HT-2 toxin, T-2 T-2 toxin, PEGDA polyethylene glycol diacrylate, ZEN zearalenone.

Providing affinity

DEFF Research Database (Denmark)

Guglielmi, Michel; Johannesen, Hl

2004-01-01

, Essex, Hertfordshire, Norfolk and Suffolk. Research found that there was a lack of identity or sense of belonging and nothing anchoring people to the region as a whole. Common affinity is somehow forced to the people of East England and thereby we came to the conclusion that a single landmark...... and potential situations but also virtual events that calls for an undeterminated process of resolution. This process is activated by the user who co-produces the actualisation as an answer to a virtual reality that we defined at the first place. The potential situations or the possible it is a fantomatic real....... The possible is like the real. It is determinated and it only lakes existence. While the possible is already made, the virtual is like a problematic which needs to be resolved and actualized. Our installations are based on high tech interactivity where we use sensors and remote communication to offer a sense...

High density and ligand affinity confer ultrasensitive signal detection by a guanylyl cyclase chemoreceptor

Science.gov (United States)

Pichlo, Magdalena; Bungert-Plümke, Stefanie; Weyand, Ingo; Seifert, Reinhard; Bönigk, Wolfgang; Strünker, Timo; Kashikar, Nachiket Dilip; Goodwin, Normann; Müller, Astrid; Körschen, Heinz G.; Collienne, Ursel; Pelzer, Patric; Van, Qui; Enderlein, Jörg; Klemm, Clementine; Krause, Eberhard; Trötschel, Christian; Poetsch, Ansgar; Kremmer, Elisabeth

2014-01-01

Guanylyl cyclases (GCs), which synthesize the messenger cyclic guanosine 3′,5′-monophosphate, control several sensory functions, such as phototransduction, chemosensation, and thermosensation, in many species from worms to mammals. The GC chemoreceptor in sea urchin sperm can decode chemoattractant concentrations with single-molecule sensitivity. The molecular and cellular underpinnings of such ultrasensitivity are not known for any eukaryotic chemoreceptor. In this paper, we show that an exquisitely high density of 3 × 105 GC chemoreceptors and subnanomolar ligand affinity provide a high ligand-capture efficacy and render sperm perfect absorbers. The GC activity is terminated within 150 ms by dephosphorylation steps of the receptor, which provides a means for precise control of the GC lifetime and which reduces “molecule noise.” Compared with other ultrasensitive sensory systems, the 10-fold signal amplification by the GC receptor is surprisingly low. The hallmarks of this signaling mechanism provide a blueprint for chemical sensing in small compartments, such as olfactory cilia, insect antennae, or even synaptic boutons. PMID:25135936

Modeling DNA affinity landscape through two-round support vector regression with weighted degree kernels

KAUST Repository

Wang, Xiaolei

2014-12-12

Background: A quantitative understanding of interactions between transcription factors (TFs) and their DNA binding sites is key to the rational design of gene regulatory networks. Recent advances in high-throughput technologies have enabled high-resolution measurements of protein-DNA binding affinity. Importantly, such experiments revealed the complex nature of TF-DNA interactions, whereby the effects of nucleotide changes on the binding affinity were observed to be context dependent. A systematic method to give high-quality estimates of such complex affinity landscapes is, thus, essential to the control of gene expression and the advance of synthetic biology. Results: Here, we propose a two-round prediction method that is based on support vector regression (SVR) with weighted degree (WD) kernels. In the first round, a WD kernel with shifts and mismatches is used with SVR to detect the importance of subsequences with different lengths at different positions. The subsequences identified as important in the first round are then fed into a second WD kernel to fit the experimentally measured affinities. To our knowledge, this is the first attempt to increase the accuracy of the affinity prediction by applying two rounds of string kernels and by identifying a small number of crucial k-mers. The proposed method was tested by predicting the binding affinity landscape of Gcn4p in Saccharomyces cerevisiae using datasets from HiTS-FLIP. Our method explicitly identified important subsequences and showed significant performance improvements when compared with other state-of-the-art methods. Based on the identified important subsequences, we discovered two surprisingly stable 10-mers and one sensitive 10-mer which were not reported before. Further test on four other TFs in S. cerevisiae demonstrated the generality of our method. Conclusion: We proposed in this paper a two-round method to quantitatively model the DNA binding affinity landscape. Since the ability to modify

Video-Based Electroshocking Platform to Identify Lamprey Ammocoete Habitats: Field Validation and New Discoveries in the Columbia River Basin

Energy Technology Data Exchange (ETDEWEB)

Arntzen, Evan V.; Mueller, Robert P.

2017-05-04

A deep water electroshocking platform (DEP), developed to characterize larval lampreys (ammocoetes) and associated habitat in depths up to 15 m, was recently tested in the field. The DEP samples 0.55 m2∙min-1 without requiring ammocoete transport to the surface. Searches were conducted at a known rearing location (mouth of the Wind River, WA) and at locations on the Cowlitz River, WA, where ammocoetes had not previously been found. At the mouth of the Wind River, video imaged ammocoetes ranged from 50 to 150 mm in water depths between 1.5 m and 4.5 m and were more common in sediments containing organic silt. Ammocoetes (n=137) were detected at 61% of locations sampled (summer) and 50% of the locations sampled (winter). Following the field verification, the DEP was used on the lower 11.7 km of the Cowlitz River, WA. Ammocoetes (n=41) were found with a detection rate of 26% at specific search locations. Cowlitz River sediment containing ammocoetes was also dominated by silt with organic material, often downstream of alluvial bars in water depths from 0.8 to 1.7 m. Test results indicated a high sampling efficiency, favorable detection rates, and little or no impact to ammocoetes and their surrounding benthic environments.

Cyr61/CCN1 displays high-affinity binding to the somatomedin B(1-44 domain of vitronectin.

Directory of Open Access Journals (Sweden)

Ivo M B Francischetti

2010-02-01

Full Text Available Cyr61 is a member of the CCN (Cyr61, connective tissue growth, NOV family of extracellular-associated (matricellular proteins that present four distinct functional modules, namely insulin-like growth factor binding protein (IGFBP, von Willebrand factor type C (vWF, thrombospondin type 1 (TSP, and C-terminal growth factor cysteine knot (CT domain. While heparin sulphate proteoglycans reportedly mediate the interaction of Cyr61 with the matrix and cell surface, the role of other extracellular associated proteins has not been revealed.In this report, surface plasmon resonance (SPR experiments and solid-phase binding assays demonstrate that recombinant Cyr61 interacts with immobilized monomeric or multimeric vitronectin (VTNC with K(D in the nanomolar range. Notably, the binding site for Cyr61 was identified as the somatomedin B domain (SMTB(1-44 of VTNC, which mediates its interaction with PAI-1, uPAR, and integrin alphav beta3. Accordingly, PAI-1 outcompetes Cyr61 for binding to immobilized SMTB(1-44, and Cyr61 attenuates uPAR-mediated U937 adhesion to VTNC. In contrast, isothermal titration calorimetry shows that Cyr61 does not display high-affinity binding for SMTB(1-44 in solution. Nevertheless, competitive ELISA revealed that multimeric VTNC, heat-modified monomeric VTNC, or SMTB(1-44 at high concentrations attenuate Cyr61 binding to immobilized VTNC, while monomeric VTNC was ineffective. Therefore, immobilization of VTNC exposes cryptic epitopes that recognize Cyr61 with high affinity, as reported for a number of antibodies, beta-endorphin, and other molecules.The finding that Cyr61 interacts with the SMTB(1-44 domain suggests that VTNC represent a point of anchorage for CCN family members to the matrix. Results are discussed in the context of the role of CCN and VTNC in matrix biology and angiogenesis.

Rapid Diagnostic Assay for Intact Influenza Virus Using a High Affinity Hemagglutinin Binding Protein.

Science.gov (United States)

Anderson, Caitlin E; Holstein, Carly A; Strauch, Eva-Maria; Bennett, Steven; Chevalier, Aaron; Nelson, Jorgen; Fu, Elain; Baker, David; Yager, Paul

2017-06-20

Influenza is a ubiquitous and recurring infection that results in approximately 500 000 deaths globally each year. Commercially available rapid diagnostic tests are based upon detection of the influenza nucleoprotein, which are limited in that they are unable to differentiate by species and require an additional viral lysis step. Sample preprocessing can be minimized or eliminated by targeting the intact influenza virus, thereby reducing assay complexity and leveraging the large number of hemagglutinin proteins on the surface of each virus. Here, we report the development of a paper-based influenza assay that targets the hemagglutinin protein; the assay employs a combination of antibodies and novel computationally designed, recombinant affinity proteins as the capture and detection agents. This system leverages the customizability of recombinant protein design to target the conserved receptor-binding pocket of the hemagglutinin protein and to match the trimeric nature of hemagglutinin for improved avidity. Using this assay, we demonstrate the first instance of intact influenza virus detection using a combination of antibody and affinity proteins within a porous network. The recombinant head region binder based assays yield superior analytical sensitivity as compared to the antibody based assay, with lower limits of detection of 3.54 × 10 7 and 1.34 × 10 7 CEID 50 /mL for the mixed and all binder stacks, respectively. Not only does this work describe the development of a novel influenza assay, it also demonstrates the power of recombinant affinity proteins for use in rapid diagnostic assays.

Extreme disorder in an ultrahigh-affinity protein complex

DEFF Research Database (Denmark)

Borgia, Alessandro; Borgia, Madeleine B; Bugge, Katrine

2018-01-01

Molecular communication in biology is mediated by protein interactions. According to the current paradigm, the specificity and affinity required for these interactions are encoded in the precise complementarity of binding interfaces. Even proteins that are disordered under physiological conditions...... with picomolar affinity, but fully retain their structural disorder, long-range flexibility and highly dynamic character. On the basis of closely integrated experiments and molecular simulations, we show that the interaction can be explained by the large opposite net charge of the two proteins, without requiring...... or that contain large unstructured regions commonly interact with well-structured binding sites on other biomolecules. Here we demonstrate the existence of an unexpected interaction mechanism: the two intrinsically disordered human proteins histone H1 and its nuclear chaperone prothymosin-α associate in a complex...

Carbonate-sensitive phytotransferrin controls high-affinity iron uptake in diatoms

Science.gov (United States)

McQuaid, Jeffrey B.; Kustka, Adam B.; Oborník, Miroslav; Horák, Aleš; McCrow, John P.; Karas, Bogumil J.; Zheng, Hong; Kindeberg, Theodor; Andersson, Andreas J.; Barbeau, Katherine A.; Allen, Andrew E.

2018-03-01

In vast areas of the ocean, the scarcity of iron controls the growth and productivity of phytoplankton. Although most dissolved iron in the marine environment is complexed with organic molecules, picomolar amounts of labile inorganic iron species (labile iron) are maintained within the euphotic zone and serve as an important source of iron for eukaryotic phytoplankton and particularly for diatoms. Genome-enabled studies of labile iron utilization by diatoms have previously revealed novel iron-responsive transcripts, including the ferric iron-concentrating protein ISIP2A, but the mechanism behind the acquisition of picomolar labile iron remains unknown. Here we show that ISIP2A is a phytotransferrin that independently and convergently evolved carbonate ion-coordinated ferric iron binding. Deletion of ISIP2A disrupts high-affinity iron uptake in the diatom Phaeodactylum tricornutum, and uptake is restored by complementation with human transferrin. ISIP2A is internalized by endocytosis, and manipulation of the seawater carbonic acid system reveals a second-order dependence on the concentrations of labile iron and carbonate ions. In P. tricornutum, the synergistic interaction of labile iron and carbonate ions occurs at environmentally relevant concentrations, revealing that carbonate availability co-limits iron uptake. Phytotransferrin sequences have a broad taxonomic distribution and are abundant in marine environmental genomic datasets, suggesting that acidification-driven declines in the concentration of seawater carbonate ions will have a negative effect on this globally important eukaryotic iron acquisition mechanism.

Global affine differential geometry of hypersurfaces

CERN Document Server

Li, An-Min; Zhao, Guosong; Hu, Zejun

2015-01-01

This book draws a colorful and widespread picture of global affine hypersurface theory up to the most recent state. Moreover, the recent development revealed that affine differential geometry- as differential geometry in general- has an exciting intersection area with other fields of interest, like partial differential equations, global analysis, convex geometry and Riemann surfaces.

Heparin-binding peptide as a novel affinity tag for purification of recombinant proteins.

Science.gov (United States)

Morris, Jacqueline; Jayanthi, Srinivas; Langston, Rebekah; Daily, Anna; Kight, Alicia; McNabb, David S; Henry, Ralph; Kumar, Thallapuranam Krishnaswamy Suresh

2016-10-01

Purification of recombinant proteins constitutes a significant part of the downstream processing in biopharmaceutical industries. Major costs involved in the production of bio-therapeutics mainly depend on the number of purification steps used during the downstream process. Affinity chromatography is a widely used method for the purification of recombinant proteins expressed in different expression host platforms. Recombinant protein purification is achieved by fusing appropriate affinity tags to either N- or C- terminus of the target recombinant proteins. Currently available protein/peptide affinity tags have proved quite useful in the purification of recombinant proteins. However, these affinity tags suffer from specific limitations in their use under different conditions of purification. In this study, we have designed a novel 34-amino acid heparin-binding affinity tag (HB-tag) for the purification of recombinant proteins expressed in Escherichia coli (E. coli) cells. HB-tag fused recombinant proteins were overexpressed in E. coli in high yields. A one-step heparin-Sepharose-based affinity chromatography protocol was developed to purify HB-fused recombinant proteins to homogeneity using a simple sodium chloride step gradient elution. The HB-tag has also been shown to facilitate the purification of target recombinant proteins from their 8 M urea denatured state(s). The HB-tag has been demonstrated to be successfully released from the fusion protein by an appropriate protease treatment to obtain the recombinant target protein(s) in high yields. Results of the two-dimensional NMR spectroscopy experiments indicate that the purified recombinant target protein(s) exist in the native conformation. Polyclonal antibodies raised against the HB-peptide sequence, exhibited high binding specificity and sensitivity to the HB-fused recombinant proteins (∼10 ng) in different crude cell extracts obtained from diverse expression hosts. In our opinion, the HB-tag provides a

Pharmacokinetics and biodistribution of a radioiodine labeled peptidomimetic ligand for high-affinity nerve growth factor receptors

Energy Technology Data Exchange (ETDEWEB)

Jung, K. H.; Kim, D. H.; Paik, J. Y.; Koh, B. H.; Bae, J. S.; Choe, Y. S.; Lee, K. H.; Kim, B. T. [Samsung Medical Center, Seoul (Korea, Republic of)

2005-07-01

Some of the obstacles for the clinical application of whole nerve growth factor (NGF) may be overcome by utilizing small molecule mimetics. We thus investigated the in vivo pharmacokinetics and biodistribution of a small cyclic peptide derived from NGF-[C(92-96)] with high receptor binding affinity. I-125 C(92-96) was labeled with the Bolton-Hunter method, and binding to TrkA/IgG chimeric protein was confirmed on a polyacrylamide gel after cross-linking. Pharmacokinetic analysis was performed in normal ICR mice intravenously injected with 0.5 MBq I-125 C(92-96) containing varying doses of C(92-96). Biodistribution studies were done at 6 h after injection. Cross-linkage analysis confirmed binding of I-125 C(92-96) to the high affinity NGF receptor, TrkA. Intravenously injected I-125 C(92-96) was cleared from the blood in a biexponential manner with an early T1/2{alpha} of 5.2 min and late T1/2{beta} of 121.3 min. Log blood-concentration decreased over time with a k-slope of 0.0025, clearance of 11.8{+-}0.5 ml/min, T1/2 of 4.1{+-}0.4 hr, and volume of distribution of 69.7{+-}4.6 ml. The pattern of elimination from the blood remained essentially unchanged regardless of the dose of added C(92-96), with dose-proportionate increases in AUCs and peak concentrations consistent with linear pharmacokinetics. Biodistribution studies demonstrated high kidney activity suggesting renal excretion of I-125 C(92-96). There were moderate levels of accumulation in the spleen, lungs and liver, followed by the myocardium and skeletal muscle, whereas brain uptake was low (< 0.2 %ID/gm). Intravenously administered C(92-96) follows linear pharmacokinetics, and is cleared from the circulation at a rate comparable to whole NGF despite its substantially smaller size. Although intravenous C(92-96) does not adequately reach brain tissue, clinically relevant doses can achieve major organ accumulation levels that may be sufficient to elicit biologic responses through NGF receptors.

Pharmacokinetics and biodistribution of a radioiodine labeled peptidomimetic ligand for high-affinity nerve growth factor receptors

International Nuclear Information System (INIS)

Jung, K. H.; Kim, D. H.; Paik, J. Y.; Koh, B. H.; Bae, J. S.; Choe, Y. S.; Lee, K. H.; Kim, B. T.

2005-01-01

Some of the obstacles for the clinical application of whole nerve growth factor (NGF) may be overcome by utilizing small molecule mimetics. We thus investigated the in vivo pharmacokinetics and biodistribution of a small cyclic peptide derived from NGF-[C(92-96)] with high receptor binding affinity. I-125 C(92-96) was labeled with the Bolton-Hunter method, and binding to TrkA/IgG chimeric protein was confirmed on a polyacrylamide gel after cross-linking. Pharmacokinetic analysis was performed in normal ICR mice intravenously injected with 0.5 MBq I-125 C(92-96) containing varying doses of C(92-96). Biodistribution studies were done at 6 h after injection. Cross-linkage analysis confirmed binding of I-125 C(92-96) to the high affinity NGF receptor, TrkA. Intravenously injected I-125 C(92-96) was cleared from the blood in a biexponential manner with an early T1/2α of 5.2 min and late T1/2β of 121.3 min. Log blood-concentration decreased over time with a k-slope of 0.0025, clearance of 11.8±0.5 ml/min, T1/2 of 4.1±0.4 hr, and volume of distribution of 69.7±4.6 ml. The pattern of elimination from the blood remained essentially unchanged regardless of the dose of added C(92-96), with dose-proportionate increases in AUCs and peak concentrations consistent with linear pharmacokinetics. Biodistribution studies demonstrated high kidney activity suggesting renal excretion of I-125 C(92-96). There were moderate levels of accumulation in the spleen, lungs and liver, followed by the myocardium and skeletal muscle, whereas brain uptake was low (< 0.2 %ID/gm). Intravenously administered C(92-96) follows linear pharmacokinetics, and is cleared from the circulation at a rate comparable to whole NGF despite its substantially smaller size. Although intravenous C(92-96) does not adequately reach brain tissue, clinically relevant doses can achieve major organ accumulation levels that may be sufficient to elicit biologic responses through NGF receptors

Enhanced binding affinity, remarkable selectivity, and high capacity of CO 2 by dual functionalization of a rht-type metal-organic framework

KAUST Repository

Li, Baiyan

2011-12-23

Open and friendly: The smallest member of the rht-type metal-organic frameworks (MOFs, see picture) constructed by a hexacarboxylate ligand with a nitrogen-rich imino triazine backbone shows a significantly enhanced gas binding affinity relative to all other isoreticular rht-type MOFs. The high adsorption capacity and remarkable selectivity of CO 2 are attributed to the high density of open metal and Lewis basic sites in the framework. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

High affinity radiopharmaceuticals based upon lansoprazole for PET imaging of aggregated tau in Alzheimer's disease and progressive supranuclear palsy: synthesis, preclinical evaluation, and lead selection.

Science.gov (United States)

Fawaz, Maria V; Brooks, Allen F; Rodnick, Melissa E; Carpenter, Garrett M; Shao, Xia; Desmond, Timothy J; Sherman, Phillip; Quesada, Carole A; Hockley, Brian G; Kilbourn, Michael R; Albin, Roger L; Frey, Kirk A; Scott, Peter J H

2014-08-20

Abnormally aggregated tau is the hallmark pathology of tauopathy neurodegenerative disorders and is a target for development of both diagnostic tools and therapeutic strategies across the tauopathy disease spectrum. Development of carbon-11- or fluorine-18-labeled radiotracers with appropriate affinity and specificity for tau would allow noninvasive quantification of tau burden using positron emission tomography (PET) imaging. We have synthesized [(18)F]lansoprazole, [(11)C]N-methyl lansoprazole, and [(18)F]N-methyl lansoprazole and identified them as high affinity radiotracers for tau with low to subnanomolar binding affinities. Herein, we report radiosyntheses and extensive preclinical evaluation with the aim of selecting a lead radiotracer for translation into human PET imaging trials. We demonstrate that [(18)F]N-methyl lansoprazole, on account of the favorable half-life of fluorine-18 and its rapid brain entry in nonhuman primates, favorable kinetics, low white matter binding, and selectivity for binding to tau over amyloid, is the lead compound for progression into clinical trials.

Occupation of low-affinity cholecystokinin (CCK) receptors by CCK activates signal transduction and stimulates amylase secretion in pancreatic acinar cells.

Science.gov (United States)

Vinayek, R; Patto, R J; Menozzi, D; Gregory, J; Mrozinski, J E; Jensen, R T; Gardner, J D

1993-03-10

Based on the effects of monensin on binding of 125I-CCK-8 and its lack of effect on CCK-8-stimulated amylase secretion we previously proposed that pancreatic acinar cells possess three classes of CCK receptors: high-affinity receptors, low-affinity receptors and very low-affinity receptors [1]. In the present study we treated pancreatic acini with carbachol to induce a complete loss of high-affinity CCK receptors and then examined the action of CCK-8 on inositol trisphosphate IP3(1,4,5), cytosolic calcium and amylase secretion in an effort to confirm and extend our previous hypothesis. We found that first incubating pancreatic acini with 10 mM carbachol decreased binding of 125I-CCK-8 measured during a second incubation by causing a complete loss of high-affinity CCK receptors with no change in the low-affinity CCK receptors. Carbachol treatment of acini, however, did not alter the action of CCK-8 on IP3(1,4,5), cytosolic calcium or amylase secretion or the action of CCK-JMV-180 on amylase secretion or on the supramaximal inhibition of amylase secretion caused by CCK-8. The present findings support our previous hypothesis that pancreatic acinar cells possess three classes of CCK receptors and suggest that high-affinity CCK receptors do not mediate the action of CCK-8 on enzyme secretion, that low-affinity CCK receptors may mediate the action of CCK on cytosolic calcium that does not involve IP3(1,4,5) and produce the upstroke of the dose-response curve for CCK-8-stimulated amylase secretion and that very low-affinity CCK receptors mediate the actions of CCK on IP3(1,4,5) and cytosolic calcium and produce the downstroke of the dose-response curve for CCK-8-stimulated amylase secretion. Moreover, CCK-JMV-180 is a full agonist for stimulating amylase secretion by acting at low-affinity CCK receptors and is an antagonist at very low-affinity CCK receptors.

New unitary affine-Virasoro constructions

International Nuclear Information System (INIS)

Halpern, M.B.; Kiritsis, E.; Obers, N.A.; Poratti, M.; Yamron, J.P.

1990-01-01

This paper reports on a quasi-systematic investigation of the Virasoro master equation. The space of all affine-Virasoro constructions is organized by K-conjugation into affine-Virasoro nests, and an estimate of the dimension of the space shows that most solutions await discovery. With consistent ansatze for the master equation, large classes of new unitary nests are constructed, including quadratic deformation nests with continuous conformal weights, and unitary irrational central charge nests, which may dominate unitary rational central charge on compact g

Development of an aptamer-based affinity purification method for vascular endothelial growth factor

Directory of Open Access Journals (Sweden)

Maren Lönne

2015-12-01

Full Text Available Since aptamers bind their targets with high affinity and specificity, they are promising alternative ligands in protein affinity purification. As aptamers are chemically synthesized oligonucleotides, they can be easily produced in large quantities regarding GMP conditions allowing their application in protein production for therapeutic purposes. Several advantages of aptamers compared to antibodies are described in general within this paper. Here, an aptamer directed against the human Vascular Endothelial Growth Factor (VEGF was used as affinity ligand for establishing a purification platform for VEGF in small scale. The aptamer was covalently immobilized on magnetic beads in a controlled orientation resulting in a functional active affinity matrix. Target binding was optimized by introduction of spacer molecules and variation of aptamer density. Further, salt-induced target elution was demonstrated as well as VEGF purification from a complex protein mixture proving the specificity of protein-aptamer binding.

PHARMACEUTICAL AND BIOMEDICAL APPLICATIONS OF AFFINITY CHROMATOGRAPHY: RECENT TRENDS AND DEVELOPMENTS

Science.gov (United States)

Hage, David S.; Anguizola, Jeanethe A.; Bi, Cong; Li, Rong; Matsuda, Ryan; Papastavros, Efthimia; Pfaunmiller, Erika; Vargas, John; Zheng, Xiwei

2012-01-01

Affinity chromatography is a separation technique that has become increasingly important in work with biological samples and pharmaceutical agents. This method is based on the use of a biologically-related agent as a stationary phase to selectively retain analytes or to study biological interactions. This review discusses the basic principles behind affinity chromatography and examines recent developments that have occurred in the use of this method for biomedical and pharmaceutical analysis. Techniques based on traditional affinity supports are discussed, but an emphasis is placed on methods in which affinity columns are used as part of HPLC systems or in combination with other analytical methods. General formats for affinity chromatography that are considered include step elution schemes, weak affinity chromatography, affinity extraction and affinity depletion. Specific separation techniques that are examined include lectin affinity chromatography, boronate affinity chromatography, immunoaffinity chromatography, and immobilized metal ion affinity chromatography. Approaches for the study of biological interactions by affinity chromatography are also presented, such as the measurement of equilibrium constants, rate constants, or competition and displacement effects. In addition, related developments in the use of immobilized enzyme reactors, molecularly imprinted polymers, dye ligands and aptamers are briefly considered. PMID:22305083

Contrast-enhanced CT with a High-Affinity Cationic Contrast Agent for Imaging ex Vivo Bovine, Intact ex Vivo Rabbit, and in Vivo Rabbit Cartilage

OpenAIRE

Stewart, Rachel C.; Bansal, Prashant N.; Entezari, Vahid; Lusic, Hrvoje; Nazarian, Rosalynn M.; Snyder, Brian D.; Grinstaff, Mark W.

2013-01-01

The high affinity of a cationic iodinated contrast agent for cartilage provides better tissue visualization, easier segmentation, higher contrast-to-noise ratios, and longer usable imaging windows and requires a lower dose of injected contrast agent compared with an anionic contrast agent.

Synthesis, Biodistribution and In vitro Evaluation of Brain Permeable High Affinity Type 2 Cannabinoid Receptor Agonists [11C]MA2 and [18F]MA3.

Science.gov (United States)

Ahamed, Muneer; van Veghel, Daisy; Ullmer, Christoph; Van Laere, Koen; Verbruggen, Alfons; Bormans, Guy M

2016-01-01

The type 2 cannabinoid receptor (CB2) is a member of the endocannabinoid system and is known for its important role in (neuro)inflammation. A PET-imaging agent that allows in vivo visualization of CB2 expression may thus allow quantification of neuroinflammation. In this paper, we report the synthesis, radiosynthesis, biodistribution and in vitro evaluation of a carbon-11 ([ 11 C]MA2) and a fluorine-18 ([ 18 F]MA3) labeled analog of a highly potent N -arylamide oxadiazole CB2 agonist (EC 50 = 0.015 nM). MA2 and MA3 behaved as potent CB2 agonist (EC 50 : 3 nM and 0.1 nM, respectively) and their in vitro binding affinity for h CB2 was found to be 87 nM and 0.8 nM, respectively. Also MA3 (substituted with a fluoro ethyl group) was found to have higher binding affinity and EC 50 values when compared to the originally reported trifluoromethyl analog 12 . [ 11 C]MA2 and [ 18 F]MA3 were successfully synthesized with good radiochemical yield, high radiochemical purity and high specific activity. In mice, both tracers were efficiently cleared from blood and all major organs by the hepatobiliary pathway and importantly these compounds showed high brain uptake. In conclusion, [ 11 C]MA2 and [ 18 F]MA3 are shown to be high potent CB2 agonists with good brain uptake, these favorable characteristics makes them potential PET probes for in vivo imaging of brain CB2 receptors. However, in view of its higher affinity and selectivity, further detailed evaluation of MA3 as a PET tracer for CB2 is warranted.

Affine stochastic mortality

NARCIS (Netherlands)

Schrager, D.F.

2006-01-01

We propose a new model for stochastic mortality. The model is based on the literature on affine term structure models. It satisfies three important requirements for application in practice: analytical tractibility, clear interpretation of the factors and compatibility with financial option pricing

An Integrative, Multi-Scale Computational Model of a Swimming Lamprey Fully Coupled to Its Fluid Environment and Incorporating Proprioceptive Feedback

Science.gov (United States)

Hamlet, C. L.; Hoffman, K.; Fauci, L.; Tytell, E.

2016-02-01

The lamprey is a model organism for both neurophysiology and locomotion studies. To study the role of sensory feedback as an organism moves through its environment, a 2D, integrative, multi-scale model of an anguilliform swimmer driven by neural activation from a central pattern generator (CPG) is constructed. The CPG in turn drives muscle kinematics and is fully coupled to the surrounding fluid. The system is numerically evolved in time using an immersed boundary framework producing an emergent swimming mode. Proprioceptive feedback to the CPG based on experimental observations adjust the activation signal as the organism interacts with its environment. Effects on the speed, stability and cost (metabolic work) of swimming due to nonlinear dependencies associated with muscle force development combined with proprioceptive feedback to neural activation are estimated and examined.

Exploiting the high-affinity phosphonate-hydroxyapatite nanoparticle interaction for delivery of radiation and drugs

International Nuclear Information System (INIS)

Ong, Hooi Tin; Loo, Joachim S. C.; Boey, Freddy Y. C.; Russell, Stephen J.; Ma Jan; Peng, Kah-Whye

2008-01-01

Hydroxyapatite is biocompatible and used in various biomedical applications. Here, we generated hydroxyapatite nanoparticles (HNPs) of various sizes (40-200 nm) and demonstrated that they can be stably loaded with drugs or radioisotopes by exploiting the high-affinity HA-(poly)phosphonate interaction. Clinically available phosphonates, clodronate, and Tc-99m-methylene-diphosphonate (Tc-99m-MDP), were efficiently loaded onto HNPs within 15 min. Biodistribution of radiolabeled HNP-MDP-Tc99m in mice was monitored non-invasively using microSPECT-CT. Imaging and dosimetry studies indicated that the HNPs, regardless of size, were quickly taken up by Kupffer cells in the liver after systemic administration into mice. Clodronate loaded onto HNPs remained biologically active and were able to result in selective depletion of Kupffer cells. This method of drug or isotope loading on HA is fast and easy as it eliminates the need for additional surface modifications of the nanoparticles

Rank Two Affine Manifolds in Genus 3

OpenAIRE

Aulicino, David; Nguyen, Duc-Manh

2016-01-01

We complete the classification of rank two affine manifolds in the moduli space of translation surfaces in genus three. Combined with a recent result of Mirzakhani and Wright, this completes the classification of higher rank affine manifolds in genus three.

Maximum-Entropy Models of Sequenced Immune Repertoires Predict Antigen-Antibody Affinity.

Directory of Open Access Journals (Sweden)

Lorenzo Asti

2016-04-01

Full Text Available The immune system has developed a number of distinct complex mechanisms to shape and control the antibody repertoire. One of these mechanisms, the affinity maturation process, works in an evolutionary-like fashion: after binding to a foreign molecule, the antibody-producing B-cells exhibit a high-frequency mutation rate in the genome region that codes for the antibody active site. Eventually, cells that produce antibodies with higher affinity for their cognate antigen are selected and clonally expanded. Here, we propose a new statistical approach based on maximum entropy modeling in which a scoring function related to the binding affinity of antibodies against a specific antigen is inferred from a sample of sequences of the immune repertoire of an individual. We use our inference strategy to infer a statistical model on a data set obtained by sequencing a fairly large portion of the immune repertoire of an HIV-1 infected patient. The Pearson correlation coefficient between our scoring function and the IC50 neutralization titer measured on 30 different antibodies of known sequence is as high as 0.77 (p-value 10-6, outperforming other sequence- and structure-based models.

Multilayer affinity adsorption of albumin on polymer brushes modified membranes in a continuous-flow system.

Science.gov (United States)

Hu, Meng-Xin; Li, Xiang; Li, Ji-Nian; Huang, Jing-Jing; Ren, Ge-Rui

2018-02-23

Polymer brushes modified surfaces have been widely used for protein immobilization and isolation. Modification of membranes with polymer brushes increases the surface concentration of affinity ligands used for protein binding. Albumin is one of the transporting proteins and shows a high affinity to bile acids. In this work, the modified membranes with cholic acid-containing polymer brushes can be facilely prepared by the immobilization of cholic acid on the poly(2-hydroxyethyl methacrylate) grafted microporous polypropylene membranes (MPPMs) for affinity adsorption of albumin. ATR/FT-IR and X-ray photoelectron spectroscopy were used to characterize the chemical composition of the modified membranes. Water contact angle measurements were used to analyze the hydrophilic/hydrophobic properties of the membrane surface. The modified MPPMs show a high affinity to albumin and have little non-specific adsorption of hemoglobin. The dynamic binding capacity of albumin in the continous-flow system increases with the cycle number and feed rate as the binding degree of cholic acid is moderate. The highest binding capacity of affinity membranes is about 52.49 g/m 2 membrane, which is about 24 times more than the monolayer binding capacity. These results reveal proteins could be captured in multilayers by the polymer brushes containing affinity ligands similar to the polymer brushes containing ion-exchange groups, which open up the potential of the polymer brushes containing affinity ligands in protein or another components separation. And the cholic acid containing polymer brushes modified membranes has the promising potential for albumin separation and purification rapidly from serum or fermented solution in medical diagnosis and bioseparation. Copyright © 2018 Elsevier B.V. All rights reserved.

High-affinity hemoglobin and blood oxygen saturation in diving emperor penguins.

Science.gov (United States)

Meir, Jessica U; Ponganis, Paul J

2009-10-01

The emperor penguin (Aptenodytes forsteri) thrives in the Antarctic underwater environment, diving to depths greater than 500 m and for durations longer than 23 min. To examine mechanisms underlying the exceptional diving ability of this species and further describe blood oxygen (O2) transport and depletion while diving, we characterized the O2-hemoglobin (Hb) dissociation curve of the emperor penguin in whole blood. This allowed us to (1) investigate the biochemical adaptation of Hb in this species, and (2) address blood O2 depletion during diving, by applying the dissociation curve to previously collected partial pressure of O2 (PO2) profiles to estimate in vivo Hb saturation (SO2) changes during dives. This investigation revealed enhanced Hb-O2 affinity (P50=28 mmHg, pH 7.5) in the emperor penguin, similar to high-altitude birds and other penguin species. This allows for increased O2 at low blood PO2 levels during diving and more complete depletion of the respiratory O2 store. SO2 profiles during diving demonstrated that arterial SO2 levels are maintained near 100% throughout much of the dive, not decreasing significantly until the final ascent phase. End-of-dive venous SO2 values were widely distributed and optimization of the venous blood O2 store resulted from arterialization and near complete depletion of venous blood O2 during longer dives. The estimated contribution of the blood O2 store to diving metabolic rate was low and highly variable. This pattern is due, in part, to the influx of O2 from the lungs into the blood during diving, and variable rates of tissue O2 uptake.

Food and value motivation: Linking consumer affinities to different types of food products.

Science.gov (United States)

de Boer, Joop; Schösler, Hanna

2016-08-01

This study uses the consumer affinity concept to examine the multiple motives that may shape consumers' relationships with food. The concept was applied in a study on four broad product types in the Netherlands, which cover a wide range of the market and may each appeal to consumers with different affinities towards foods. These product types may be denoted as 'conventional', 'efficient', 'gourmet' and 'pure'. A comparative analysis, based on Higgins' Regulatory Focus Theory, was performed to examine whether food-related value motivations could explain different consumer affinities for these product types. The affinities of consumers were measured by means of a non-verbal, visual presentation of four samples of food products in a nationwide survey (n = 742) among consumers who were all involved in food purchasing and/or cooking. The affinities found could be predicted fairly well from a number of self-descriptions relating to food and eating, which expressed different combinations of type of value motivation and involvement with food. The analysis demonstrated the contrasting role of high and low involvement as well as the potential complementarity of promotion- and prevention-focused value motivation. It is suggested that knowledge of the relationships between product types, consumer affinities and value motivation can help improve the effectiveness of interventions that seek to promote healthy and sustainable diets in developed countries. Copyright © 2016 Elsevier Ltd. All rights reserved.

Dynamics of Open Systems with Affine Maps

International Nuclear Information System (INIS)

Zhang Da-Jian; Liu Chong-Long; Tong Dian-Min

2015-01-01

Many quantum systems of interest are initially correlated with their environments and the reduced dynamics of open systems are an interesting while challenging topic. Affine maps, as an extension of completely positive maps, are a useful tool to describe the reduced dynamics of open systems with initial correlations. However, it is unclear what kind of initial state shares an affine map. In this study, we give a sufficient condition of initial states, in which the reduced dynamics can always be described by an affine map. Our result shows that if the initial states of the combined system constitute a convex set, and if the correspondence between the initial states of the open system and those of the combined system, defined by taking the partial trace, is a bijection, then the reduced dynamics of the open system can be described by an affine map. (paper)

USING MICROSCALE THERMOPHORESIS TO EASILY MEASURE BINDING AFFINITY

Directory of Open Access Journals (Sweden)

Dennis Breitsprecher*

2018-03-01

Full Text Available While it’s very common for biologists and chemists to test whether or not two molecules interact with each other, it’s much more useful to gather information on the nature of that interaction. How strong is it? How long will it last? What does that mean for its biological function? One way to answer these questions is to study affinity. Binding affinity is defined as the strength of the binding interaction between a single biomolecule to its binding partner, or ligand, and it can be quantifiably measured, providing information on whether or not molecules are interacting, as well as assigning a value to the affinity. When measuring binding affinity, there are several parameters to look at, but the dissociation constant (Kd, which defines the likelihood that an interaction between two molecules will break, is a very common measurement. The smaller the dissociation constant, the more tightly bound the ligand is, and the higher the affinity is between the two molecules.

Single-Step Affinity Purification for Fungal Proteomics ▿ †

OpenAIRE

Liu, Hui-Lin; Osmani, Aysha H.; Ukil, Leena; Son, Sunghun; Markossian, Sarine; Shen, Kuo-Fang; Govindaraghavan, Meera; Varadaraj, Archana; Hashmi, Shahr B.; De Souza, Colin P.; Osmani, Stephen A.

2010-01-01

A single-step protein affinity purification protocol using Aspergillus nidulans is described. Detailed protocols for cell breakage, affinity purification, and depending on the application, methods for protein release from affinity beads are provided. Examples defining the utility of the approaches, which should be widely applicable, are included.

Blood O2 affinity of a large polar elasmobranch, the Greenland shark Somniosus microcephalus

DEFF Research Database (Denmark)

Herbert, N.A.; Skov, Peter Vilhelm; Tirsgaard, B.

2017-01-01

The Greenland shark (Somniosus microcephalus. Bloch & Schneider 1801) is a polar elasmobranch that is hypothesised to possess a unique metabolic physiology due to its extreme large size, the cold waters it inhabits and its slow swimming lifestyle. Our results therefore provide the first insight...... into the metabolic physiology of this unique shark, with a focus on blood O2 affinity. An evaluation of blood O2 affinity at 2 °C using tonometry revealed a P50 of 11.7 mmHg at a PCO2 of 2.25 mmHg and a Bohr effect (binding sensitivity of blood to pH, ϕ = Δlog P50/ΔpH) of −0.26. A comparative evaluation of blood O2...... affinity across elasmobranch fishes suggests that S. microcephalus has a high blood O2 affinity (i.e., low P50) and a small Bohr effect but these are common traits in sluggish elasmobranch fishes, with little evidence for any relationship of blood O2 affinity to the low metabolic rates, low environmental...

The Structure of Affine Buildings

CERN Document Server

Weiss, Richard M

2009-01-01

In The Structure of Affine Buildings, Richard Weiss gives a detailed presentation of the complete proof of the classification of Bruhat-Tits buildings first completed by Jacques Tits in 1986. The book includes numerous results about automorphisms, completions, and residues of these buildings. It also includes tables correlating the results in the locally finite case with the results of Tits's classification of absolutely simple algebraic groups defined over a local field. A companion to Weiss's The Structure of Spherical Buildings, The Structure of Affine Buildings is organized around the clas

The dynamics of metric-affine gravity

International Nuclear Information System (INIS)

Vitagliano, Vincenzo; Sotiriou, Thomas P.; Liberati, Stefano

2011-01-01

Highlights: → The role and the dynamics of the connection in metric-affine theories is explored. → The most general second order action does not lead to a dynamical connection. → Including higher order invariants excites new degrees of freedom in the connection. → f(R) actions are also discussed and shown to be a non- representative class. - Abstract: Metric-affine theories of gravity provide an interesting alternative to general relativity: in such an approach, the metric and the affine (not necessarily symmetric) connection are independent quantities. Furthermore, the action should include covariant derivatives of the matter fields, with the covariant derivative naturally defined using the independent connection. As a result, in metric-affine theories a direct coupling involving matter and connection is also present. The role and the dynamics of the connection in such theories is explored. We employ power counting in order to construct the action and search for the minimal requirements it should satisfy for the connection to be dynamical. We find that for the most general action containing lower order invariants of the curvature and the torsion the independent connection does not carry any dynamics. It actually reduces to the role of an auxiliary field and can be completely eliminated algebraically in favour of the metric and the matter field, introducing extra interactions with respect to general relativity. However, we also show that including higher order terms in the action radically changes this picture and excites new degrees of freedom in the connection, making it (or parts of it) dynamical. Constructing actions that constitute exceptions to this rule requires significant fine tuned and/or extra a priori constraints on the connection. We also consider f(R) actions as a particular example in order to show that they constitute a distinct class of metric-affine theories with special properties, and as such they cannot be used as representative toy

Interactions of dopaminergic agonists and antagonists with dopaminergic D3 binding sites in rat striatum. Evidence that [3H]dopamine can label a high affinity agonist-binding state of the D1 dopamine receptor

International Nuclear Information System (INIS)

Leff, S.E.; Creese, I.

1985-01-01

The interactions of dopaminergic agonists and antagonists with 3 H-agonist labeled D3 dopaminergic binding sites of rat striatum have been characterized by radioligand-binding techniques. When the binding of [ 3 H]dopamine and [ 3 H]apomorphine to D2 dopamine receptors is blocked by the inclusion of D2 selective concentrations of unlabeled spiroperidol or domperidone, these ligands appear to label selectively the previously termed D3 binding site. Antagonist/[ 3 H]dopamine competition curves are of uniformly steep slope (nH . 1.0), suggesting the presence of a single D3 binding site. The relative potencies of antagonists to inhibit D3 specific [ 3 H]dopamine binding are significantly correlated with their potencies to block D1 dopamine receptors as measured by the inhibition of both dopamine-stimulated adenylate cyclase and [ 3 H]flupentixol-binding activities. The affinities of agonists to inhibit D3 specific [ 3 H]dopamine binding are also correlated with estimates of these agonists affinities for the high affinity binding component of agonist/[ 3 H]flupentixol competition curves. Both D3 specific [ 3 H] dopamine binding and the high affinity agonist-binding component of dopamine/[ 3 H]flupentixol competition curves show a similar sensitivity to guanine nucleotides. Taken together, these data strongly suggest that the D3 binding site is related to a high affinity agonist-binding state of the D1 dopamine receptor

Spectral affinity in protein networks.

Science.gov (United States)

Voevodski, Konstantin; Teng, Shang-Hua; Xia, Yu

2009-11-29

Protein-protein interaction (PPI) networks enable us to better understand the functional organization of the proteome. We can learn a lot about a particular protein by querying its neighborhood in a PPI network to find proteins with similar function. A spectral approach that considers random walks between nodes of interest is particularly useful in evaluating closeness in PPI networks. Spectral measures of closeness are more robust to noise in the data and are more precise than simpler methods based on edge density and shortest path length. We develop a novel affinity measure for pairs of proteins in PPI networks, which uses personalized PageRank, a random walk based method used in context-sensitive search on the Web. Our measure of closeness, which we call PageRank Affinity, is proportional to the number of times the smaller-degree protein is visited in a random walk that restarts at the larger-degree protein. PageRank considers paths of all lengths in a network, therefore PageRank Affinity is a precise measure that is robust to noise in the data. PageRank Affinity is also provably related to cluster co-membership, making it a meaningful measure. In our experiments on protein networks we find that our measure is better at predicting co-complex membership and finding functionally related proteins than other commonly used measures of closeness. Moreover, our experiments indicate that PageRank Affinity is very resilient to noise in the network. In addition, based on our method we build a tool that quickly finds nodes closest to a queried protein in any protein network, and easily scales to much larger biological networks. We define a meaningful way to assess the closeness of two proteins in a PPI network, and show that our closeness measure is more biologically significant than other commonly used methods. We also develop a tool, accessible at http://xialab.bu.edu/resources/pnns, that allows the user to quickly find nodes closest to a queried vertex in any protein

Genetically based low oxygen affinities of felid hemoglobins: lack of biochemical adaptation to high-altitude hypoxia in the snow leopard

Science.gov (United States)

Janecka, Jan E.; Nielsen, Simone S. E.; Andersen, Sidsel D.; Hoffmann, Federico G.; Weber, Roy E.; Anderson, Trevor; Storz, Jay F.; Fago, Angela

2015-01-01

ABSTRACT Genetically based modifications of hemoglobin (Hb) function that increase blood–O2 affinity are hallmarks of hypoxia adaptation in vertebrates. Among mammals, felid Hbs are unusual in that they have low intrinsic O2 affinities and reduced sensitivities to the allosteric cofactor 2,3-diphosphoglycerate (DPG). This combination of features compromises the acclimatization capacity of blood–O2 affinity and has led to the hypothesis that felids have a restricted physiological niche breadth relative to other mammals. In seeming defiance of this conjecture, the snow leopard (Panthera uncia) has an extraordinarily broad elevational distribution and occurs at elevations above 6000 m in the Himalayas. Here, we characterized structural and functional variation of big cat Hbs and investigated molecular mechanisms of Hb adaptation and allosteric regulation that may contribute to the extreme hypoxia tolerance of the snow leopard. Experiments revealed that purified Hbs from snow leopard and African lion exhibited equally low O2 affinities and DPG sensitivities. Both properties are primarily attributable to a single amino acid substitution, β2His→Phe, which occurred in the common ancestor of Felidae. Given the low O2 affinity and reduced regulatory capacity of feline Hbs, the extreme hypoxia tolerance of snow leopards must be attributable to compensatory modifications of other steps in the O2-transport pathway. PMID:26246610

High Affinity Radiopharmaceuticals Based Upon Lansoprazole for PET Imaging of Aggregated Tau in Alzheimer’s Disease and Progressive Supranuclear Palsy: Synthesis, Preclinical Evaluation, and Lead Selection

Science.gov (United States)

2014-01-01

Abnormally aggregated tau is the hallmark pathology of tauopathy neurodegenerative disorders and is a target for development of both diagnostic tools and therapeutic strategies across the tauopathy disease spectrum. Development of carbon-11- or fluorine-18-labeled radiotracers with appropriate affinity and specificity for tau would allow noninvasive quantification of tau burden using positron emission tomography (PET) imaging. We have synthesized [18F]lansoprazole, [11C]N-methyl lansoprazole, and [18F]N-methyl lansoprazole and identified them as high affinity radiotracers for tau with low to subnanomolar binding affinities. Herein, we report radiosyntheses and extensive preclinical evaluation with the aim of selecting a lead radiotracer for translation into human PET imaging trials. We demonstrate that [18F]N-methyl lansoprazole, on account of the favorable half-life of fluorine-18 and its rapid brain entry in nonhuman primates, favorable kinetics, low white matter binding, and selectivity for binding to tau over amyloid, is the lead compound for progression into clinical trials. PMID:24896980

Multiprocessor Real-Time Scheduling with Hierarchical Processor Affinities

OpenAIRE

Bonifaci , Vincenzo; Brandenburg , Björn; D'Angelo , Gianlorenzo; Marchetti-Spaccamela , Alberto

2016-01-01

International audience; Many multiprocessor real-time operating systems offer the possibility to restrict the migrations of any task to a specified subset of processors by setting affinity masks. A notion of " strong arbitrary processor affinity scheduling " (strong APA scheduling) has been proposed; this notion avoids schedulability losses due to overly simple implementations of processor affinities. Due to potential overheads, strong APA has not been implemented so far in a real-time operat...

Influence of bone affinity on the skeletal distribution of fluorescently labeled bisphosphonates in vivo.

Science.gov (United States)

Roelofs, Anke J; Stewart, Charlotte A; Sun, Shuting; Błażewska, Katarzyna M; Kashemirov, Boris A; McKenna, Charles E; Russell, R Graham G; Rogers, Michael J; Lundy, Mark W; Ebetino, Frank H; Coxon, Fraser P

2012-04-01

Bisphosphonates are widely used antiresorptive drugs that bind to calcium. It has become evident that these drugs have differing affinities for bone mineral; however, it is unclear whether such differences affect their distribution on mineral surfaces. In this study, fluorescent conjugates of risedronate, and its lower-affinity analogues deoxy-risedronate and 3-PEHPC, were used to compare the localization of compounds with differing mineral affinities in vivo. Binding to dentine in vitro confirmed differences in mineral binding between compounds, which was influenced predominantly by the characteristics of the parent compound but also by the choice of fluorescent tag. In growing rats, all compounds preferentially bound to forming endocortical as opposed to resorbing periosteal surfaces in cortical bone, 1 day after administration. At resorbing surfaces, lower-affinity compounds showed preferential binding to resorption lacunae, whereas the highest-affinity compound showed more uniform labeling. At forming surfaces, penetration into the mineralizing osteoid was found to inversely correlate with mineral affinity. These differences in distribution at resorbing and forming surfaces were not observed at quiescent surfaces. Lower-affinity compounds also showed a relatively higher degree of labeling of osteocyte lacunar walls and labeled lacunae deeper within cortical bone, indicating increased penetration of the osteocyte canalicular network. Similar differences in mineralizing surface and osteocyte network penetration between high- and low-affinity compounds were evident 7 days after administration, with fluorescent conjugates at forming surfaces buried under a new layer of bone. Fluorescent compounds were incorporated into these areas of newly formed bone, indicating that "recycling" had occurred, albeit at very low levels. Taken together, these findings indicate that the bone mineral affinity of bisphosphonates is likely to influence their distribution within the

Specific, high affinity receptors for insulin-like growth factor II in the rat kidney glomerulus

International Nuclear Information System (INIS)

Haskell, J.F.; Pillion, D.J.; Meezan, E.

1988-01-01

Rat renal glomeruli were isolated by a technique involving kidney perfusion with a solution containing magnetic iron oxide particles, followed by homogenization, sieving, and concentration over a strong magnet. Isolated glomeruli were treated with 1% Triton X-100 to solubilize plasma membrane components, while insoluble basement membrane components were removed by centrifugation. [ 125 I]Insulin-like growth factor II (IGF-II) binding to this preparation was competitively inhibited by increasing amounts of unlabeled IGF-II, with 50% inhibition at an IGF-II concentration of 1 ng/ml. [ 125 I]IGF-II was covalently cross-linked with disuccinimidyl suberate to its receptor in rat renal glomeruli and a specific high mol wt (255,000) band could be identified on autoradiograms of dodecyl sulfate-polyacrylamide gels. [ 125 I]IGF-II binding and cross-linking to this band was inhibited by a polyclonal antibody against the type II IGF receptor. These results demonstrate for the first time that the isolated rat renal glomerulus contains a high affinity receptor for IGF-II

Simulated electron affinity tuning in metal-insulator-metal (MIM) diodes

Science.gov (United States)

Mistry, Kissan; Yavuz, Mustafa; Musselman, Kevin P.

2017-05-01

Metal-insulator-metal diodes for rectification applications must exhibit high asymmetry, nonlinearity, and responsivity. Traditional methods of improving these figures of merit have consisted of increasing insulator thickness, adding multiple insulator layers, and utilizing a variety of metal contact combinations. However, these methods have come with the price of increasing the diode resistance and ultimately limiting the operating frequency to well below the terahertz regime. In this work, an Airy Function Transfer Matrix simulation method was used to observe the effect of tuning the electron affinity of the insulator as a technique to decrease the diode resistance. It was shown that a small increase in electron affinity can result in a resistance decrease in upwards of five orders of magnitude, corresponding to an increase in operating frequency on the same order. Electron affinity tuning has a minimal effect on the diode figures of merit, where asymmetry improves or remains unaffected and slight decreases in nonlinearity and responsivity are likely to be greatly outweighed by the improved operating frequency of the diode.

Phylogenetic distribution of a male pheromone that may exploit a nonsexual preference in lampreys.

Science.gov (United States)

Buchinger, T J; Bussy, U; Li, K; Wang, H; Huertas, M; Baker, C F; Jia, L; Hayes, M C; Li, W; Johnson, N S

2017-12-01

Pheromones are among the most important sexual signals used by organisms throughout the animal kingdom. However, few are identified in vertebrates, leaving the evolutionary mechanisms underlying vertebrate pheromones poorly understood. Pre-existing biases in receivers' perceptual systems shape visual and auditory signalling systems, but studies on how receiver biases influence the evolution of pheromone communication remain sparse. The lamprey Petromyzon marinus uses a relatively well-understood suite of pheromones and offers a unique opportunity to study the evolution of vertebrate pheromone communication. Previous studies indicate that male signalling with the mating pheromone 3-keto petromyzonol sulphate (3kPZS) may exploit a nonsexual attraction to juvenile-released 3kPZS that guides migration into productive rearing habitat. Here, we infer the distribution of male signalling with 3kPZS using a phylogenetic comparison comprising six of 10 genera and two of three families. Our results indicate that only P. marinus and Ichthyomyzon castaneus release 3kPZS at high rates. Olfactory and behavioural assays with P. marinus, I. castaneus and a subset of three other species that do not use 3kPZS as a sexual signal indicate that male signalling might have driven the evolution of female adaptations to detect 3kPZS with specific olfactory mechanisms and respond to 3kPZS with targeted attraction relevant during mate search. We postulate that 3kPZS communication evolved independently in I. castaneus and P. marinus, but cannot eliminate the alternative that other species lost 3kPZS communication. Regardless, our results represent a rare macroevolutionary investigation of a vertebrate pheromone and provide insight into the evolutionary mechanisms underlying pheromone communication. © 2017 European Society For Evolutionary Biology. Journal of Evolutionary Biology © 2017 European Society For Evolutionary Biology.

Synthesis, biodistribution and in vitro evaluation of brain permeable high affinity type 2 cannabinoid receptor agonists [11C]MA2 and [18F]MA3

Directory of Open Access Journals (Sweden)

Muneer Ahamed

2016-09-01

Full Text Available Abstract The type 2 cannabinoid receptor (CB2 is a member of the endocannabinoid system and is known for its important role in (neuroinflammation. A PET-imaging agent that allows in vivo visualization of CB2 expression may thus allow quantification of neuroinflammation. In this paper, we report the synthesis, radiosynthesis, biodistribution and in vitro evaluation of a carbon-11 ([11C]MA2 and a fluorine-18 ([18F]MA3 labeled analogue of a highly potent N-arylamide oxadiazole CB2 agonist (EC50 = 0.015 nM. MA2 and MA3 behaved as potent CB2 agonist (EC50: 3 nM and 0.1 nM, respectively and their in vitro binding affinity for hCB2 was found to be 87 nM and 0.8 nM, respectively. Also MA3 (substituted with a fluoro ethyl group was found to have higher binding affinity and EC50 values when compared to the originally reported trifluoromethyl analogue 12. [11C]MA2 and [18F]MA3 were successfully synthesized with good radiochemical yield, high radiochemical purity and high specific activity. In mice, both tracers were efficiently cleared from blood and all major organs by the hepatobiliary pathway and importantly these compounds showed high brain uptake. In conclusion, [11C]MA2 and [18F]MA3 are shown to be high potent CB2 agonists with good brain uptake, these favorable characteristics makes them potential PET probes for in vivo imaging of brain CB2 receptors. However in view of its higher affinity and selectivity, further detailed evaluation of MA3 as a PET tracer for CB2 is warranted.

Affine fractal functions as bases of continuous funtions | Navascues ...

African Journals Online (AJOL)

The objective of the present paper is the study of affine transformations of the plane, which provide self-affine curves as attractors. The properties of these curves depend decisively of the coefficients of the system of affinities involved. The corresponding functions are continuous on a compact interval. If the scale factors are ...

Antibody Affinity Maturation in Fishes—Our Current Understanding

Directory of Open Access Journals (Sweden)

Brad G. Magor

2015-07-01