Laccase engineering: from rational design to directed evolution.

Science.gov (United States)

Mate, Diana M; Alcalde, Miguel

2015-01-01

Laccases are multicopper oxidoreductases considered by many in the biotechonology field as the ultimate "green catalysts". This is mainly due to their broad substrate specificity and relative autonomy (they use molecular oxygen from air as an electron acceptor and they only produce water as by-product), making them suitable for a wide array of applications: biofuel production, bioremediation, organic synthesis, pulp biobleaching, textiles, the beverage and food industries, biosensor and biofuel cell development. Since the beginning of the 21st century, specific features of bacterial and fungal laccases have been exhaustively adapted in order to reach the industrial demands for high catalytic activity and stability in conjunction with reduced production cost. Among the goals established for laccase engineering, heterologous functional expression, improved activity and thermostability, tolerance to non-natural media (organic solvents, ionic liquids, physiological fluids) and resistance to different types of inhibitors are all challenges that have been met, while obtaining a more comprehensive understanding of laccase structure-function relationships. In this review we examine the most significant advances in this exciting research area in which rational, semi-rational and directed evolution approaches have been employed to ultimately convert laccases into high value-added biocatalysts. Copyright © 2014 Elsevier Inc. All rights reserved.

TGP, an extremely stable, non-aggregating fluorescent protein created by structure-guided surface engineering

Science.gov (United States)

Close, Devin W.; Don Paul, Craig; Langan, Patricia S.; Wilce, Matthew C.J.; Traore, Daouda A.K.; Halfmann, Randal; Rocha, Reginaldo C.; Waldo, Geoffery S.; Payne, Riley J.; Rucker, Joseph B.; Prescott, Mark; Bradbury, Andrew R.M.

2014-01-01

In this paper we describe the engineering and X-ray crystal structure of Thermal Green Protein (TGP), an extremely stable, highly soluble, non-aggregating green fluorescent protein. TGP is a soluble variant of the fluorescent protein eCGP123, which despite being highly stable, has proven to be aggregation-prone. The X-ray crystal structure of eCGP123, also determined within the context of this paper, was used to carry out rational surface engineering to improve its solubility, leading to TGP. The approach involved simultaneously eliminating crystal lattice contacts while increasing the overall negative charge of the protein. Despite intentional disruption of lattice contacts and introduction of high entropy glutamate side chains, TGP crystallized readily in a number of different conditions and the X-ray crystal structure of TGP was determined to 1.9 Å resolution. The structural reasons for the enhanced stability of TGP and eCGP123 are discussed. We demonstrate the utility of using TGP as a fusion partner in various assays and significantly, in amyloid assays in which the standard fluorescent protein, EGFP, is undesirable because of aberrant oligomerization. PMID:25287913

State-of-the-art protein engineering approaches using biological macromolecules: A review from immobilization to implementation view point.

Science.gov (United States)

Bilal, Muhammad; Iqbal, Hafiz M N; Guo, Shuqi; Hu, Hongbo; Wang, Wei; Zhang, Xuehong

2018-03-01

Over the past years, technological and scientific advances have proven biocatalysis as a sustainable alternative than traditional chemical catalysis including organo- or metallocatalysis. In this context, immobilization based approaches represent simple but effective routes for engineering enzyme catalysts with higher activities than wild-type derivatives. Many enzymes including oxidoreductases have been engineered by rational and directed evolution, to realize the catalytic activity, enantioselectivity, and stability attributes which are essential for their biotechnological exploitation. Induce yet stable activity in enzyme catalysis offer new insights and motivation to engineer efficient catalysts for practical and commercial purposes. It has now become possible to envisage substrate accessibility to the catalytic site of the enzyme by current computational capabilities that reduce the experimental work related to the enzyme selection, screening, and engineering. Herein, state-of-the-art protein engineering approaches for improving enzymatic activities including chemical modification, directed evolution, and rational design or their combination methods are discussed. The emphasis is also given to the applications of the resulting tailored catalysts ranging from fine chemicals to novel pharmaceutical compounds that use biocatalysts as a vital step. Copyright © 2017 Elsevier B.V. All rights reserved.

Protein engineering for metabolic engineering: current and next-generation tools

Science.gov (United States)

Marcheschi, Ryan J.; Gronenberg, Luisa S.; Liao, James C.

2014-01-01

Protein engineering in the context of metabolic engineering is increasingly important to the field of industrial biotechnology. As the demand for biologically-produced food, fuels, chemicals, food additives, and pharmaceuticals continues to grow, the ability to design and modify proteins to accomplish new functions will be required to meet the high productivity demands for the metabolism of engineered organisms. This article reviews advances of selecting, modeling, and engineering proteins to improve or alter their activity. Some of the methods have only recently been developed for general use and are just beginning to find greater application in the metabolic engineering community. We also discuss methods of generating random and targeted diversity in proteins to generate mutant libraries for analysis. Recent uses of these techniques to alter cofactor use, produce non-natural amino acids, alcohols, and carboxylic acids, and alter organism phenotypes are presented and discussed as examples of the successful engineering of proteins for metabolic engineering purposes. PMID:23589443

Artificially Engineered Protein Polymers.

Science.gov (United States)

Yang, Yun Jung; Holmberg, Angela L; Olsen, Bradley D

2017-06-07

Modern polymer science increasingly requires precise control over macromolecular structure and properties for engineering advanced materials and biomedical systems. The application of biological processes to design and synthesize artificial protein polymers offers a means for furthering macromolecular tunability, enabling polymers with dispersities of ∼1.0 and monomer-level sequence control. Taking inspiration from materials evolved in nature, scientists have created modular building blocks with simplified monomer sequences that replicate the function of natural systems. The corresponding protein engineering toolbox has enabled the systematic development of complex functional polymeric materials across areas as diverse as adhesives, responsive polymers, and medical materials. This review discusses the natural proteins that have inspired the development of key building blocks for protein polymer engineering and the function of these elements in material design. The prospects and progress for scalable commercialization of protein polymers are reviewed, discussing both technology needs and opportunities.

Protein engineering techniques gateways to synthetic protein universe

CERN Document Server

Poluri, Krishna Mohan

2017-01-01

This brief provides a broad overview of protein-engineering research, offering a glimpse of the most common experimental methods. It also presents various computational programs with applications that are widely used in directed evolution, computational and de novo protein design. Further, it sheds light on the advantages and pitfalls of existing methodologies and future perspectives of protein engineering techniques.

Protein Engineering: Case Studies of Commercialized Engineered Products

Science.gov (United States)

Walsh, Gary

2007-01-01

Programs in biochemistry invariably encompass the principles of protein engineering. Students often display increased understanding and enthusiasm when theoretical concepts are underpinned by practical example. Herein are presented five case studies, each focusing upon a commercial protein product engineered to enhance its application-relevant…

Protein design for pathway engineering.

Science.gov (United States)

Eriksen, Dawn T; Lian, Jiazhang; Zhao, Huimin

2014-02-01

Design and construction of biochemical pathways has increased the complexity of biosynthetically-produced compounds when compared to single enzyme biocatalysis. However, the coordination of multiple enzymes can introduce a complicated set of obstacles to overcome in order to achieve a high titer and yield of the desired compound. Metabolic engineering has made great strides in developing tools to optimize the flux through a target pathway, but the inherent characteristics of a particular enzyme within the pathway can still limit the productivity. Thus, judicious protein design is critical for metabolic and pathway engineering. This review will describe various strategies and examples of applying protein design to pathway engineering to optimize the flux through the pathway. The proteins can be engineered for altered substrate specificity/selectivity, increased catalytic activity, reduced mass transfer limitations through specific protein localization, and reduced substrate/product inhibition. Protein engineering can also be expanded to design biosensors to enable high through-put screening and to customize cell signaling networks. These strategies have successfully engineered pathways for significantly increased productivity of the desired product or in the production of novel compounds. Copyright © 2013 Elsevier Inc. All rights reserved.

An ontology-based search engine for protein-protein interactions.

Science.gov (United States)

Park, Byungkyu; Han, Kyungsook

2010-01-18

Keyword matching or ID matching is the most common searching method in a large database of protein-protein interactions. They are purely syntactic methods, and retrieve the records in the database that contain a keyword or ID specified in a query. Such syntactic search methods often retrieve too few search results or no results despite many potential matches present in the database. We have developed a new method for representing protein-protein interactions and the Gene Ontology (GO) using modified Gödel numbers. This representation is hidden from users but enables a search engine using the representation to efficiently search protein-protein interactions in a biologically meaningful way. Given a query protein with optional search conditions expressed in one or more GO terms, the search engine finds all the interaction partners of the query protein by unique prime factorization of the modified Gödel numbers representing the query protein and the search conditions. Representing the biological relations of proteins and their GO annotations by modified Gödel numbers makes a search engine efficiently find all protein-protein interactions by prime factorization of the numbers. Keyword matching or ID matching search methods often miss the interactions involving a protein that has no explicit annotations matching the search condition, but our search engine retrieves such interactions as well if they satisfy the search condition with a more specific term in the ontology.

Protein engineering approaches to chemical biotechnology.

Science.gov (United States)

Chen, Zhen; Zeng, An-Ping

2016-12-01

Protein engineering for the improvement of properties of biocatalysts and for the generation of novel metabolic pathways plays more and more important roles in chemical biotechnology aiming at the production of chemicals from biomass. Although widely used in single-enzyme catalysis process, protein engineering is only being increasingly explored in recent years to achieve more complex in vitro and in vivo biocatalytic processes. This review focuses on major contributions of protein engineering to chemical biotechnology in the field of multi-enzymatic cascade catalysis and metabolic engineering. Especially, we discuss and highlight recent strategies for combining pathway design and protein engineering for the production of novel products. Copyright © 2016. Published by Elsevier Ltd.

Protein engineering and its applications in food industry.

Science.gov (United States)

Kapoor, Swati; Rafiq, Aasima; Sharma, Savita

2017-07-24

Protein engineering is a young discipline that has been branched out from the field of genetic engineering. Protein engineering is based on the available knowledge about the proteins structure/function(s), tools/instruments, software, bioinformatics database, available cloned gene, knowledge about available protein, vectors, recombinant strains and other materials that could lead to change in the protein backbone. Protein produced properly from genetic engineering process means a protein that is able to fold correctly and to do particular function(s) efficiently even after being subjected to engineering practices. Protein is modified through its gene or chemically. However, modification of protein through gene is easier. There is no specific limitation of Protein Engineering tools; any technique that can lead to change the protein constituent of amino acid and result in the modification of protein structure/function is in the frame of Protein Engineering. Meanwhile, there are some common tools used to reach a specific target. More active industrial and pharmaceutical based proteins have been invented by the field of Protein Engineering to introduce new function as well as to change its interaction with surrounding environment. A variety of protein engineering applications have been reported in the literature. These applications range from biocatalysis for food and industry to environmental, medical and nanobiotechnology applications. Successful combinations of various protein engineering methods had led to successful results in food industries and have created a scope to maintain the quality of finished product after processing.

Integrating enzyme immobilization and protein engineering: An alternative path for the development of novel and improved industrial biocatalysts.

Science.gov (United States)

Bernal, Claudia; Rodríguez, Karen; Martínez, Ronny

2018-06-09

Enzyme immobilization often achieves reusable biocatalysts with improved operational stability and solvent resistance. However, these modifications are generally associated with a decrease in activity or detrimental modifications in catalytic properties. On the other hand, protein engineering aims to generate enzymes with increased performance at specific conditions by means of genetic manipulation, directed evolution and rational design. However, the achieved biocatalysts are generally generated as soluble enzymes, -thus not reusable- and their performance under real operational conditions is uncertain. Combined protein engineering and enzyme immobilization approaches have been employed as parallel or consecutive strategies for improving an enzyme of interest. Recent reports show efforts on simultaneously improving both enzymatic and immobilization components through genetic modification of enzymes and optimizing binding chemistry for site-specific and oriented immobilization. Nonetheless, enzyme engineering and immobilization are usually performed as separate workflows to achieve improved biocatalysts. In this review, we summarize and discuss recent research aiming to integrate enzyme immobilization and protein engineering and propose strategies to further converge protein engineering and enzyme immobilization efforts into a novel "immobilized biocatalyst engineering" research field. We believe that through the integration of both enzyme engineering and enzyme immobilization strategies, novel biocatalysts can be obtained, not only as the sum of independently improved intrinsic and operational properties of enzymes, but ultimately tailored specifically for increased performance as immobilized biocatalysts, potentially paving the way for a qualitative jump in the development of efficient, stable biocatalysts with greater real-world potential in challenging bioprocess applications. Copyright © 2018. Published by Elsevier Inc.

TRITON: graphic software for rational engineering of enzymes.

Science.gov (United States)

Damboský, J; Prokop, M; Koca, J

2001-01-01

Engineering of the catalytic properties of enzymes requires knowledge about amino acid residues interacting with the transition state of the substrate. TRITON is a graphic software package for modelling enzymatic reactions for the analysis of essential interactions between the enzyme and its substrate and for in silico construction of protein mutants. The reactions are modelled using semi-empirical quantum-mechanic methods and the protein mutants are constructed by homology modelling. The users are guided through the calculation and data analysis by wizards.

L-Cysteine Production in Escherichia coli Based on Rational Metabolic Engineering and Modular Strategy.

Science.gov (United States)

Liu, Han; Fang, Guochen; Wu, Hui; Li, Zhimin; Ye, Qin

2018-05-01

L-cysteine is an amino acid with important physiological functions and has a wide range of applications in medicine, food, animal feed, and cosmetics industry. In this study, the L-cysteine synthesis in Escherichia coliEscherichia coli is divided into four modules: the transport module, sulfur module, precursor module, and degradation module. The engineered strain LH03 (overexpression of the feedback-insensitive cysE and the exporter ydeD in JM109) accumulated 45.8 mg L -1 of L-cysteine in 48 hr with yield of 0.4% g/g glucose. Further modifications of strains and culture conditions which based on the rational metabolic engineering and modular strategy improved the L-cysteine biosynthesis significantly. The engineered strain LH06 (with additional overexpression of serA, serC, and serB and double mutant of tnaA and sdaA in LH03) produced 620.9 mg L -1 of L-cysteine with yield of 6.0% g/g glucose, which increased the production by 12 times and the yield by 14 times more than those of LH03 in the original condition. In fed-batch fermentation performed in a 5-L reactor, the concentration of L-cysteine achieved 5.1 g L -1 in 32 hr. This work demonstrates that the combination of rational metabolic engineering and module strategy is a promising approach for increasing the L-cysteine production in E. coli. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Rationally engineered polymeric cisplatin nanoparticles for improved antitumor efficacy

International Nuclear Information System (INIS)

Paraskar, Abhimanyu; Soni, Shivani; Basu, Sudipta; Srivats, Shyam; Roy, Rituparna Sinha; Sengupta, Shiladitya; Amarasiriwardena, Chitra J; Lupoli, Nicola

2011-01-01

The use of cisplatin, a first line chemotherapy for most cancers, is dose-limited due to nephrotoxicity. While this toxicity can be addressed through nanotechnology, previous attempts at engineering cisplatin nanoparticles have been limited by the impact on the potency of cisplatin. Here we report the rational engineering of a novel cisplatin nanoparticle by harnessing a novel polyethylene glycol-functionalized poly-isobutylene-maleic acid (PEG-PIMA) copolymer, which can complex with cis-platinum (II) through a monocarboxylato and a coordinate bond. We show that this complex self-assembles into a nanoparticle, and exhibits an IC 50 = 0.77 ± 0.11 μM comparable to that of free cisplatin (IC 50 = 0.44 ± 0.09 μM). The nanoparticles are internalized into the endolysosomal compartment of cancer cells, and release cisplatin in a pH-dependent manner. Furthermore, the nanoparticles exhibit significantly improved antitumor efficacy in a 4T1 breast cancer model in vivo, with limited nephrotoxicity, which can be explained by preferential biodistribution in the tumor with reduced kidney concentrations. Our results suggest that the PEG-PIMA-cisplatin nanoparticle can emerge as an attractive solution to the challenges in cisplatin chemotherapy.

Investigation of the influence of the content of plant crude protein in the ration on the utilisation of urea in dairy cattle. 5

International Nuclear Information System (INIS)

Motyl, T.; Barej, W.; Kulasek, G.; Bergner, H.; Goersch, R.

1984-01-01

The incorporation of urea- 15 N (given as an intraruminal drench or infusion) into plasma urea and protein of dairy cows fed isoenergetic rations with different levels of plant protein (9, 11, 12, 14, 15, and 17% in dry matter) was investigated. A nonlinear and asymptotic dependence between the plasma concentration of urea and protein level in the ration was stated. The availability of dietary urea- 15 N for plasma urea for 48 hours after administration was lowest in cows fed with low protein rations (9 and 11% of plant protein). On the contrary the highest incorporation of urea- 15 N into plasma protein of these animals was observed. The possible explanation of these results is presented. (author)

Rationalization and Design of the Complementarity Determining Region Sequences in an Antibody-Antigen Recognition Interface

Science.gov (United States)

Chen, Ing-Chien; Lee, Yu-Ching; Chen, Jun-Bo; Tsai, Keng-Chang; Chen, Ching-Tai; Chang, Jeng-Yih; Yang, Ei-Wen; Hsu, Po-Chiang; Jian, Jhih-Wei; Hsu, Hung-Ju; Chang, Hung-Ju; Hsu, Wen-Lian; Huang, Kai-Fa; Ma, Alex Che; Yang, An-Suei

2012-01-01

Protein-protein interactions are critical determinants in biological systems. Engineered proteins binding to specific areas on protein surfaces could lead to therapeutics or diagnostics for treating diseases in humans. But designing epitope-specific protein-protein interactions with computational atomistic interaction free energy remains a difficult challenge. Here we show that, with the antibody-VEGF (vascular endothelial growth factor) interaction as a model system, the experimentally observed amino acid preferences in the antibody-antigen interface can be rationalized with 3-dimensional distributions of interacting atoms derived from the database of protein structures. Machine learning models established on the rationalization can be generalized to design amino acid preferences in antibody-antigen interfaces, for which the experimental validations are tractable with current high throughput synthetic antibody display technologies. Leave-one-out cross validation on the benchmark system yielded the accuracy, precision, recall (sensitivity) and specificity of the overall binary predictions to be 0.69, 0.45, 0.63, and 0.71 respectively, and the overall Matthews correlation coefficient of the 20 amino acid types in the 24 interface CDR positions was 0.312. The structure-based computational antibody design methodology was further tested with other antibodies binding to VEGF. The results indicate that the methodology could provide alternatives to the current antibody technologies based on animal immune systems in engineering therapeutic and diagnostic antibodies against predetermined antigen epitopes. PMID:22457753

Nitrogen balance, microbial protein synthesis and blood metabolites in fattening of male Bali cattle fed ration with different protein levels in smallholder farms

Directory of Open Access Journals (Sweden)

P. K. Tahuk

2018-03-01

Full Text Available Research was aimed to determine nitrogen balance, microbial protein synthesis, and blood metabolites of male Bali cattle fattening fed ration with different protein level in smallholder farms North Central Timor, Province of East Timor Tenggara, Indonesia. The cattle used were 18 heads aged 2 to 2.5 years with initial body weight of 229.86±12.46 kg. The cattle were randomly divided into three treatment groups. The T0 group was given feed the same as traditional fattening cattle practices by farmers,T1 group fed ration containing 12% crude protein (CP and 72% total digestible nutrients (TDN, andT2 group fedration containing 15% CP and 72%TDN. Cattle were fed individually for 90 days and drinkingwater ad libitum. The data were analyzedby analysis of variance.Results of research indicated the nitrogen balance, and blood urea nitrogen between T1 and T2 were relatively similar, but those were higher (P<0.05 than T0 . In contrast, microbial proteins synthesis, and blood glucose at 0, 4, and 6 hours before and after feeding were relatively similar between the groups. Blood glucose of T2 at 2 hours after intake were higher (P <0.05 than T0, but was not different with T1 . It can be concluded, that the fattening maleBali cattle fed ration containing 12% CP and 72% TDNimprovedthe nitrogen balance and blood metabolites, butit was no positive effect on the microbial proteins and N synthesis.

Molecular basis of glyphosate resistance: Different approaches through protein engineering

Science.gov (United States)

Pollegioni, Loredano; Schonbrunn, Ernst; Siehl, Daniel

2011-01-01

Glyphosate (N-phosphonomethyl-glycine) is the most-used herbicide in the world: glyphosate-based formulations exhibit broad-spectrum herbicidal activity with minimal human and environmental toxicity. The extraordinary success of this simple small molecule is mainly due to the high specificity of glyphosate towards the plant enzyme enolpyruvylshikimate-3-phosphate synthase in the shikimate pathway leading to biosynthesis of aromatic amino acids. Starting in 1996, transgenic glyphosate-resistant plants were introduced thus allowing the application of the herbicide to the crop (post-emergence) to remove emerged weeds without crop damage. This review focuses on the evolution of mechanisms of resistance to glyphosate as obtained through natural diversity, the gene shuffling approach to molecular evolution, and a rational, structure-based approach to protein engineering. In addition, we offer rationale for the means by which the modifications made have had their intended effect. PMID:21668647

Rational design of new materials using recombinant structural proteins: Current state and future challenges.

Science.gov (United States)

Sutherland, Tara D; Huson, Mickey G; Rapson, Trevor D

2018-01-01

Sequence-definable polymers are seen as a prerequisite for design of future materials, with many polymer scientists regarding such polymers as the holy grail of polymer science. Recombinant proteins are sequence-defined polymers. Proteins are dictated by DNA templates and therefore the sequence of amino acids in a protein is defined, and molecular biology provides tools that allow redesign of the DNA as required. Despite this advantage, proteins are underrepresented in materials science. In this publication we investigate the advantages and limitations of using proteins as templates for rational design of new materials. Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.

Introduction to current and future protein therapeutics: a protein engineering perspective.

Science.gov (United States)

Carter, Paul J

2011-05-15

Protein therapeutics and its enabling sister discipline, protein engineering, have emerged since the early 1980s. The first protein therapeutics were recombinant versions of natural proteins. Proteins purposefully modified to increase their clinical potential soon followed with enhancements derived from protein or glycoengineering, Fc fusion or conjugation to polyethylene glycol. Antibody-based drugs subsequently arose as the largest and fastest growing class of protein therapeutics. The rationale for developing better protein therapeutics with enhanced efficacy, greater safety, reduced immunogenicity or improved delivery comes from the convergence of clinical, scientific, technological and commercial drivers that have identified unmet needs and provided strategies to address them. Future protein drugs seem likely to be more extensively engineered to improve their performance, e.g., antibodies and Fc fusion proteins with enhanced effector functions or extended half-life. Two old concepts for improving antibodies, namely antibody-drug conjugates and bispecific antibodies, have advanced to the cusp of clinical success. As for newer protein therapeutic platform technologies, several engineered protein scaffolds are in early clinical development and offer differences and some potential advantages over antibodies. Copyright © 2011 Elsevier Inc. All rights reserved.

Improving protein mass and cumulative body weight gain of local chicken fed ration fortified with a combination of Lactobacillus sp. and dahlia inulin

Science.gov (United States)

Wahyuni, H. I.; Suthama, N.; Mangisah, I.; Krismiyanto, L.

2018-01-01

The research aimed to evaluate meat calcium and protein content of local chicken fed diet fortified with a combination of Lactobacillus sp and Dahlia Inulin. One hundred and twenty birds of 4 months old local chicken with average body weight of 1001 g were assigned in a completely randomized design with 4 treatments and 5 replications. The treatments were the farmer formulated ration (FF) and the improved ration (IR), fortified with 1.2% inulin and 1.2 ml Lactobacillus sp. (FFIL and IRIL). Parameters were calcium retention, protein coefficient digestibility, meat calcium and protein mass, and cumulative body weight gain. The results showed that all parameters were significantly affected by dietary treatments. The improved ration resulted in higher calcium retention and protein coefficient digestibility than the farmer formulated ration when fed by both with and without fortification of dahlia inulin and Lactobacillus sp. Meat protein mass of chicken fed by both FR and IR fortified with dahlia inulin and Lactobacillus sp. showed higher value than chicken fed by unfortified FR and IR. Cumulative body weight gain of chicken fed by both FR and IR fortified with dahlia inulin and Lactobacillus sp. also showed higher value than chicken fed by without fortification. In conclusion, both FR and IR fortified with dahlia inulin and Lactobacillus sp. improved meat protein mass and cumulative body weight gain, especially the farmer formulated ration was pronouncedly improved by fortification of Lactobacillus sp. and dahlia inulin.

Recombinant protein scaffolds for tissue engineering

International Nuclear Information System (INIS)

Werkmeister, Jerome A; Ramshaw, John A M

2012-01-01

New biological materials for tissue engineering are now being developed using common genetic engineering capabilities to clone and express a variety of genetic elements that allow cost-effective purification and scaffold fabrication from these recombinant proteins, peptides or from chimeric combinations of these. The field is limitless as long as the gene sequences are known. The utility is dependent on the ease, product yield and adaptability of these protein products to the biomedical field. The development of recombinant proteins as scaffolds, while still an emerging technology with respect to commercial products, is scientifically superior to current use of natural materials or synthetic polymer scaffolds, in terms of designing specific structures with desired degrees of biological complexities and motifs. In the field of tissue engineering, next generation scaffolds will be the key to directing appropriate tissue regeneration. The initial period of biodegradable synthetic scaffolds that provided shape and mechanical integrity, but no biological information, is phasing out. The era of protein scaffolds offers distinct advantages, particularly with the combination of powerful tools of molecular biology. These include, for example, the production of human proteins of uniform quality that are free of infectious agents and the ability to make suitable quantities of proteins that are found in low quantity or are hard to isolate from tissue. For the particular needs of tissue engineering scaffolds, fibrous proteins like collagens, elastin, silks and combinations of these offer further advantages of natural well-defined structural scaffolds as well as endless possibilities of controlling functionality by genetic manipulation. (topical review)

Using Nigella sativa meal as a substitute source for vegetable protein in rations of native growing calves

Directory of Open Access Journals (Sweden)

A. K. Nasser

2011-01-01

Full Text Available The present study was carried out on 15 growing local bull calves of about 150-200 kg, live body weight and 10-12 months old to investigate the effect of substituting soyabean meal as concentrate feed mixture protein by Nigella sativa meal (NSM at 0 , 60 and 100%. Animals were divided into 3 groups of 5 calves each, according to their live body weight for performing feeding trials. All groups of animals were fed iso-nitrogen (15% CP and iso-caloric (2.7 Mcal/kg. ME diets. Experimental rations were offered at 2.5% of live body weight with 1% of wheat straw. At the end of the feeding trial, which lasted for 105 days, blood samples were collected from all calves to estimate the total protein, albumin, globulin, triglyceride and cholesterol. Digestibility trial was carried out on three animals of each group to investigate the nutritional value of rations. Economical study was also carried out on experimental animals. Results indicated that there was an improvement in feed intake by 13 and 14% for groups fed a ration containing NSM compared with the group fed the control one. No significant differences were between groups of calves in total body weight gain and blood parameters. The feed conversion ratio improved by 12% for the group of calves fed control ration as compared with other groups. The same cost of producing 1 kg live body weight gain was found. Substituting soybean meal protein at 60 and 100% by NSM protein significantly improved crude fiber, ether extract, EE, and the values of digestion coefficient. It was concluded that NSM could be substituted instead of soyabean meal for growing local calves with out adverse effects on their performance.

Rational selection and engineering of exogenous principal sigma factor (σ(HrdB)) to increase teicoplanin production in an industrial strain of Actinoplanes teichomyceticus.

Science.gov (United States)

Wang, Haiyong; Yang, Liu; Wu, Kuo; Li, Guanghui

2014-01-16

Transcriptional engineering has presented a strong ability of phenotypic improvement in microorganisms. However, it could not be directly applied to Actinoplanes teichomyceticus L-27 because of the paucity of endogenous transcription factors in the strain. In this study, exogenous transcription factors were rationally selected and transcriptional engineering was carried out to increase the productivity of teicoplanin in L-27. It was illuminated that the σ(HrdB) molecules shared strong similarity of amino acid sequences among some genera of actinomycetes. Combining this advantage with the ability of transcriptional engineering, exogenous sigma factor σ(HrdB) molecules were rationally selected and engineered to improve L-27. hrdB genes from Actinoplanes missouriensis 431, Micromonospora aurantiaca ATCC 27029 and Salinispora arenicola CNS-205 were selected based on molecular evolutionary analysis. Random mutagenesis, DNA shuffling and point mutation were subsequently performed to generate diversified mutants. A recombinant was identified through screening program, yielding 5.3 mg/ml of teicoplanin, over 2-fold compared to that of L-27. More significantly, the engineered strain presented a good performance in 500-l pilot scale fermentation, which meant its valuable potential application in industry. Through rational selection and engineering of exogenous transcriptional factor, we have extended the application of transcriptional engineering. To our knowledge, it is the first time to focus on the related issue. In addition, possessing the advantage of efficient metabolic perturbation in transcription level, this strategy could be useful in analyzing metabolic and physiological mechanisms of strains, especially those with the only information on taxonomy.

Engineering Protein Hydrogels Using SpyCatcher-SpyTag Chemistry.

Science.gov (United States)

Gao, Xiaoye; Fang, Jie; Xue, Bin; Fu, Linglan; Li, Hongbin

2016-09-12

Constructing hydrogels from engineered proteins has attracted significant attention within the material sciences, owing to their myriad potential applications in biomedical engineering. Developing efficient methods to cross-link tailored protein building blocks into hydrogels with desirable mechanical, physical, and functional properties is of paramount importance. By making use of the recently developed SpyCatcher-SpyTag chemistry, we successfully engineered protein hydrogels on the basis of engineered tandem modular elastomeric proteins. Our resultant protein hydrogels are soft but stable, and show excellent biocompatibility. As the first step, we tested the use of these hydrogels as a drug carrier, as well as in encapsulating human lung fibroblast cells. Our results demonstrate the robustness of the SpyCatcher-SpyTag chemistry, even when the SpyTag (or SpyCatcher) is flanked by folded globular domains. These results demonstrate that SpyCatcher-SpyTag chemistry can be used to engineer protein hydrogels from tandem modular elastomeric proteins that can find applications in tissue engineering, in fundamental mechano-biological studies, and as a controlled drug release vehicle.

Investigations of the influence of the content of crude plant protein in the ration on the utilisation of urea in dairy cattle. 2

International Nuclear Information System (INIS)

Voigt, J.; Piatkowski, B.; Krawielitzki, R.; Adam, K.

1984-01-01

The metabolism of 15 N-urea in dairy cows was investigated in dependence on the crude protein content of the rations. With the energy concentration remaining unchanged, the rations contained 10.7(I), 13.7(II) and 17.1(III)% plant crude protein and, after the supplementation of 150 g urea per animal and day, a total of 13.8, 16.7 and 20.2% crude protein in the dry matter. The urea was intraruminally infused during the feeding in the morning and the evening. In the morning feeding of each 1st measuring day it was labelled with 27.5 atom-% 15 N-excess ( 15 N'). The degree of labelling with 15 N' of the N fraction of rumen fluid, contents of the duodenum, feces and milk, precipitable with trichloric acetic acid (TCA) decreased with the rising protein level of the ration. This effect was bigger than could be expected considering the low 15 N' quota in the total N of the ration. In the sequence I..III, 52.7, 32.2 and 30.6% of the 15 N' amount taken in passed the duodenal re-entrant cannula in TCA-precipitable form within 72 hours after 15 N application. 33.3, 21.9 and 22.6% were apparently absorbed in the intestines as TCA-precipitable N within 120 h after the 15 N' application. In the same period 31.7, 43.1 and 72.8% of the 15 N' taken in were excreted in urine. 12.3, 9.6 and 5.8% of the applied 15 N' were found in milk protein. One can conclude that the utilization of urea N decreases with the rising level of crude protein in the ration and that, however, urea N is still biochemically utilized when there is an excess of plant N in the ration. (author)

Minor milk constituents are affected by protein concentration and forage digestibility in the feed ration

DEFF Research Database (Denmark)

Larsen, Torben; Alstrup, Lene; Weisbjerg, Martin Riis

2016-01-01

The present study was conducted in order to investigate if selected minor milk components would be indicative for the nutritional situation of the cow. Forty-eight dairy cows were offered a high digestible ration vs. a lower digestible ration combined with 2 protein levels in a 4 × 4 Latin square...... design. Milk glucose, glucose-6-phosphate, cholesterol, triacylglycerides (TAG), uric acid and β-hydroxybutyrate (BHBA) were measured and correlated mutually and towards other milking parameters (yield, h since last milking, days in milk (DIM), urea, etc). The variation range of the suggested variables...... were broad, a fact that may support their utilisation as predictive parameters. The content of milk metabolites was significantly affected by the change in rations as milk glucose, glucose-6-phosphate, uric acid, and the ratio cholesterol: triacylglycerides increased with higher energy intake while...

Combining random gene fission and rational gene fusion to discover near-infrared fluorescent protein fragments that report on protein-protein interactions.

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Pandey, Naresh; Nobles, Christopher L; Zechiedrich, Lynn; Maresso, Anthony W; Silberg, Jonathan J

2015-05-15

Gene fission can convert monomeric proteins into two-piece catalysts, reporters, and transcription factors for systems and synthetic biology. However, some proteins can be challenging to fragment without disrupting function, such as near-infrared fluorescent protein (IFP). We describe a directed evolution strategy that can overcome this challenge by randomly fragmenting proteins and concomitantly fusing the protein fragments to pairs of proteins or peptides that associate. We used this method to create libraries that express fragmented IFP as fusions to a pair of associating peptides (IAAL-E3 and IAAL-K3) and proteins (CheA and CheY) and screened for fragmented IFP with detectable near-infrared fluorescence. Thirteen novel fragmented IFPs were identified, all of which arose from backbone fission proximal to the interdomain linker. Either the IAAL-E3 and IAAL-K3 peptides or CheA and CheY proteins could assist with IFP fragment complementation, although the IAAL-E3 and IAAL-K3 peptides consistently yielded higher fluorescence. These results demonstrate how random gene fission can be coupled to rational gene fusion to create libraries enriched in fragmented proteins with AND gate logic that is dependent upon a protein-protein interaction, and they suggest that these near-infrared fluorescent protein fragments will be suitable as reporters for pairs of promoters and protein-protein interactions within whole animals.

Protein engineering and the use of molecular modeling and simulation: the case of heterodimeric Fc engineering.

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Spreter Von Kreudenstein, Thomas; Lario, Paula I; Dixit, Surjit B

2014-01-01

Computational and structure guided methods can make significant contributions to the development of solutions for difficult protein engineering problems, including the optimization of next generation of engineered antibodies. In this paper, we describe a contemporary industrial antibody engineering program, based on hypothesis-driven in silico protein optimization method. The foundational concepts and methods of computational protein engineering are discussed, and an example of a computational modeling and structure-guided protein engineering workflow is provided for the design of best-in-class heterodimeric Fc with high purity and favorable biophysical properties. We present the engineering rationale as well as structural and functional characterization data on these engineered designs. Copyright © 2013 Elsevier Inc. All rights reserved.

Exploring the Feasibility of Using Silage-Based Feed with Alternative Sources of Protein in Organic Pig Rations

Directory of Open Access Journals (Sweden)

Ruth C. Clements

2013-12-01

Full Text Available Current regulations for organic pig and poultry production systems permit feed ingredients of non-organic origin at an inclusion rate of up to 5 per cent. This is primarily due to concerns that there is an insufficient supply of organic protein on the European Union market, in terms of quality and quantity, to meet the nutritional requirements of pigs and poultry raised on organic farms. However, 100 per cent organic diets for monogastric livestock will become compulsory in the EU from 1 January 2018, and there is therefore a need to develop sustainable feeding strategies based on organic feeds. This feed trial conducted in the UK explores the feasibility of using a silage-based feeding system for Gloucester Old Spot pigs, and compares the inclusion of soya, beans and peas as protein sources in terms of pig growth performance. No significant difference in the pen mean daily live weight gain was observed during the grower phase (pen mean age of 11-14 weeks between the diet groups. However, during the finisher phase (pen mean age of 15-22 weeks, pigs on the soya and pea rations had significantly faster growth rates than pigs fed the bean ration. It is speculated that the slight shortfall in growth rate observed in the pigs fed the bean ration may be offset by the lower cost of production of beans in the UK. This feasibility trial demonstrates that a 100 per cent organic diet for pigs using alternative, locally-grown sources of protein as part of a forage-based ration can provide a viable alternative to a soya-based diet.

Deep sequencing methods for protein engineering and design.

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Wrenbeck, Emily E; Faber, Matthew S; Whitehead, Timothy A

2017-08-01

The advent of next-generation sequencing (NGS) has revolutionized protein science, and the development of complementary methods enabling NGS-driven protein engineering have followed. In general, these experiments address the functional consequences of thousands of protein variants in a massively parallel manner using genotype-phenotype linked high-throughput functional screens followed by DNA counting via deep sequencing. We highlight the use of information rich datasets to engineer protein molecular recognition. Examples include the creation of multiple dual-affinity Fabs targeting structurally dissimilar epitopes and engineering of a broad germline-targeted anti-HIV-1 immunogen. Additionally, we highlight the generation of enzyme fitness landscapes for conducting fundamental studies of protein behavior and evolution. We conclude with discussion of technological advances. Copyright © 2016 Elsevier Ltd. All rights reserved.

Development of rations for the enhanced survival of salmon

International Nuclear Information System (INIS)

Ewing, R.D.; Lagasse, J.P.

1990-12-01

The nutritional quality of feed plays an important role in determining the health and ''fitness'' of smolts. Commercial fish meal, the major source of protein in salmon rations, may be reduced in quality from poor drying techniques during manufacture. Dietary stress in the hatchery may result. This investigation tests the hypothesis that protein quality of fish rations can influence the survival of smolts and the ultimate return of adults. The test involves a comparison between performances of coho (Oncorhynchus kisutch) and chinook salmon (O. tshawytscha) reared on rations containing very high quality protein derived from vacuum dried meals and those of fish reared on commercial rations, with commercial fish meal as a source of protein. Survival and return of several brood years of test and control fish are used to measure the influence of ration on survival. This report includes recovery data from these marked fish collected 1982 through September 1990

Biocatalysts: application and engineering for industrial purposes.

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Jemli, Sonia; Ayadi-Zouari, Dorra; Hlima, Hajer Ben; Bejar, Samir

2016-01-01

Enzymes are widely applied in various industrial applications and processes, including the food and beverage, animal feed, textile, detergent and medical industries. Enzymes screened from natural origins are often engineered before entering the market place because their native forms do not meet the requirements for industrial application. Protein engineering is concerned with the design and construction of novel enzymes with tailored functional properties, including stability, catalytic activity, reaction product inhibition and substrate specificity. Two broad approaches have been used for enzyme engineering, namely, rational design and directed evolution. The powerful and revolutionary techniques so far developed for protein engineering provide excellent opportunities for the design of industrial enzymes with specific properties and production of high-value products at lower production costs. The present review seeks to highlight the major fields of enzyme application and to provide an updated overview on previous protein engineering studies wherein natural enzymes were modified to meet the operational conditions required for industrial application.

Improving the success and impact of the metabolic engineering design, build, test, learn cycle by addressing proteins of unknown function.

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Jarboe, Laura R

2018-01-04

Rational, predictive metabolic engineering of organisms requires an ability to associate biological activity to the corresponding gene(s). Despite extensive advances in the 20 years since the Escherichia coli genome was published, there are still gaps in our knowledge of protein function. The substantial amount of data that has been published, such as: omics-level characterization in a myriad of conditions; genome-scale libraries; and evolution and genome sequencing, provide means of identifying and prioritizing proteins for characterization. This review describes the scale of this knowledge gap, demonstrates the benefit of addressing the knowledge gap, and demonstrates the availability of interesting candidates for characterization. Copyright © 2017 Elsevier Ltd. All rights reserved.

Rational design and evolutional fine tuning of Saccharomyces cerevisiae for biomass breakdown.

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Hasunuma, Tomohisa; Ishii, Jun; Kondo, Akihiko

2015-12-01

Conferring biomass hydrolysis activity on yeast through genetic engineering has paved the way for the development of groundbreaking processes for producing liquid fuels and commodity chemicals from lignocellulosic biomass. However, the overproduction and misfolding of heterologous and endogenous proteins can trigger cellular stress, increasing the metabolic burden and retarding growth. Improving the efficiency of lignocellulosic breakdown requires engineering of yeast secretory pathway based on system-wide metabolic analysis as well as DNA constructs for enhanced cellulase gene expression with advanced molecular biology tools. Also, yeast is subjected to severe stress due to toxic compounds generated during lignocellulose pretreatment in consolidated saccharification and fermentation processes. The prospect for development of robust yeast strains makes combining evolutionary and rational engineering strategies. Copyright © 2015 Elsevier Ltd. All rights reserved.

Protein engineering of CYP105s for their industrial uses.

Science.gov (United States)

Yasuda, Kaori; Sugimoto, Hiroshi; Hayashi, Keiko; Takita, Teisuke; Yasukawa, Kiyoshi; Ohta, Miho; Kamakura, Masaki; Ikushiro, Shinichi; Shiro, Yoshitsugu; Sakaki, Toshiyuki

2018-01-01

Cytochrome P450 enzymes belonging to the CYP105 family are predominantly found in bacteria belonging to the phylum Actinobacteria and the order Actinomycetales. In this review, we focused on the protein engineering of P450s belonging to the CYP105 family for industrial use. Two Arg substitutions to Ala of CYP105A1 enhanced its vitamin D 3 25- and 1α-hydroxylation activities by 400 and 100-fold, respectively. The coupling efficiency between product formation and NADPH oxidation was largely improved by the R84A mutation. The quintuple mutant Q87W/T115A/H132L/R194W/G294D of CYP105AB3 showed a 20-fold higher activity than the wild-type enzyme. Amino acids at positions 87 and 191 were located at the substrate entrance channel, and that at position 294 was located close to the heme group. Semi-rational engineering of CYP105A3 selected the best performing mutant, T85F/T119S/V194N/N363Y, for producing pravastatin. The T119S and N363Y mutations synergistically had remarkable effects on the interaction between CYP105A3 and putidaredoxin. Although wild-type CYP105AS1 hydroxylated compactin to 6-epi-pravastatin, the quintuple mutant I95T/Q127R/A180V/L236I/A265N converted almost all compactin to pravastatin. Five amino acid substitutions by two rounds of mutagenesis almost completely changed the stereo-selectivity of CYP105AS1. These results strongly suggest that the protein engineering of CYP105 enzymes greatly increase their industrial utility. This article is part of a Special Issue entitled: Cytochrome P450 biodiversity and biotechnology, edited by Erika Plettner, Gianfranco Gilardi, Luet Wong, Vlada Urlacher, Jared Goldstone. Copyright © 2017 Elsevier B.V. All rights reserved.

Towards a rational approach for heavy-atom derivative screening in protein crystallography

International Nuclear Information System (INIS)

Agniswamy, Johnson; Joyce, M. Gordon; Hammer, Carl H.; Sun, Peter D.

2008-01-01

Heavy-atom derivatization is routinely used in protein structure determination and is thus of critical importance in structural biology. In order to replace the current trial-and-error heavy-atom derivative screening with a knowledge-based rational derivative-selection method, the reactivity of more than 40 heavy-atom compounds over a wide range of buffer and pH values was systematically examined using peptides which contained a single reactive amino-acid residue. Heavy-atom derivatization is routinely used in protein structure determination and is thus of critical importance in structural biology. In order to replace the current trial-and-error heavy-atom derivative screening with a knowledge-based rational derivative-selection method, the reactivity of more than 40 heavy-atom compounds over a wide range of buffer and pH values was systematically examined using peptides which contained a single reactive amino-acid residue. Met-, Cys- and His-containing peptides were derivatized against Hg, Au and Pt compounds, while Tyr-, Glu-, Asp-, Asn- and Gln-containing peptides were assessed against Pb compounds. A total of 1668 reactive conditions were examined using mass spectrometry and were compiled into heavy-atom reactivity tables. The results showed that heavy-atom derivatization reactions are highly linked to buffer and pH, with the most accommodating buffer being MES at pH 6. A group of 21 compounds were identified as most successful irrespective of ligand or buffer/pH conditions. To assess the applicability of the peptide heavy-atom reactivity to proteins, lysozyme crystals were derivatized with a list of peptide-reactive compounds that included both known and new compounds for lysozyme derivatization. The results showed highly consistent heavy-atom reactivities between the peptides and lysozyme

Protein engineering for biofuel production: Recent development

Directory of Open Access Journals (Sweden)

Nisha Singh

2016-09-01

Full Text Available The unstable and unsure handiness of crude oil sources moreover the rising price of fuels have shifted international efforts to utilize renewable resources for the assembly of greener energy and a replacement which might additionally meet the high energy demand of the globe. Biofuels represent a sustainable, renewable, and also the solely predictable energy supply to fossil fuels. During the green production of Biofuels, several in vivo processes place confidence in the conversion of biomass to sugars by engineered enzymes, and the subsequent conversion of sugars to chemicals via designed proteins in microbial production hosts. Enzymes are indispensable within the effort to provide fuels in an ecologically friendly manner. They have the potential to catalyze reactions with high specificity and potency while not using dangerous chemicals. Nature provides an in depth assortment of enzymes, however usually these should be altered to perform desired functions in needed conditions. Presently available enzymes like cellulose are subject to tight induction and regulation systems and additionally suffer inhibition from numerous end products. Therefore, more impregnable and economical catalyst preparations ought to be developed for the enzymatic method to be more economical. Approaches like protein engineering, reconstitution of protein mixtures and bio prospecting for superior enzymes are gaining importance. Advances in enzyme engineering allow the planning and/or directed evolution of enzymes specifically tailored for such industrial applications. Recent years have seen the production of improved enzymes to help with the conversion of biomass into fuels. The assembly of the many of those fuels is feasible due to advances in protein engineering. This review discusses the distinctive challenges that protein engineering faces in the method of changing lignocellulose to biofuels and the way they're addressed by recent advances in this field.

An Engineered Split Intein for Photoactivated Protein Trans-Splicing.

Directory of Open Access Journals (Sweden)

Stanley Wong

Full Text Available Protein splicing is mediated by inteins that auto-catalytically join two separated protein fragments with a peptide bond. Here we engineered a genetically encoded synthetic photoactivatable intein (named LOVInC, by using the light-sensitive LOV2 domain from Avena sativa as a switch to modulate the splicing activity of the split DnaE intein from Nostoc punctiforme. Periodic blue light illumination of LOVInC induced protein splicing activity in mammalian cells. To demonstrate the broad applicability of LOVInC, synthetic protein systems were engineered for the light-induced reassembly of several target proteins such as fluorescent protein markers, a dominant positive mutant of RhoA, caspase-7, and the genetically encoded Ca2+ indicator GCaMP2. Spatial precision of LOVInC was demonstrated by targeting activity to specific mammalian cells. Thus, LOVInC can serve as a general platform for engineering light-based control for modulating the activity of many different proteins.

Rational design of an EGF-IL18 fusion protein: Implication for developing tumor therapeutics

International Nuclear Information System (INIS)

Lu Jianxin; Peng Ying; Meng Zhefeng; Jin Liqin; Lu Yongsui; Guan Minxin

2005-01-01

Interleukin-18 (IL-18) is a proinflammatory cytokine. This protein has a role in regulating immune responses and exhibits significant anti-tumor activities. Epidermal growth factor (EGF) is an important growth factor that plays a central role in the regulation of cell cycle and differentiation. It was proposed that a targeted delivery of IL-18 by generation of IL-18-EGF fusion protein might decrease adverse effects and result in enhancing cytotoxic and antitumor activities. In the present study, a fusion protein, consisting of EGFR binding domain fused to human IL-18 mature peptide via a linker peptide of (Gly 4 Ser) 3, was constructed and expressed in the insect cell line Sf9 using Bac-to-Bac baculovirus expression system. We showed that the purified recombinant fusion protein induced similar levels of IFN-γ to that of native IL-18 protein in human PBMC in the presence of ConA. Furthermore, EGF receptor competitive test in human epithelial cancer A431 cell line showed that EGF-IL18 fusion protein can specifically bind with EGFR by competing with native EGF protein. These suggest that this rationally designed protein can be further developed as novel tumor therapeutics

Overcoming barriers to membrane protein structure determination.

Science.gov (United States)

Bill, Roslyn M; Henderson, Peter J F; Iwata, So; Kunji, Edmund R S; Michel, Hartmut; Neutze, Richard; Newstead, Simon; Poolman, Bert; Tate, Christopher G; Vogel, Horst

2011-04-01

After decades of slow progress, the pace of research on membrane protein structures is beginning to quicken thanks to various improvements in technology, including protein engineering and microfocus X-ray diffraction. Here we review these developments and, where possible, highlight generic new approaches to solving membrane protein structures based on recent technological advances. Rational approaches to overcoming the bottlenecks in the field are urgently required as membrane proteins, which typically comprise ~30% of the proteomes of organisms, are dramatically under-represented in the structural database of the Protein Data Bank.

Virus-Like Particle Engineering: From Rational Design to Versatile Applications.

Science.gov (United States)

Ding, Xuanwei; Liu, Dong; Booth, George; Gao, Wei; Lu, Yuan

2018-05-01

As mimicking natural virus structures, virus-like particles (VLPs) have evolved to become a widely accepted technology used for humans which are safe, highly efficacious, and profitable. Several remarkable advantages have been achieved to revolutionize the molecule delivery for diverse applications in nanotechnology, biotechnology, and medicine. Here, the rational structure design, manufacturing process, functionalization strategy, and emerging applications of VLPs is reviewed. The situation and challenges in the VLP engineering, the key development orientation, and future applications have been discussed. To develop a good VLP design concept, the virus/VLP-host interactions need to be examined and the screening methods of the VLP stabilization factors need to be established. The functionalization toolbox can be expanded to fabricate smart, robust, and multifunctional VLPs. Novel robust VLP manufacturing platforms are required to deliver vaccines in resource-poor regions with a significant reduction in the production time and cost. The future applications of VLPs are always driven by the development of emerging technologies and new requirements of modern life. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Engineering proteins for environmental applications

NARCIS (Netherlands)

Janssen, Dick B.; Schanstra, Joost P.

1994-01-01

Recently, significant new insight has been obtained into the structure and catalytic mechanism of enzymes that convert environmental pollutants. Recent advances in protein engineering make it possible to use this information for improving the catalytic performance of such enzymes to achieve

Rationally reduced libraries for combinatorial pathway optimization minimizing experimental effort.

Science.gov (United States)

Jeschek, Markus; Gerngross, Daniel; Panke, Sven

2016-03-31

Rational flux design in metabolic engineering approaches remains difficult since important pathway information is frequently not available. Therefore empirical methods are applied that randomly change absolute and relative pathway enzyme levels and subsequently screen for variants with improved performance. However, screening is often limited on the analytical side, generating a strong incentive to construct small but smart libraries. Here we introduce RedLibs (Reduced Libraries), an algorithm that allows for the rational design of smart combinatorial libraries for pathway optimization thereby minimizing the use of experimental resources. We demonstrate the utility of RedLibs for the design of ribosome-binding site libraries by in silico and in vivo screening with fluorescent proteins and perform a simple two-step optimization of the product selectivity in the branched multistep pathway for violacein biosynthesis, indicating a general applicability for the algorithm and the proposed heuristics. We expect that RedLibs will substantially simplify the refactoring of synthetic metabolic pathways.

Pharmacokinetic and pharmacodynamic considerations for the next generation protein therapeutics.

Science.gov (United States)

Shah, Dhaval K

2015-10-01

Increasingly sophisticated protein engineering efforts have been undertaken lately to generate protein therapeutics with desired properties. This has resulted in the discovery of the next generation of protein therapeutics, which include: engineered antibodies, immunoconjugates, bi/multi-specific proteins, antibody mimetic novel scaffolds, and engineered ligands/receptors. These novel protein therapeutics possess unique physicochemical properties and act via a unique mechanism-of-action, which collectively makes their pharmacokinetics (PK) and pharmacodynamics (PD) different than other established biological molecules. Consequently, in order to support the discovery and development of these next generation molecules, it becomes important to understand the determinants controlling their PK/PD. This review discusses the determinants that a PK/PD scientist should consider during the design and development of next generation protein therapeutics. In addition, the role of systems PK/PD models in enabling rational development of the next generation protein therapeutics is emphasized.

Simultaneous inhibition of aberrant cancer kinome using rationally designed polymer-protein core-shell nanomedicine.

Science.gov (United States)

Chandran, Parwathy; Gupta, Neha; Retnakumari, Archana Payickattu; Malarvizhi, Giridharan Loghanathan; Keechilat, Pavithran; Nair, Shantikumar; Koyakutty, Manzoor

2013-11-01

Simultaneous inhibition of deregulated cancer kinome using rationally designed nanomedicine is an advanced therapeutic approach. Herein, we have developed a polymer-protein core-shell nanomedicine to inhibit critically aberrant pro-survival kinases (mTOR, MAPK and STAT5) in primitive (CD34(+)/CD38(-)) Acute Myeloid Leukemia (AML) cells. The nanomedicine consists of poly-lactide-co-glycolide core (~250 nm) loaded with mTOR inhibitor, everolimus, and albumin shell (~25 nm thick) loaded with MAPK/STAT5 inhibitor, sorafenib and the whole construct was surface conjugated with monoclonal antibody against CD33 receptor overexpressed in AML. Electron microscopy confirmed formation of core-shell nanostructure (~290 nm) and flow cytometry and confocal studies showed enhanced cellular uptake of targeted nanomedicine. Simultaneous inhibition of critical kinases causing synergistic lethality against leukemic cells, without affecting healthy blood cells, was demonstrated using immunoblotting, cytotoxicity and apoptosis assays. This cell receptor plus multi-kinase targeted core-shell nanomedicine was found better specific and tolerable compared to current clinical regime of cytarabine and daunorubicin. These authors demonstrate simultaneous inhibition of critical kinases causing synergistic lethality against leukemic cells, without affecting healthy blood cells by using rationally designed polymer-protein core-shell nanomedicine, provoding an advanced method to eliminate cancer cells, with the hope of future therapeutic use. Copyright © 2013 Elsevier Inc. All rights reserved.

Rational redesign of neutral endopeptidase binding to merlin and moesin proteins

Science.gov (United States)

Niv, Masha Y; Iida, Katsuyuki; Zheng, Rong; Horiguchi, Akio; Shen, Ruoqian; Nanus, David M

2009-01-01

Neutral endopeptidase (NEP) is a 90- to 110-kDa cell-surface peptidase that is normally expressed by numerous tissues but whose expression is lost or reduced in a variety of malignancies. The anti-tumorigenic function of NEP is mediated not only by its catalytic activity but also through direct protein–protein interactions of its cytosolic region with several binding partners, including Lyn kinase, PTEN, and ezrin/radixin/moesin (ERM) proteins. We have previously shown that mutation of the K19K20K21 basic cluster in NEPs' cytosolic region to residues QNI disrupts binding to the ERM proteins. Here we show that the ERM-related protein merlin (NF2) does not bind NEP or its cytosolic region. Using experimental data, threading, and sequence analysis, we predicted the involvement of moesin residues E159Q160 in binding to the NEP cytosolic domain. Mutation of these residues to NL (to mimic the corresponding N159L160 residues in the nonbinder merlin) disrupted moesin binding to NEP. Mutation of residues N159L160Y161K162M163 in merlin to the corresponding moesin residues resulted in NEP binding to merlin. This engineered NEP peptide–merlin interaction was diminished by the QNI mutation in NEP, supporting the role of the NEP basic cluster in binding. We thus identified the region of interaction between NEP and moesin, and engineered merlin into a NEP-binding protein. These data form the basis for further exploration of the details of NEP-ERM binding and function. PMID:19388049

Protein consensus-based surface engineering (ProCoS): a computer-assisted method for directed protein evolution.

Science.gov (United States)

Shivange, Amol V; Hoeffken, Hans Wolfgang; Haefner, Stefan; Schwaneberg, Ulrich

2016-12-01

Protein consensus-based surface engineering (ProCoS) is a simple and efficient method for directed protein evolution combining computational analysis and molecular biology tools to engineer protein surfaces. ProCoS is based on the hypothesis that conserved residues originated from a common ancestor and that these residues are crucial for the function of a protein, whereas highly variable regions (situated on the surface of a protein) can be targeted for surface engineering to maximize performance. ProCoS comprises four main steps: ( i ) identification of conserved and highly variable regions; ( ii ) protein sequence design by substituting residues in the highly variable regions, and gene synthesis; ( iii ) in vitro DNA recombination of synthetic genes; and ( iv ) screening for active variants. ProCoS is a simple method for surface mutagenesis in which multiple sequence alignment is used for selection of surface residues based on a structural model. To demonstrate the technique's utility for directed evolution, the surface of a phytase enzyme from Yersinia mollaretii (Ymphytase) was subjected to ProCoS. Screening just 1050 clones from ProCoS engineering-guided mutant libraries yielded an enzyme with 34 amino acid substitutions. The surface-engineered Ymphytase exhibited 3.8-fold higher pH stability (at pH 2.8 for 3 h) and retained 40% of the enzyme's specific activity (400 U/mg) compared with the wild-type Ymphytase. The pH stability might be attributed to a significantly increased (20 percentage points; from 9% to 29%) number of negatively charged amino acids on the surface of the engineered phytase.

Rational design of highly potent HIV-1 fusion inhibitory proteins: Implication for developing antiviral therapeutics

International Nuclear Information System (INIS)

Ni Ling; Gao, George F.; Tien Po

2005-01-01

Recombinant protein containing one heptad-repeat 1 (HR1) segment and one HR2 segment of the HIV-1 gp41 (HR1-HR2) has been shown to fold into thermally stable six-helix bundle, representing the fusogenic core of gp41. In this study, we have used the fusogenic core as a scaffold to design HIV-1 fusion inhibitory proteins by linking another HR1 to the C terminus of HR1-HR2 (HR121) or additional HR2 to the N terminus of HR1-HR2 (HR212). Both recombinant proteins could be abundantly and solubly expressed and easily purified, exhibiting high stability and potent inhibitory activity on HIV-1 fusion with IC 50 values of 16.2 ± 2.8 and 2.8 ± 0.63 nM, respectively. These suggest that these rationally designed proteins can be further developed as novel anti-HIV-1 therapeutics

Combined protein construct and synthetic gene engineering for heterologous protein expression and crystallization using Gene Composer

Directory of Open Access Journals (Sweden)

Walchli John

2009-04-01

Full Text Available Abstract Background With the goal of improving yield and success rates of heterologous protein production for structural studies we have developed the database and algorithm software package Gene Composer. This freely available electronic tool facilitates the information-rich design of protein constructs and their engineered synthetic gene sequences, as detailed in the accompanying manuscript. Results In this report, we compare heterologous protein expression levels from native sequences to that of codon engineered synthetic gene constructs designed by Gene Composer. A test set of proteins including a human kinase (P38α, viral polymerase (HCV NS5B, and bacterial structural protein (FtsZ were expressed in both E. coli and a cell-free wheat germ translation system. We also compare the protein expression levels in E. coli for a set of 11 different proteins with greatly varied G:C content and codon bias. Conclusion The results consistently demonstrate that protein yields from codon engineered Gene Composer designs are as good as or better than those achieved from the synonymous native genes. Moreover, structure guided N- and C-terminal deletion constructs designed with the aid of Gene Composer can lead to greater success in gene to structure work as exemplified by the X-ray crystallographic structure determination of FtsZ from Bacillus subtilis. These results validate the Gene Composer algorithms, and suggest that using a combination of synthetic gene and protein construct engineering tools can improve the economics of gene to structure research.

PIA: An Intuitive Protein Inference Engine with a Web-Based User Interface.

Science.gov (United States)

Uszkoreit, Julian; Maerkens, Alexandra; Perez-Riverol, Yasset; Meyer, Helmut E; Marcus, Katrin; Stephan, Christian; Kohlbacher, Oliver; Eisenacher, Martin

2015-07-02

Protein inference connects the peptide spectrum matches (PSMs) obtained from database search engines back to proteins, which are typically at the heart of most proteomics studies. Different search engines yield different PSMs and thus different protein lists. Analysis of results from one or multiple search engines is often hampered by different data exchange formats and lack of convenient and intuitive user interfaces. We present PIA, a flexible software suite for combining PSMs from different search engine runs and turning these into consistent results. PIA can be integrated into proteomics data analysis workflows in several ways. A user-friendly graphical user interface can be run either locally or (e.g., for larger core facilities) from a central server. For automated data processing, stand-alone tools are available. PIA implements several established protein inference algorithms and can combine results from different search engines seamlessly. On several benchmark data sets, we show that PIA can identify a larger number of proteins at the same protein FDR when compared to that using inference based on a single search engine. PIA supports the majority of established search engines and data in the mzIdentML standard format. It is implemented in Java and freely available at https://github.com/mpc-bioinformatics/pia.

Rosetta comparative modeling for library design: Engineering alternative inducer specificity in a transcription factor.

Science.gov (United States)

Jha, Ramesh K; Chakraborti, Subhendu; Kern, Theresa L; Fox, David T; Strauss, Charlie E M

2015-07-01

Structure-based rational mutagenesis for engineering protein functionality has been limited by the scarcity and difficulty of obtaining crystal structures of desired proteins. On the other hand, when high-throughput selection is possible, directed evolution-based approaches for gaining protein functionalities have been random and fortuitous with limited rationalization. We combine comparative modeling of dimer structures, ab initio loop reconstruction, and ligand docking to select positions for mutagenesis to create a library focused on the ligand-contacting residues. The rationally reduced library requirement enabled conservative control of the substitutions by oligonucleotide synthesis and bounding its size within practical transformation efficiencies (∼ 10(7) variants). This rational approach was successfully applied on an inducer-binding domain of an Acinetobacter transcription factor (TF), pobR, which shows high specificity for natural effector molecule, 4-hydroxy benzoate (4HB), but no native response to 3,4-dihydroxy benzoate (34DHB). Selection for mutants with high transcriptional induction by 34DHB was carried out at the single-cell level under flow cytometry (via green fluorescent protein expression under the control of pobR promoter). Critically, this selection protocol allows both selection for induction and rejection of constitutively active mutants. In addition to gain-of-function for 34DHB induction, the selected mutants also showed enhanced sensitivity and response for 4HB (native inducer) while no sensitivity was observed for a non-targeted but chemically similar molecule, 2-hydroxy benzoate (2HB). This is unique application of the Rosetta modeling protocols for library design to engineer a TF. Our approach extends applicability of the Rosetta redesign protocol into regimes without a priori precision structural information. © 2015 Wiley Periodicals, Inc.

Fn3 proteins engineered to recognize tumor biomarker mesothelin internalize upon binding.

Directory of Open Access Journals (Sweden)

Allison R Sirois

Full Text Available Mesothelin is a cell surface protein that is overexpressed in numerous cancers, including breast, ovarian, lung, liver, and pancreatic tumors. Aberrant expression of mesothelin has been shown to promote tumor progression and metastasis through interaction with established tumor biomarker CA125. Therefore, molecules that specifically bind to mesothelin have potential therapeutic and diagnostic applications. However, no mesothelin-targeting molecules are currently approved for routine clinical use. While antibodies that target mesothelin are in development, some clinical applications may require a targeting molecule with an alternative protein fold. For example, non-antibody proteins are more suitable for molecular imaging and may facilitate diverse chemical conjugation strategies to create drug delivery complexes. In this work, we engineered variants of the fibronectin type III domain (Fn3 non-antibody protein scaffold to bind to mesothelin with high affinity, using directed evolution and yeast surface display. Lead engineered Fn3 variants were solubly produced and purified from bacterial culture at high yield. Upon specific binding to mesothelin on human cancer cell lines, the engineered Fn3 proteins internalized and co-localized to early endosomes. To our knowledge, this is the first report of non-antibody proteins engineered to bind mesothelin. The results validate that non-antibody proteins can be engineered to bind to tumor biomarker mesothelin, and encourage the continued development of engineered variants for applications such as targeted diagnostics and therapeutics.

Integrating protein engineering with process design for biocatalysis

DEFF Research Database (Denmark)

Woodley, John M.

2017-01-01

Biocatalysis uses enzymes for chemical synthesis and production, offering selective, safe and sustainable catalysis. While today the majority of applications are in the pharmaceutical sector, new opportunities are arising every day in other industry sectors, where production costs become a more...... important driver. In the early applications of the technology, it was necessary to design processes to match the properties of the biocatalyst. With the advent of protein engineering, organic chemists started to develop and improve enzymes to suit their needs. Likewise in industry, although not widespread......, a new paradigm was already implemented several years ago to engineer enzymes to suit process needs. Today, a new era is entered, where the effectiveness with which such integrated protein and process engineering is achieved becomes critical to implementation. In this paper, the development of a tool...

Rational engineering of p-hydroxybenzoate hydroxylase to enable efficient gallic acid synthesis via a novel artificial biosynthetic pathway.

Science.gov (United States)

Chen, Zhenya; Shen, Xiaolin; Wang, Jian; Wang, Jia; Yuan, Qipeng; Yan, Yajun

2017-11-01

Gallic acid (GA) is a naturally occurring phytochemical that has strong antioxidant and antibacterial activities. It is also used as a potential platform chemical for the synthesis of diverse high-value compounds. Hydrolytic degradation of tannins by acids, bases or microorganisms serves as a major way for GA production, which however, might cause environmental pollution and low yield and efficiency. Here, we report a novel approach for efficient microbial production of GA. First, structure-based rational engineering of PobA, a p-hydroxybenzoate hydroxylase from Pseudomonas aeruginosa, generated a new mutant, Y385F/T294A PobA, which displayed much higher activity toward 3,4-dihydroxybenzoic acid (3,4-DHBA) than the wild-type and any other reported mutants. Remarkably, expression of this mutant in Escherichia coli enabled generation of 1149.59 mg/L GA from 1000 mg/L 4-hydroxybenzoic acid (4-HBA), representing a 93% molar conversion ratio. Based on that, we designed and reconstituted a novel artificial biosynthetic pathway of GA and achieved 440.53 mg/L GA production from simple carbon sources in E. coli. Further enhancement of precursor supply through reinforcing shikimate pathway was able to improve GA de novo production to 1266.39 mg/L in shake flasks. Overall, this study not only led to the development of a highly active PobA variant for hydroxylating 3,4-DHBA into GA via structure-based protein engineering approach, but also demonstrated a promising pathway for bio-based manufacturing of GA and its derived compounds. Biotechnol. Bioeng. 2017;114: 2571-2580. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

The development and application of engineered proteins for bioremediation

Energy Technology Data Exchange (ETDEWEB)

Trewhella, J. [ed.

1995-09-26

Clean up of the toxic legacy of the Cold War is projected to be the most expensive domestic project the nation has yet undertaken. Remediation of the Department of Energy and Department of Defense toxic waste sites alone are projected to cost {approximately}$1 trillion over a 20-30 year period. New, cost effective technologies are needed to attack this enormous problem. Los Alamos has put together a cross-divisional team of scientist to develop science based bioremediation technology to work toward this goal. In the team we have expertise in: (1) molecular, ecosystem and transport modeling; (2) genetic and protein engineering; (3) microbiology and microbial ecology; (4) structural biology; and (5) bioinorganic chemistry. This document summarizes talks at a workshop of different aspects of bioremediation technology including the following: Introducing novel function into a Heme enzyme: engineering by excavation; cytochrome P-450: ideal systems for bioremediation?; selection and development of bacterial strains for in situ remediation of cholorinated solvents; genetic analysis and preparation of toluene ortho-monooxygenase for field application in remediation of trichloroethylene; microbial ecology and diversity important to bioremediation; engineering haloalkane dehalogenase for bioremediation; enzymes for oxidative biodegradation; indigenous bacteria as hosts for engineered proteins; performance of indigenous bacterial, hosting engineered proteins in microbial communities.

Modulating Cytotoxic Effector Functions by Fc Engineering to Improve Cancer Therapy.

Science.gov (United States)

Kellner, Christian; Otte, Anna; Cappuzzello, Elisa; Klausz, Katja; Peipp, Matthias

2017-09-01

In the last two decades, monoclonal antibodies have revolutionized the therapy of cancer patients. Although antibody therapy has continuously been improved, still a significant number of patients do not benefit from antibody therapy. Therefore, rational optimization of the antibody molecule by Fc engineering represents a major area of translational research to further improve this potent therapeutic option. Monoclonal antibodies are able to trigger a variety of effector mechanisms. Especially Fc-mediated effector functions such as antibody-dependent cell-mediated cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), and complement- dependent cytotoxicity (CDC) are considered important in antibody therapy of cancer. Novel mechanistic insights into the action of monoclonal antibodies allowed the development of various Fc engineering approaches to modulate antibodies' effector functions. Strategies in modifying the Fc glycosylation profile (Fc glyco-engineering) or approaches in engineering the protein backbone (Fc protein engineering) have been intensively evaluated. In the current review, Fc engineering strategies resulting in improved ADCC, ADCP and CDC activity are summarized and discussed.

Association of total-mixed-ration chemical composition with milk, fat, and protein yield lactation curves at the individual level

NARCIS (Netherlands)

Caccamo, M.; Veerkamp, R.F.; Licitra, G.; Petriglieri, R.; Terra, La F.; Pozzebon, A.; Ferguson, J.D.

2012-01-01

The objective of this study was to examine the effect of the chemical composition of a total mixed ration (TMR) tested quarterly from March 2006 through December 2008 for milk, fat, and protein yield curves for 27 herds in Ragusa, Sicily. Before this study, standard yield curves were generated on

Protein engineering in designing tailored enzymes and microorganisms for biofuels production

Science.gov (United States)

Wen, Fei; Nair, Nikhil U; Zhao, Huimin

2009-01-01

Summary Lignocellulosic biofuels represent a sustainable, renewable, and the only foreseeable alternative energy source to transportation fossil fuels. However, the recalcitrant nature of lignocellulose poses technical hurdles to an economically viable biorefinery. Low enzymatic hydrolysis efficiency and low productivity, yield, and titer of biofuels are among the top cost contributors. Protein engineering has been used to improve the performances of lignocellulose-degrading enzymes, as well as proteins involved in biofuel synthesis pathways. Unlike its great success seen in other industrial applications, protein engineering has achieved only modest results in improving the lignocellulose-to-biofuels efficiency. This review will discuss the unique challenges that protein engineering faces in the process of converting lignocellulose to biofuels and how they are addressed by recent advances in this field. PMID:19660930

An evolution-based strategy for engineering allosteric regulation

Science.gov (United States)

Pincus, David; Resnekov, Orna; Reynolds, Kimberly A.

2017-04-01

Allosteric regulation provides a way to control protein activity at the time scale of milliseconds to seconds inside the cell. An ability to engineer synthetic allosteric systems would be of practical utility for the development of novel biosensors, creation of synthetic cell signaling pathways, and design of small molecule pharmaceuticals with regulatory impact. To this end, we outline a general approach—termed rational engineering of allostery at conserved hotspots (REACH)—to introduce novel regulation into a protein of interest by exploiting latent allostery that has been hard-wired by evolution into its structure. REACH entails the use of statistical coupling analysis (SCA) to identify ‘allosteric hotspots’ on protein surfaces, the development and implementation of experimental assays to test hotspots for functionality, and a toolkit of allosteric modulators to impinge on endogenous cellular circuitry. REACH can be broadly applied to rewire cellular processes to respond to novel inputs.

Use of a protein engineering strategy to overcome limitations in the production of "Difficult to Express" recombinant proteins.

Science.gov (United States)

Hussain, Hirra; Fisher, David I; Abbott, W Mark; Roth, Robert G; Dickson, Alan J

2017-10-01

Certain recombinant proteins are deemed "difficult to express" in mammalian expression systems requiring significant cell and/or process engineering to abrogate expression bottlenecks. With increasing demand for the production of recombinant proteins in mammalian cells, low protein yields can have significant consequences for industrial processes. To investigate the molecular mechanisms that restrict expression of recombinant proteins, naturally secreted model proteins were analyzed from the tissue inhibitors of metalloproteinase (TIMP) protein family. In particular, TIMP-2 and TIMP-3 were subjected to detailed study. TIMP proteins share significant sequence homology (∼50% identity and ∼70% similarity in amino acid sequence). However, they show marked differences in secretion in mammalian expression systems despite this extensive sequence homology. Using these two proteins as models, this study characterized the molecular mechanisms responsible for poor recombinant protein production. Our results reveal that both TIMP-2 and TIMP-3 are detectable at mRNA and protein level within the cell but only TIMP-2 is secreted effectively into the extracellular medium. Analysis of protein localization and the nature of intracellular protein suggest TIMP-3 is severely limited in its post-translational processing. To overcome this challenge, modification of the TIMP-3 sequence to include a furin protease-cleavable pro-sequence resulted in secretion of the modified TIMP-3 protein, however, incomplete processing was observed. Based on the TIMP-3 data, the protein engineering approach was optimized and successfully applied in combination with cell engineering, the overexpression of furin, to another member of the TIMP protein family (the poorly expressed TIMP-4). Use of the described protein engineering strategy resulted in successful secretion of poorly (TIMP-4) and non-secreted (TIMP-3) targets, and presents a novel strategy to enhance the production of "difficult" recombinant

A tailored biocatalyst achieved by the rational anchoring of imidazole groups on a natural polymer: furnishing a potential artificial nuclease by sustainable materials engineering.

Science.gov (United States)

Ferreira, José G L; Grein-Iankovski, Aline; Oliveira, Marco A S; Simas-Tosin, Fernanda F; Riegel-Vidotti, Izabel C; Orth, Elisa S

2015-04-11

Foreseeing the development of artificial enzymes by sustainable materials engineering, we rationally anchored reactive imidazole groups on gum arabic, a natural biocompatible polymer. The tailored biocatalyst GAIMZ demonstrated catalytic activity (>10(5)-fold) in dephosphorylation reactions with recyclable features and was effective in cleaving plasmid DNA, comprising a potential artificial nuclease.

A Rational Method for Ranking Engineering Programs.

Science.gov (United States)

Glower, Donald D.

1980-01-01

Compares two methods for ranking academic programs, the opinion poll v examination of career successes of the program's alumni. For the latter, "Who's Who in Engineering" and levels of research funding provided data. Tables display resulting data and compare rankings by the two methods for chemical engineering and civil engineering. (CS)

Protein design in systems metabolic engineering for industrial strain development.

Science.gov (United States)

Chen, Zhen; Zeng, An-Ping

2013-05-01

Accelerating the process of industrial bacterial host strain development, aimed at increasing productivity, generating new bio-products or utilizing alternative feedstocks, requires the integration of complementary approaches to manipulate cellular metabolism and regulatory networks. Systems metabolic engineering extends the concept of classical metabolic engineering to the systems level by incorporating the techniques used in systems biology and synthetic biology, and offers a framework for the development of the next generation of industrial strains. As one of the most useful tools of systems metabolic engineering, protein design allows us to design and optimize cellular metabolism at a molecular level. Here, we review the current strategies of protein design for engineering cellular synthetic pathways, metabolic control systems and signaling pathways, and highlight the challenges of this subfield within the context of systems metabolic engineering. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

A real-time all-atom structural search engine for proteins.

Science.gov (United States)

Gonzalez, Gabriel; Hannigan, Brett; DeGrado, William F

2014-07-01

Protein designers use a wide variety of software tools for de novo design, yet their repertoire still lacks a fast and interactive all-atom search engine. To solve this, we have built the Suns program: a real-time, atomic search engine integrated into the PyMOL molecular visualization system. Users build atomic-level structural search queries within PyMOL and receive a stream of search results aligned to their query within a few seconds. This instant feedback cycle enables a new "designability"-inspired approach to protein design where the designer searches for and interactively incorporates native-like fragments from proven protein structures. We demonstrate the use of Suns to interactively build protein motifs, tertiary interactions, and to identify scaffolds compatible with hot-spot residues. The official web site and installer are located at http://www.degradolab.org/suns/ and the source code is hosted at https://github.com/godotgildor/Suns (PyMOL plugin, BSD license), https://github.com/Gabriel439/suns-cmd (command line client, BSD license), and https://github.com/Gabriel439/suns-search (search engine server, GPLv2 license).

Trans-species Engineering of Glycosylated Therapeutic Proteins

DEFF Research Database (Denmark)

Yang, Zhang

important to address. Whenever glycosylation has been found to be an important PTM for function or bioactivity, human therapeutics have generally been produced in mammalian Chinese hamster ovary (CHO) cell line. Oglycosylation is one of the most complex regulated PTMs of proteins but also one of the least...... understood. Currently, mammalian cells are required for human O-glycosylation. Increasing efforts have been devoted to engineering non-mammalian cells for production of recombinant proteins with “human-like” glycosylation. Substantial success has been achieved with designed N-glycosylation in both lower......Recombinant expression of therapeutic proteins is one of the major tasks in modern biomedicine. One of the most important factors with respect to therapeutic use in human is posttranslational modifications (PTMs) of the recombinant proteins, of which protein glycosylation is by far the most...

Rational Design of Disulfide Bonds Increases Thermostability of a Mesophilic 1,3-1,4-β-Glucanase from Bacillus terquilensis.

Directory of Open Access Journals (Sweden)

Chengtuo Niu

Full Text Available 1,3-1,4-β-glucanase is an important biocatalyst in brewing industry and animal feed industry, while its low thermostability often reduces its application performance. In this study, the thermostability of a mesophilic β-glucanase from Bacillus terquilensis was enhanced by rational design and engineering of disulfide bonds in the protein structure. Protein spatial configuration was analyzed to pre-exclude the residues pairs which negatively conflicted with the protein structure and ensure the contact of catalytic center. The changes in protein overall and local flexibility among the wild-type enzyme and the designated mutants were predicted to select the potential disulfide bonds for enhancement of thermostability. Two residue pairs (N31C-T187C and P102C-N125C were chosen as engineering targets and both of them were proved to significantly enhance the protein thermostability. After combinational mutagenesis, the double mutant N31C-T187C/P102C-N125C showed a 48.3% increase in half-life value at 60°C and a 4.1°C rise in melting temperature (Tm compared to wild-type enzyme. The catalytic property of N31C-T187C/P102C-N125C mutant was similar to that of wild-type enzyme. Interestingly, the optimal pH of double mutant was shifted from pH6.5 to pH6.0, which could also increase its industrial application. By comparison with mutants with single-Cys substitutions, the introduction of disulfide bonds and the induced new hydrogen bonds were proved to result in both local and overall rigidification and should be responsible for the improved thermostability. Therefore, the introduction of disulfide bonds for thermostability improvement could be rationally and highly-effectively designed by combination with spatial configuration analysis and molecular dynamics simulation.

Standby Gasoline Rationing Plan. Contingency gasoline rationing regulations

Energy Technology Data Exchange (ETDEWEB)

1979-02-01

The Economic Regulatory Administration issues final rules with respect to standby gasoline rationing. The plan is designed for and would be used only in the event of a severe gasoline shortage. The plan provides that eligibility for ration allotments will be primarily on the basis of motor vehicle registrations. DOE will mail government ration checks to the parties named in a national vehicle registration file to be maintained by DOE. Ration recipients may cash these checks for ration coupons at various designated coupon issuance points. Retail outlets and other suppliers will be required to redeem the ration coupons received in exchange for gasoline sold. Supplemental gas will be given to high-priority activities. A ration banking system will be established with two separate and distinct of ration accounts: retail outlets and other suppliers will open redemption accounts for the deposit of redeemed ration rights; and individuals or firms may open ration rights accounts, which will operate in much the same manner as monetary checking accounts. A white market will be permitted for the sale of transfer of ration rights. A percentage of the total ration rights to be issued will be reserved for distribution to the states as a State Ration Reserve, to be used by the states primarily for the relief of hardship. A National Ration Reserave will also be established. All sections of the Standby Gasoline Rationing Regulations are analyzed. (MCW)

Lentiviral Delivery of Proteins for Genome Engineering.

Science.gov (United States)

Cai, Yujia; Mikkelsen, Jacob Giehm

2016-01-01

Viruses have evolved to traverse cellular barriers and travel to the nucleus by mechanisms that involve active transport through the cytoplasm and viral quirks to resist cellular restriction factors and innate immune responses. Virus-derived vector systems exploit the capacity of viruses to ferry genetic information into cells, and now - more than three decades after the discovery of HIV - lentiviral vectors based on HIV-1 have become instrumental in biomedical research and gene therapies that require genomic insertion of transgenes. By now, the efficacy of lentiviral gene delivery to stem cells, cells of the immune system including T cells, hepatic cells, and many other therapeutically relevant cell types is well established. Along with nucleic acids, HIV-1 virions carry the enzymatic tools that are essential for early steps of infection. Such capacity to package enzymes, even proteins of nonviral origin, has unveiled new ways of exploiting cellular intrusion of HIV-1. Based on early findings demonstrating the packaging of heterologous proteins into virus particles as part of the Gag and GagPol polypeptides, we have established lentiviral protein transduction for delivery of DNA transposases and designer nucleases. This strategy for delivering genome-engineering proteins facilitates high enzymatic activity within a short time frame and may potentially improve the safety of genome editing. Exploiting the full potential of lentiviral vectors, incorporation of foreign protein can be combined with the delivery of DNA transposons or a donor sequence for homology-directed repair in so-called 'all-in-one' lentiviral vectors. Here, we briefly describe intracellular restrictions that may affect lentiviral gene and protein delivery and review the current status of lentiviral particles as carriers of tool kits for genome engineering.

Respon Pertumbuhan Ayam Lokal Pedaging terhadap Suplementasi Protein Isolasi Biji-bijian (PIB dan Perbedaan Level Protein Ransum

Directory of Open Access Journals (Sweden)

M. Aman Yaman

2009-10-01

Full Text Available The response of local meat chicken growth to supplementation of isolated grain protein and the difference in ration protein level ABSTRACT. A research which aims to determine the response of local meat chicken growth of protein supplementation with Isolation Grains Protein (IGB and the difference in ration protein level has been conducted in the Laboratory of Experimental Farm, Animal Husbandry Department, Faculty of Agriculture, Syiah Kuala University-Darussalam, Banda Aceh for 90 days. This study used a completely randomized design factorial with 2 factors, consisting of factors namely male gender (JJ and female (JB and the ration is a combination of factors and levels IGB in the ration, ie: treatment A: 17% protein and 0.4% IGB; treatment B 19% protein and 0.6% IGB and treatment C 21% protein and 0.8% IGB. Each combination consisted of 4 replications and each replication consists of 5 chickens. Parameters observed in the study were weight gain, achievement of final weight, consumption, conversion and efficiency of ration. DOC used a derivative result of selection of local meat chicken which are in the process of selection. Data acquired and processed by ANOVA. The results showed that supplementation of IGB and ration protein level difference was significantly effect (P <0.01 on weight gain, final weight, rate of consumption, conversion efficiency of rations and rations, but there is no interaction effect between sex and ration factors . The highest weight gain obtained in the male local chicken achieved by feeding a ration B (93.23 grams, while the hen rations achieved by providing treatment C (63.86 grams / week. The highest final body weight of male chicken on treatment B (1491.5 gram/90 days and hens in treatment C (1061.5 gram/90 days. However, the highest ration consumption in both male and female local chickens obtained from the ration A. Feed conversion value and the best feed efficiency obtained in treatment B for the treatment of

Production of biopharmaceutical proteins by yeast: Advances through metabolic engineering

DEFF Research Database (Denmark)

Nielsen, Jens

2013-01-01

Production of recombinant proteins for use as pharmaceuticals, so-called biopharmaceuticals, is a multi-billion dollar industry. Many different cell factories are used for the production of biopharmaceuticals, but the yeast Saccharomyces cerevisiae is an important cell factory as it is used for p...... production. The involvement of directed metabolic engineering through the integration of tools from genetic engineering, systems biology and mathematical modeling, is also discussed....... by yeast are human serum albumin, hepatitis vaccines and virus like particles used for vaccination against human papillomavirus. Here is given a brief overview of biopharmaceutical production by yeast and it is discussed how the secretory pathway can be engineered to ensure more efficient protein...

Rational and efficient preparation of a chimeric protein containing a tandem dimer of thrombopoietin mimetic peptide fused to human growth hormone in Escherichia coli.

Science.gov (United States)

Wang, Song; Shen, Mingqiang; Xu, Yang; Chen, Fang; Chen, Mo; Chen, Shilei; Wang, Aiping; Zhang, Zhou; Ran, Xinze; Cheng, Tianmin; Su, Yongping; Wang, Junping

2013-04-01

The 14-mer thrombopoietin mimetic peptide (TMP), especially in the form of dimer, displayed potent megakaryocytopoiesis activity in vitro. However, it is difficult to prepare such short peptide with high bioactivity through gene-engineering approaches. In this study, a chimeric protein containing a tandem dimer of TMP (dTMP) fused to human growth hormone (hGH), a kind of hematopoietic growth factor that activates the same signal pathways as thrombopoietin, was produced in Escherichia coli by soluble expression. By rational utilization of the XmnI and EcoRV restriction sites, a PCR fragment encoding dTMP-GH was inserted into the plasmid vector pMAL-p2X at the position right after Xa factor cleavage site, in frame with maltose-binding protein (MBP) gene. Under optimized conditions, a high-level expression of soluble MBP-dTMP-GH fusion protein was obtained. By application of amylose resin chromatography, Xa factor digestion, hydrophobic chromatography followed by gel filtration, the dTMP-GH fusion protein was separated. Finally, a relatively high yield of dTMP-GH fusion protein with high purity (>98%) and without redundant amino acid was achieved, as identified by high-performance liquid chromatography, mass spectrometry, and amino acid sequencing. The functional assays showed that dTMP-GH could promote the proliferation of megakaryoblast cells and maturation of murine megakaryocytes derived from bone marrow, in a dose-dependent manner. Moreover, an enhanced effect of dTMP-GH on megakaryocytopoiesis was found as compared with equimolar concentration of dTMP and rhGH. This work provides a new avenue to generate thrombopoietic agents based on TMP.

Modelling of microbial polyhydroxyalkanoate surface binding protein PhaP for rational mutagenesis.

Science.gov (United States)

Zhao, Hongyu; Yao, Zhenyu; Chen, Xiangbin; Wang, Xinquan; Chen, Guo-Qiang

2017-11-01

Phasins are unusual amphiphilic proteins that bind to microbial polyhydroxyalkanoate (PHA) granules in nature and show great potential for various applications in biotechnology and medicine. Despite their remarkable diversity, only the crystal structure of PhaP A h from Aeromonas hydrophila has been solved to date. Based on the structure of PhaP A h , homology models of PhaP A z from Azotobacter sp. FA-8 and PhaP TD from Halomonas bluephagenesis TD were successfully established, allowing rational mutagenesis to be conducted to enhance the stability and surfactant properties of these proteins. PhaP A z mutants, including PhaP A z Q38L and PhaP A z Q78L, as well as PhaP TD mutants, including PhaP TD Q38M and PhaP TD Q72M, showed better emulsification properties and improved thermostability (6-10°C higher melting temperatures) compared with their wild-type homologues under the same conditions. Importantly, the established PhaP homology-modelling approach, based on the high-resolution structure of PhaP A h , can be generalized to facilitate the study of other PhaP members. © 2017 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

Biotechnology Conference: Protein Engineering Held in Oxford, United Kingdom on 5-8 April 1987.

Science.gov (United States)

1987-07-27

engineered by protein engineering was reported by J. new variants which are now being checked. Brange (Novo Research Institute, Bags- Studies of a cassette...to Brange . Therefore, multidomain protein consisting of five Brange and his group applied protein en- putative domains: the fribonectin finger

Rationality of limited rationality : some aggregate implications

OpenAIRE

Uri M. Possen; Mikko Puhakka

1994-01-01

In this paper we let economic agents choose whether to become fully rational or stay boundedly rational. Boundedly rational agents are less sophisticated in their information processing abilities. It is costly to acquire information needed to become fully rational, and thus not all agents are willing to incur those costs. We then explore the aggregate effects of endogenizing the decision whether the agent should or should not become fully rational in handling information. Since fully and boun...

Biomineralization of Engineered Spider Silk Protein-Based Composite Materials for Bone Tissue Engineering

Directory of Open Access Journals (Sweden)

John G. Hardy

2016-07-01

Full Text Available Materials based on biodegradable polyesters, such as poly(butylene terephthalate (PBT or poly(butylene terephthalate-co-poly(alkylene glycol terephthalate (PBTAT, have potential application as pro-regenerative scaffolds for bone tissue engineering. Herein, the preparation of films composed of PBT or PBTAT and an engineered spider silk protein, (eADF4(C16, that displays multiple carboxylic acid moieties capable of binding calcium ions and facilitating their biomineralization with calcium carbonate or calcium phosphate is reported. Human mesenchymal stem cells cultured on films mineralized with calcium phosphate show enhanced levels of alkaline phosphatase activity suggesting that such composites have potential use for bone tissue engineering.

Integrating the protein and metabolic engineering toolkits for next-generation chemical biosynthesis.

Science.gov (United States)

Pirie, Christopher M; De Mey, Marjan; Jones Prather, Kristala L; Ajikumar, Parayil Kumaran

2013-04-19

Through microbial engineering, biosynthesis has the potential to produce thousands of chemicals used in everyday life. Metabolic engineering and synthetic biology are fields driven by the manipulation of genes, genetic regulatory systems, and enzymatic pathways for developing highly productive microbial strains. Fundamentally, it is the biochemical characteristics of the enzymes themselves that dictate flux through a biosynthetic pathway toward the product of interest. As metabolic engineers target sophisticated secondary metabolites, there has been little recognition of the reduced catalytic activity and increased substrate/product promiscuity of the corresponding enzymes compared to those of central metabolism. Thus, fine-tuning these enzymatic characteristics through protein engineering is paramount for developing high-productivity microbial strains for secondary metabolites. Here, we describe the importance of protein engineering for advancing metabolic engineering of secondary metabolism pathways. This pathway integrated enzyme optimization can enhance the collective toolkit of microbial engineering to shape the future of chemical manufacturing.

Residue-specific incorporation of noncanonical amino acids for protein engineering

NARCIS (Netherlands)

van Eldijk, Mark B.; van Hest, Jan C.M.; Lemke, E.A.

2018-01-01

The incorporation of noncanonical amino acids has given protein chemists access to an expanded repertoire of amino acids. This methodology has significantly broadened the scope of protein engineering allowing introduction of amino acids with non-native functionalities, such as bioorthogonal reactive

Engineering growth factors for regenerative medicine applications.

Energy Technology Data Exchange (ETDEWEB)

Mitchell, Aaron C.; Briquez, Priscilla S.; Hubbell, Jeffrey A.; Cochran, Jennifer R.

2016-01-15

Growth factors are important morphogenetic proteins that instruct cell behavior and guide tissue repair and renewal. Although their therapeutic potential holds great promise in regenerative medicine applications, translation of growth factors into clinical treatments has been hindered by limitations including poor protein stability, low recombinant expression yield, and suboptimal efficacy. This review highlights current tools, technologies, and approaches to design integrated and effective growth factor-based therapies for regenerative medicine applications. The first section describes rational and combinatorial protein engineering approaches that have been utilized to improve growth factor stability, expression yield, biodistribution, and serum half-life, or alter their cell trafficking behavior or receptor binding affinity. The second section highlights elegant biomaterial-based systems, inspired by the natural extracellular matrix milieu, that have been developed for effective spatial and temporal delivery of growth factors to cell surface receptors. Although appearing distinct, these two approaches are highly complementary and involve principles of molecular design and engineering to be considered in parallel when developing optimal materials for clinical applications.

Rational Design of an Ultrasensitive Quorum-Sensing Switch.

Science.gov (United States)

Zeng, Weiqian; Du, Pei; Lou, Qiuli; Wu, Lili; Zhang, Haoqian M; Lou, Chunbo; Wang, Hongli; Ouyang, Qi

2017-08-18

One of the purposes of synthetic biology is to develop rational methods that accelerate the design of genetic circuits, saving time and effort spent on experiments and providing reliably predictable circuit performance. We applied a reverse engineering approach to design an ultrasensitive transcriptional quorum-sensing switch. We want to explore how systems biology can guide synthetic biology in the choice of specific DNA sequences and their regulatory relations to achieve a targeted function. The workflow comprises network enumeration that achieves the target function robustly, experimental restriction of the obtained candidate networks, global parameter optimization via mathematical analysis, selection and engineering of parts based on these calculations, and finally, circuit construction based on the principles of standardization and modularization. The performance of realized quorum-sensing switches was in good qualitative agreement with the computational predictions. This study provides practical principles for the rational design of genetic circuits with targeted functions.

A rapid, ensemble and free energy based method for engineering protein stabilities.

Science.gov (United States)

Naganathan, Athi N

2013-05-02

Engineering the conformational stabilities of proteins through mutations has immense potential in biotechnological applications. It is, however, an inherently challenging problem given the weak noncovalent nature of the stabilizing interactions. In this regard, we present here a robust and fast strategy to engineer protein stabilities through mutations involving charged residues using a structure-based statistical mechanical model that accounts for the ensemble nature of folding. We validate the method by predicting the absolute changes in stability for 138 experimental mutations from 16 different proteins and enzymes with a correlation of 0.65 and importantly with a success rate of 81%. Multiple point mutants are predicted with a higher success rate (90%) that is validated further by comparing meosphile-thermophile protein pairs. In parallel, we devise a methodology to rapidly engineer mutations in silico which we benchmark against experimental mutations of ubiquitin (correlation of 0.95) and check for its feasibility on a larger therapeutic protein DNase I. We expect the method to be of importance as a first and rapid step to screen for protein mutants with specific stability in the biotechnology industry, in the construction of stability maps at the residue level (i.e., hot spots), and as a robust tool to probe for mutations that enhance the stability of protein-based drugs.

Rudolf Diesel – The Rational Inventor of a Heat Engine

Indian Academy of Sciences (India)

München) where one of his professors was a young Carl von Linde who is famous ... into English in 1894 and is called “the theory and construction of a rational ... Diesel and everyone at the exhibition were absolutely right in predicting that this.

Engineered Proteins Program Mammalian Cells to Target Inflammatory Disease Sites.

Science.gov (United States)

Qudrat, Anam; Mosabbir, Abdullah Al; Truong, Kevin

2017-06-22

Disease sites in atherosclerosis and cancer feature cell masses (e.g., plaques/tumors), a low pH extracellular microenvironment, and various pro-inflammatory cytokines such as tumor necrosis factor α (TNFα). The ability to engineer a cell to seek TNFα sources allows for targeted therapeutic delivery. To accomplish this, here we introduced a system of proteins: an engineered TNFα chimeric receptor (named TNFR1chi), a previously engineered Ca 2+ -activated RhoA (named CaRQ), vesicular stomatitis virus glycoprotein G (VSVG), and thymidine kinase. Upon binding TNFα, TNFR1chi generates a Ca 2+ signal that in turn activates CaRQ-mediated non-apoptotic blebs that allow migration toward the TNFα source. Next, the addition of VSVG, upon low pH induction, causes membrane fusion of the engineered and TNFα source cells. Finally, after ganciclovir treatment cells undergo death via the thymidine kinase suicide mechanism. Hence, we assembled a system of proteins that forms the basis of engineering a cell to target inflammatory disease sites characterized by TNFα secretion and a low-pH microenvironment. Copyright © 2017 Elsevier Ltd. All rights reserved.

Engineered proteins with PUF scaffold to manipulate RNA metabolism

Science.gov (United States)

Wang, Yang; Wang, Zefeng; Tanaka Hall, Traci M.

2013-01-01

Pumilio/fem-3 mRNA binding factor (FBF) proteins are characterized by a sequence-specific RNA-binding domain. This unique single-stranded RNA recognition module, whose sequence specificity can be reprogrammed, has been fused with functional modules to engineer protein factors with various functions. Here we summarize the advancement in developing RNA regulatory tools and opportunities for the future. PMID:23731364

A Rational Engineering Strategy for Designing Protein A-Binding Camelid Single-Domain Antibodies

Science.gov (United States)

Henry, Kevin A.; Sulea, Traian; van Faassen, Henk; Hussack, Greg; Purisima, Enrico O.; MacKenzie, C. Roger; Arbabi-Ghahroudi, Mehdi

2016-01-01

Staphylococcal protein A (SpA) and streptococcal protein G (SpG) affinity chromatography are the gold standards for purifying monoclonal antibodies (mAbs) in therapeutic applications. However, camelid VHH single-domain Abs (sdAbs or VHHs) are not bound by SpG and only sporadically bound by SpA. Currently, VHHs require affinity tag-based purification, which limits their therapeutic potential and adds considerable complexity and cost to their production. Here we describe a simple and rapid mutagenesis-based approach designed to confer SpA binding upon a priori non-SpA-binding VHHs. We show that SpA binding of VHHs is determined primarily by the same set of residues as in human mAbs, albeit with an unexpected degree of tolerance to substitutions at certain core and non-core positions and some limited dependence on at least one residue outside the SpA interface, and that SpA binding could be successfully introduced into five VHHs against three different targets with no adverse effects on expression yield or antigen binding. Next-generation sequencing of llama, alpaca and dromedary VHH repertoires suggested that species differences in SpA binding may result from frequency variation in specific deleterious polymorphisms, especially Ile57. Thus, the SpA binding phenotype of camelid VHHs can be easily modulated to take advantage of tag-less purification techniques, although the frequency with which this is required may depend on the source species. PMID:27631624

Engineering Synthetic Proteins to Generate Ca2+ Signals in Mammalian Cells.

Science.gov (United States)

Qudrat, Anam; Truong, Kevin

2017-03-17

The versatility of Ca 2+ signals allows it to regulate diverse cellular processes such as migration, apoptosis, motility and exocytosis. In some receptors (e.g., VEGFR2), Ca 2+ signals are generated upon binding their ligand(s) (e.g., VEGF-A). Here, we employed a design strategy to engineer proteins that generate a Ca 2+ signal upon binding various extracellular stimuli by creating fusions of protein domains that oligomerize to the transmembrane domain and the cytoplasmic tail of the VEGFR2. To test the strategy, we created chimeric proteins that generate Ca 2+ signals upon stimulation with various extracellular stimuli (e.g., rapamycin, EDTA or extracellular free Ca 2+ ). By coupling these chimeric proteins that generate Ca 2+ signals with proteins that respond to Ca 2+ signals, we rewired, for example, dynamic cellular blebbing to increases in extracellular free Ca 2+ . Thus, using this design strategy, it is possible to engineer proteins to generate a Ca 2+ signal to rewire a wide range of extracellular stimuli to a wide range of Ca 2+ -activated processes.

ProtaBank: A repository for protein design and engineering data.

Science.gov (United States)

Wang, Connie Y; Chang, Paul M; Ary, Marie L; Allen, Benjamin D; Chica, Roberto A; Mayo, Stephen L; Olafson, Barry D

2018-03-25

We present ProtaBank, a repository for storing, querying, analyzing, and sharing protein design and engineering data in an actively maintained and updated database. ProtaBank provides a format to describe and compare all types of protein mutational data, spanning a wide range of properties and techniques. It features a user-friendly web interface and programming layer that streamlines data deposition and allows for batch input and queries. The database schema design incorporates a standard format for reporting protein sequences and experimental data that facilitates comparison of results across different data sets. A suite of analysis and visualization tools are provided to facilitate discovery, to guide future designs, and to benchmark and train new predictive tools and algorithms. ProtaBank will provide a valuable resource to the protein engineering community by storing and safeguarding newly generated data, allowing for fast searching and identification of relevant data from the existing literature, and exploring correlations between disparate data sets. ProtaBank invites researchers to contribute data to the database to make it accessible for search and analysis. ProtaBank is available at https://protabank.org. © 2018 The Authors Protein Science published by Wiley Periodicals, Inc. on behalf of The Protein Society.

Coupling ligand recognition to protein folding in an engineered variant of rabbit ileal lipid binding protein.

Science.gov (United States)

Kouvatsos, Nikolaos; Meldrum, Jill K; Searle, Mark S; Thomas, Neil R

2006-11-28

We have engineered a variant of the beta-clam shell protein ILBP which lacks the alpha-helical motif that caps the central binding cavity; the mutant protein is sufficiently destabilised that it is unfolded under physiological conditions, however, it unexpectedly binds its natural bile acid substrates with high affinity forming a native-like beta-sheet rich structure and demonstrating strong thermodynamic coupling between ligand binding and protein folding.

Novel blood protein based scaffolds for cardiovascular tissue engineering

Directory of Open Access Journals (Sweden)

Kuhn Antonia I.

2016-09-01

Full Text Available A major challenge in cardiovascular tissue engineering is the fabrication of scaffolds, which provide appropriate morphological and mechanical properties while avoiding undesirable immune reactions. In this study electrospinning was used to fabricate scaffolds out of blood proteins for cardiovascular tissue engineering. Lyophilised porcine plasma was dissolved in deionised water at a final concentration of 7.5% m/v and blended with 3.7% m/v PEO. Electrospinning resulted in homogeneous fibre morphologies with a mean fibre diameter of 151 nm, which could be adapted to create macroscopic shapes (mats, tubes. Cross-linking with glutaraldehyde vapour improved the long-term stability of protein based scaffolds in comparison to untreated scaffolds, resulting in a mass loss of 41% and 96% after 28 days of incubation in aqueous solution, respectively.

Tuning calcite morphology and growth acceleration by a rational design of highly stable protein-mimetics

Science.gov (United States)

Chen, Chun-Long; Qi, Jiahui; Tao, Jinhui; Zuckermann, Ronald N.; DeYoreo, James J.

2014-01-01

In nature, proteins play a significant role in biomineral formation. One of the ultimate goals of bioinspired materials science is to develop highly stable synthetic molecules that mimic the function of these natural proteins by controlling crystal formation. Here, we demonstrate that both the morphology and the degree of acceleration or inhibition observed during growth of calcite in the presence of peptoids can be rationally tuned by balancing the electrostatic and hydrophobic interactions, with hydrophobic interactions playing the dominant role. While either strong electrostatic or hydrophobic interactions inhibit growth and reduces expression of the {104} faces, correlations between peptoid-crystal binding energies and observed changes in calcite growth indicate moderate electrostatic interactions allow peptoids to weakly adsorb while moderate hydrophobic interactions cause disruption of surface-adsorbed water layers, leading to growth acceleration with retained expression of the {104} faces. This study provides fundamental principles for designing peptoids as crystallization promoters, and offers a straightforward screening method based on macroscopic crystal morphology. Because peptoids are sequence-specific, highly stable, and easily synthesized, peptoid-enhanced crystallization offers a broad range of potential applications. PMID:25189418

De novo design and engineering of functional metal and porphyrin-binding protein domains

Science.gov (United States)

Everson, Bernard H.

In this work, I describe an approach to the rational, iterative design and characterization of two functional cofactor-binding protein domains. First, a hybrid computational/experimental method was developed with the aim of algorithmically generating a suite of porphyrin-binding protein sequences with minimal mutual sequence information. This method was explored by generating libraries of sequences, which were then expressed and evaluated for function. One successful sequence is shown to bind a variety of porphyrin-like cofactors, and exhibits light- activated electron transfer in mixed hemin:chlorin e6 and hemin:Zn(II)-protoporphyrin IX complexes. These results imply that many sophisticated functions such as cofactor binding and electron transfer require only a very small number of residue positions in a protein sequence to be fixed. Net charge and hydrophobic content are important in determining protein solubility and stability. Accordingly, rational modifications were made to the aforementioned design procedure in order to improve its overall success rate. The effects of these modifications are explored using two `next-generation' sequence libraries, which were separately expressed and evaluated. Particular modifications to these design parameters are demonstrated to effectively double the purification success rate of the procedure. Finally, I describe the redesign of the artificial di-iron protein DF2 into CDM13, a single chain di-Manganese four-helix bundle. CDM13 acts as a functional model of natural manganese catalase, exhibiting a kcat of 0.08s-1 under steady-state conditions. The bound manganese cofactors have a reduction potential of +805 mV vs NHE, which is too high for efficient dismutation of hydrogen peroxide. These results indicate that as a high-potential manganese complex, CDM13 may represent a promising first step toward a polypeptide model of the Oxygen Evolving Complex of the photosynthetic enzyme Photosystem II.

Engineering self-assembled bioreactors from protein microcompartments

Energy Technology Data Exchange (ETDEWEB)

Savage, David [Univ. of California, Berkeley, CA (United States)

2016-10-12

The goals of this research are to understand how organisms such as bacteria segregate certain metabolic processes inside of specific structures, or “microcompartments,” in the cell and apply this knowledge to develop novel engineered microcompartments for use in nanotechnology and metabolic engineering. For example, in some bacteria, self-assembling protein microcompartments called carboxysomes encapsulate the enzymes involved in carbon fixation, enabling the cell to utilize carbon dioxide more effectively than if the enzymes were free in the cell. The proposed research will determine how structures such as carboxysomes assemble and function in bacteria and develop a means for creating novel, synthetic microcompartments for optimizing production of specific energy-rich compounds.

Hot-spot analysis for drug discovery targeting protein-protein interactions.

Science.gov (United States)

Rosell, Mireia; Fernández-Recio, Juan

2018-04-01

Protein-protein interactions are important for biological processes and pathological situations, and are attractive targets for drug discovery. However, rational drug design targeting protein-protein interactions is still highly challenging. Hot-spot residues are seen as the best option to target such interactions, but their identification requires detailed structural and energetic characterization, which is only available for a tiny fraction of protein interactions. Areas covered: In this review, the authors cover a variety of computational methods that have been reported for the energetic analysis of protein-protein interfaces in search of hot-spots, and the structural modeling of protein-protein complexes by docking. This can help to rationalize the discovery of small-molecule inhibitors of protein-protein interfaces of therapeutic interest. Computational analysis and docking can help to locate the interface, molecular dynamics can be used to find suitable cavities, and hot-spot predictions can focus the search for inhibitors of protein-protein interactions. Expert opinion: A major difficulty for applying rational drug design methods to protein-protein interactions is that in the majority of cases the complex structure is not available. Fortunately, computational docking can complement experimental data. An interesting aspect to explore in the future is the integration of these strategies for targeting PPIs with large-scale mutational analysis.

Efficient expression of SRK intracellular domain by a modeling-based protein engineering.

Science.gov (United States)

Murase, Kohji; Hirano, Yoshinori; Takayama, Seiji; Hakoshima, Toshio

2017-03-01

S-locus protein kinase (SRK) is a receptor kinase that plays a critical role in self-recognition in the Brassicaceae self-incompatibility (SI) response. SRK is activated by binding of its ligand S-locus protein 11 (SP11) and subsequently induced phosphorylation of the intracellular kinase domain. However, a detailed activation mechanism of SRK is still largely unknown because of the difficulty in stably expressing SRK recombinant proteins. Here, we performed modeling-based protein engineering of the SRK kinase domain for stable expression in Escherichia coli. The engineered SRK intracellular domain was expressed about 54-fold higher production than wild type SRK, without loss of the kinase activity, suggesting it could be useful for further biochemical and structural studies. Copyright © 2016 Elsevier Inc. All rights reserved.

Rationalization: A Bibliography.

Science.gov (United States)

Pedrini, D. T.; Pedrini, Bonnie C.

Rationalization was studied by Sigmund Freud and was specifically labeled by Ernest Jones. Rationalization ought to be differentiated from rational, rationality, logical analysis, etc. On the one hand, rationalization is considered a defense mechanism, on the other hand, rationality is not. Haan has done much work with self-report inventories and…

Engineering protein scaffolds for protein separation, biocatalysis and nanotechnology applications

Science.gov (United States)

Liu, Fang

Globally, there is growing appreciation for developing a sustainable economy that uses eco-efficient bio-processes. Biotechnology provides an increasing range of tools for industry to help reduce cost and improve environmental performance. Inspired by the naturally evolved machineries of protein scaffolds and their binding ligands, synthetic protein scaffolds were engineered based on cohesin-dockerin interactions and metal chelating peptides to tackle the challenges and make improvements in three specific areas: (1) protein purification, (2) biofuel cells, and (3) nanomaterial synthesis. The first objective was to develop efficient and cost-effective non-chromatographic purification processes to purify recombinant proteins in an effort to meet the dramatically growing market of protein drugs. In our design, the target protein was genetically fused with a dockerin domain from Clostridium thermocellum and direct purification and recovery was achieved using thermo-responsive elastin-like polypeptide (ELP) scaffold containing the cohesin domain from the same species. By exploiting the highly specific interaction between the dockerin and cohesin domain and the reversible aggregation property of ELP, highly purified and active dockerin-tagged proteins, such as endoglucanase CelA, chloramphenicol acetyl transferase (CAT) and enhanced green fluorescence protein (EGFP), were recovered directly from crude cell extracts in a single purification step with yields achieving over 90%. Incorporation of a self-cleaving intein domain enabled rapid removal of the affinity tag from the target proteins by another cycle of thermal precipitation. The purification cost can be further reduced by regenerating and recycling the ELP-cohesin capturing scaffolds. However, due to the high binding affinity between cohesin and dockerin domains, the bound dockerin-intein tag cannot be completely disassociated from ELP-cohesin scaffold after binding. Therefore, a truncated dockerin with the calcium

Cognitive science as an interface between rational and mechanistic explanation.

Science.gov (United States)

Chater, Nick

2014-04-01

Cognitive science views thought as computation; and computation, by its very nature, can be understood in both rational and mechanistic terms. In rational terms, a computation solves some information processing problem (e.g., mapping sensory information into a description of the external world; parsing a sentence; selecting among a set of possible actions). In mechanistic terms, a computation corresponds to causal chain of events in a physical device (in engineering context, a silicon chip; in biological context, the nervous system). The discipline is thus at the interface between two very different styles of explanation--as the papers in the current special issue well illustrate, it explores the interplay of rational and mechanistic forces. Copyright © 2014 Cognitive Science Society, Inc.

A Simple Combinatorial Codon Mutagenesis Method for Targeted Protein Engineering.

Science.gov (United States)

Belsare, Ketaki D; Andorfer, Mary C; Cardenas, Frida S; Chael, Julia R; Park, Hyun June; Lewis, Jared C

2017-03-17

Directed evolution is a powerful tool for optimizing enzymes, and mutagenesis methods that improve enzyme library quality can significantly expedite the evolution process. Here, we report a simple method for targeted combinatorial codon mutagenesis (CCM). To demonstrate the utility of this method for protein engineering, CCM libraries were constructed for cytochrome P450 BM3 , pfu prolyl oligopeptidase, and the flavin-dependent halogenase RebH; 10-26 sites were targeted for codon mutagenesis in each of these enzymes, and libraries with a tunable average of 1-7 codon mutations per gene were generated. Each of these libraries provided improved enzymes for their respective transformations, which highlights the generality, simplicity, and tunability of CCM for targeted protein engineering.

Soy Protein Scaffold Biomaterials for Tissue Engineering and Regenerative Medicine

Science.gov (United States)

Chien, Karen B.

Developing functional biomaterials using highly processable materials with tailorable physical and bioactive properties is an ongoing challenge in tissue engineering. Soy protein is an abundant, natural resource with potential use for regenerative medicine applications. Preliminary studies show that soy protein can be physically modified and fabricated into various biocompatible constructs. However, optimized soy protein structures for tissue regeneration (i.e. 3D porous scaffolds) have not yet been designed. Furthermore, little work has established the in vivo biocompatibility of implanted soy protein and the benefit of using soy over other proteins including FDA-approved bovine collagen. In this work, freeze-drying and 3D printing fabrication processes were developed using commercially available soy protein to create porous scaffolds that improve cell growth and infiltration compared to other soy biomaterials previously reported. Characterization of scaffold structure, porosity, and mechanical/degradation properties was performed. In addition, the behavior of human mesenchymal stem cells seeded on various designed soy scaffolds was analyzed. Biological characterization of the cell-seeded scaffolds was performed to assess feasibility for use in liver tissue regeneration. The acute and humoral response of soy scaffolds implanted in an in vivo mouse subcutaneous model was also investigated. All fabricated soy scaffolds were modified using thermal, chemical, and enzymatic crosslinking to change properties and cell growth behavior. 3D printing allowed for control of scaffold pore size and geometry. Scaffold structure, porosity, and degradation rate significantly altered the in vivo response. Freeze-dried soy scaffolds had similar biocompatibility as freeze-dried collagen scaffolds of the same protein content. However, the soy scaffolds degraded at a much faster rate, minimizing immunogenicity. Interestingly, subcutaneously implanted soy scaffolds affected blood

Rational Decision Making as Performative Praxis: Explaining Rationality's Éternel Retour

OpenAIRE

Cabantous, L.; Gond, J-P.

2011-01-01

Organizational theorists built their knowledge of decision making through a progressive critique of rational choice theory. Their positioning towards rationality, however, is at odds with the observation of rationality persistence in organizational life. This paper addresses this paradox. It proposes a new perspective on rationality that allows the theorizing of the production of rational decisions by organizations. To account for rationality's éternel retour, we approach rational decision ma...

Engineering of an E. coli outer membrane protein FhuA with increased channel diameter

Directory of Open Access Journals (Sweden)

Dworeck Tamara

2011-08-01

Full Text Available Abstract Background Channel proteins like FhuA can be an alternative to artificial chemically synthesized nanopores. To reach such goals, channel proteins must be flexible enough to be modified in their geometry, i.e. length and diameter. As continuation of a previous study in which we addressed the lengthening of the channel, here we report the increasing of the channel diameter by genetic engineering. Results The FhuA Δ1-159 diameter increase has been obtained by doubling the amino acid sequence of the first two N-terminal β-strands, resulting in variant FhuA Δ1-159 Exp. The total number of β-strands increased from 22 to 24 and the channel surface area is expected to increase by ~16%. The secondary structure analysis by circular dichroism (CD spectroscopy shows a high β-sheet content, suggesting the correct folding of FhuA Δ1-159 Exp. To further prove the FhuA Δ1-159 Exp channel functionality, kinetic measurement using the HRP-TMB assay (HRP = Horse Radish Peroxidase, TMB = 3,3',5,5'-tetramethylbenzidine were conducted. The results indicated a 17% faster diffusion kinetic for FhuA Δ1-159 Exp as compared to FhuA Δ1-159, well correlated to the expected channel surface area increase of ~16%. Conclusion In this study using a simple "semi rational" approach the FhuA Δ1-159 diameter was enlarged. By combining the actual results with the previous ones on the FhuA Δ1-159 lengthening a new set of synthetic nanochannels with desired lengths and diameters can be produced, broadening the FhuA Δ1-159 applications. As large scale protein production is possible our approach can give a contribution to nanochannel industrial applications.

Development of a High-Efficiency Transformation Method and Implementation of Rational Metabolic Engineering for the Industrial Butanol Hyperproducer Clostridium saccharoperbutylacetonicum Strain N1-4.

Science.gov (United States)

Herman, Nicolaus A; Li, Jeffrey; Bedi, Ripika; Turchi, Barbara; Liu, Xiaoji; Miller, Michael J; Zhang, Wenjun

2017-01-15

While a majority of academic studies concerning acetone, butanol, and ethanol (ABE) production by Clostridium have focused on Clostridium acetobutylicum, other members of this genus have proven to be effective industrial workhorses despite the inability to perform genetic manipulations on many of these strains. To further improve the industrial performance of these strains in areas such as substrate usage, solvent production, and end product versatility, transformation methods and genetic tools are needed to overcome the genetic intractability displayed by these species. In this study, we present the development of a high-efficiency transformation method for the industrial butanol hyperproducer Clostridium saccharoperbutylacetonicum strain N1-4 (HMT) ATCC 27021. Following initial failures, we found that the key to creating a successful transformation method was the identification of three distinct colony morphologies (types S, R, and I), which displayed significant differences in transformability. Working with the readily transformable type I cells (transformation efficiency, 1.1 × 10 6 CFU/μg DNA), we performed targeted gene deletions in C. saccharoperbutylacetonicum N1-4 using a homologous recombination-mediated allelic exchange method. Using plasmid-based gene overexpression and targeted knockouts of key genes in the native acetone-butanol-ethanol (ABE) metabolic pathway, we successfully implemented rational metabolic engineering strategies, yielding in the best case an engineered strain (Clostridium saccharoperbutylacetonicum strain N1-4/pWIS13) displaying an 18% increase in butanol titers and 30% increase in total ABE titer (0.35 g ABE/g sucrose) in batch fermentations. Additionally, two engineered strains overexpressing aldehyde/alcohol dehydrogenases (encoded by adh11 and adh5) displayed 8.5- and 11.8-fold increases (respectively) in batch ethanol production. This paper presents the first steps toward advanced genetic engineering of the industrial butanol

Engineered elastomeric proteins with dual elasticity can be controlled by a molecular regulator.

Science.gov (United States)

Cao, Yi; Li, Hongbin

2008-08-01

Elastomeric proteins are molecular springs that confer excellent mechanical properties to many biological tissues and biomaterials. Depending on the role performed by the tissue or biomaterial, elastomeric proteins can behave as molecular springs or shock absorbers. Here we combine single-molecule atomic force microscopy and protein engineering techniques to create elastomeric proteins that can switch between two distinct types of mechanical behaviour in response to the binding of a molecular regulator. The proteins are mechanically labile by design and behave as entropic springs with an elasticity that is governed by their configurational entropy. However, when a molecular regulator binds to the protein, it switches into a mechanically stable state and can act as a shock absorber. These engineered proteins effectively mimic and combine the two extreme forms of elastic behaviour found in natural elastomeric proteins, and thus represent a new type of smart nanomaterial that will find potential applications in nanomechanics and material sciences.

Probing Enzyme-Surface Interactions via Protein Engineering and Single-Molecule Techniques

Science.gov (United States)

2017-06-26

SECURITY CLASSIFICATION OF: The overall objective of this research was to exploit protein engineering and fluorescence single-molecule methods to...enhance our understanding of the interaction of proteins and surfaces. Given this objective, the specific aims of this research were to: 1) exploit the...incorporation of unnatural amino acids in proteins to introduce single-molecule probes (i.e., fluorophores for fluorescence resonance energy transfer

Cell-free protein synthesis enabled rapid prototyping for metabolic engineering and synthetic biology

Directory of Open Access Journals (Sweden)

Lihong Jiang

2018-06-01

Full Text Available Advances in metabolic engineering and synthetic biology have facilitated the manufacturing of many valuable-added compounds and commodity chemicals using microbial cell factories in the past decade. However, due to complexity of cellular metabolism, the optimization of metabolic pathways for maximal production represents a grand challenge and an unavoidable barrier for metabolic engineering. Recently, cell-free protein synthesis system (CFPS has been emerging as an enabling alternative to address challenges in biomanufacturing. This review summarizes the recent progresses of CFPS in rapid prototyping of biosynthetic pathways and genetic circuits (biosensors to speed up design-build-test (DBT cycles of metabolic engineering and synthetic biology. Keywords: Cell-free protein synthesis, Metabolic pathway optimization, Genetic circuits, Metabolic engineering, Synthetic biology

Gene composer: database software for protein construct design, codon engineering, and gene synthesis.

Science.gov (United States)

Lorimer, Don; Raymond, Amy; Walchli, John; Mixon, Mark; Barrow, Adrienne; Wallace, Ellen; Grice, Rena; Burgin, Alex; Stewart, Lance

2009-04-21

To improve efficiency in high throughput protein structure determination, we have developed a database software package, Gene Composer, which facilitates the information-rich design of protein constructs and their codon engineered synthetic gene sequences. With its modular workflow design and numerous graphical user interfaces, Gene Composer enables researchers to perform all common bio-informatics steps used in modern structure guided protein engineering and synthetic gene engineering. An interactive Alignment Viewer allows the researcher to simultaneously visualize sequence conservation in the context of known protein secondary structure, ligand contacts, water contacts, crystal contacts, B-factors, solvent accessible area, residue property type and several other useful property views. The Construct Design Module enables the facile design of novel protein constructs with altered N- and C-termini, internal insertions or deletions, point mutations, and desired affinity tags. The modifications can be combined and permuted into multiple protein constructs, and then virtually cloned in silico into defined expression vectors. The Gene Design Module uses a protein-to-gene algorithm that automates the back-translation of a protein amino acid sequence into a codon engineered nucleic acid gene sequence according to a selected codon usage table with minimal codon usage threshold, defined G:C% content, and desired sequence features achieved through synonymous codon selection that is optimized for the intended expression system. The gene-to-oligo algorithm of the Gene Design Module plans out all of the required overlapping oligonucleotides and mutagenic primers needed to synthesize the desired gene constructs by PCR, and for physically cloning them into selected vectors by the most popular subcloning strategies. We present a complete description of Gene Composer functionality, and an efficient PCR-based synthetic gene assembly procedure with mis-match specific endonuclease

Gene Composer: database software for protein construct design, codon engineering, and gene synthesis

Directory of Open Access Journals (Sweden)

Mixon Mark

2009-04-01

Full Text Available Abstract Background To improve efficiency in high throughput protein structure determination, we have developed a database software package, Gene Composer, which facilitates the information-rich design of protein constructs and their codon engineered synthetic gene sequences. With its modular workflow design and numerous graphical user interfaces, Gene Composer enables researchers to perform all common bio-informatics steps used in modern structure guided protein engineering and synthetic gene engineering. Results An interactive Alignment Viewer allows the researcher to simultaneously visualize sequence conservation in the context of known protein secondary structure, ligand contacts, water contacts, crystal contacts, B-factors, solvent accessible area, residue property type and several other useful property views. The Construct Design Module enables the facile design of novel protein constructs with altered N- and C-termini, internal insertions or deletions, point mutations, and desired affinity tags. The modifications can be combined and permuted into multiple protein constructs, and then virtually cloned in silico into defined expression vectors. The Gene Design Module uses a protein-to-gene algorithm that automates the back-translation of a protein amino acid sequence into a codon engineered nucleic acid gene sequence according to a selected codon usage table with minimal codon usage threshold, defined G:C% content, and desired sequence features achieved through synonymous codon selection that is optimized for the intended expression system. The gene-to-oligo algorithm of the Gene Design Module plans out all of the required overlapping oligonucleotides and mutagenic primers needed to synthesize the desired gene constructs by PCR, and for physically cloning them into selected vectors by the most popular subcloning strategies. Conclusion We present a complete description of Gene Composer functionality, and an efficient PCR-based synthetic gene

Engineering of soybean seed storage proteins

International Nuclear Information System (INIS)

Dickinson, C.D.; Floener, L.A.; Evans, R.P.; Nielsen, N.C.

1987-01-01

Protein engineering is one approach to the improvement of seed quality. With this in mind, a rapid in vitro system has been developed to assay the effect structural modifications have on the assembly of glycinin and β-conglycinin subunit complexes. Transcription plasmids were constructed for production of synthetic glycinin and β-conglycinin mRNAs by SP6 RNA-polymerase. Radiolabeled translation products from these messages were tested for their ability to form complexes. Gy4 and Gy5 proglycinins (group-2 subunits) and the a-subunit of β-conglycinin self-assembled into trimers. Proglycinin Gy2 (group-1 subunit) did not self-assemble, but assembled into mixed trimers in combination with Gy4 proglycinin. No assembly was observed for preproglycinins Gyl and Gy4, or for a Gy4 proglycinin which lacked 27 amino acids in a highly conserved internal sequence. Insertion of alternating MET-ARG residues in predicted turn regions of a hypervariable sequence in Gy4 proglycinin were tolerated when the string was short but inhibited trimer assembly as it became longer. The response to several different long deletions in this hypervariable region have also been tested. Different levels of trimer assembly were obtained and may depend on the secondary structures of the regions being joined in the engineered subunits. This system will be useful to study the assembly of storage protein complexes and to screen against modifications that interfere with subunit assembly

Cell Factory Engineering

DEFF Research Database (Denmark)

Davy, Anne Mathilde; Kildegaard, Helene Faustrup; Andersen, Mikael Rørdam

2017-01-01

focused on individual strategies or cell types, but collectively they fall under the broad umbrella of a growing field known as cell factory engineering. Here we condense >130 reviews and key studies in the art into a meta-review of cell factory engineering. We identified 33 generic strategies......-review provides general strategy guides for the broad range of applications of rational engineering of cell factories....

Nuclear techniques to determine microbial protein synthesis and productive performance of barki lambs fed rations containing some medicinal plants

International Nuclear Information System (INIS)

Mohamed, M.M.S.

2009-01-01

This study included two experiments, the first experiment was carried out in vitro to evaluate the effect of adding two levels of Lemongrass or Rosemary in ruminant rations on microbial protein synthesis using radio active sulfur S 35 . While, the second experiment was to study the effect of Lemongrass (CC) and Rosemary (RO) as feed additives in rations of lambs on feed intake, nutrient digestibility, some parameters of blood and rumen activity. Meanwhile, body weight and economical efficiency were studied. Twenty five of Barki male lambs with average body weight of 19.8 kg ± 1 kg and 3- 4 months of age were divided into 5 similar groups (5 lambs each). The first group (control) (R1) was fed on a concentrate feed mixture (CFM) plus rice straw (RS). While, R2 and R3 were fed as R1 ration supplemented with 100 or 200 mg Lemongrass /kg LBW/d respectively. Meantime, R4 and R5 were fed as R1 ration supplemented with 100 or 200 mg Rosemary /kg LBW/d respectively.The results indicated that more microbial protein synthesis was noticed with 4 mg of Lemongrass followed by 2 mg Rosemary, 2 mg Lemongrass and control which was higher than 4 mg Rosemary/ 0.5 g concentrate mixture. The differences were not statistically significant. The dry matter intake (DMI) was not significantly different for R4 and R3 when compared with R1 (control) and it significantly decreased in R5 and R2 compared with R1. The digestibilities of DM, OM, CP, EE and NFE in the supplemented groups were not significantly differing compared with R1. The digestibility of CF was significantly increased in R2 and R4 compared with R1 and there were no significant differences for R3 and R5 compared with R1. There were no significant differences in nutritive values as TDN, DCP and SV among all supplemented groups compared with R1. Rumen liquor TVFA,s was not significantly differ at zero time, but it decreased at 3 h and 6 h with all additives compared with the control with no significant differences among all

Rational synthetic pathway refactoring of natural products biosynthesis in actinobacteria.

Science.gov (United States)

Tan, Gao-Yi; Liu, Tiangang

2017-01-01

Natural products (NPs) and their derivatives are widely used as frontline treatments for many diseases. Actinobacteria spp. are used to produce most of NP antibiotics and have also been intensively investigated for NP production, derivatization, and discovery. However, due to the complicated transcriptional and metabolic regulation of NP biosynthesis in Actinobacteria, especially in the cases of genome mining and heterologous expression, it is often difficult to rationally and systematically engineer synthetic pathways to maximize biosynthetic efficiency. With the emergence of new tools and methods in metabolic engineering, the synthetic pathways of many chemicals, such as fatty acids and biofuels, in model organisms (e.g. Escherichia coli ), have been refactored to realize precise and flexible control of production. These studies also offer a promising approach for synthetic pathway refactoring in Actinobacteria. In this review, the great potential of Actinobacteria as a microbial cell factory for biosynthesis of NPs is discussed. To this end, recent progress in metabolic engineering of NP synthetic pathways in Actinobacteria are summarized and strategies and perspectives to rationally and systematically refactor synthetic pathways in Actinobacteria are highlighted. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Genome scale engineering techniques for metabolic engineering.

Science.gov (United States)

Liu, Rongming; Bassalo, Marcelo C; Zeitoun, Ramsey I; Gill, Ryan T

2015-11-01

Metabolic engineering has expanded from a focus on designs requiring a small number of genetic modifications to increasingly complex designs driven by advances in genome-scale engineering technologies. Metabolic engineering has been generally defined by the use of iterative cycles of rational genome modifications, strain analysis and characterization, and a synthesis step that fuels additional hypothesis generation. This cycle mirrors the Design-Build-Test-Learn cycle followed throughout various engineering fields that has recently become a defining aspect of synthetic biology. This review will attempt to summarize recent genome-scale design, build, test, and learn technologies and relate their use to a range of metabolic engineering applications. Copyright © 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

In-depth analysis of protein inference algorithms using multiple search engines and well-defined metrics.

Science.gov (United States)

Audain, Enrique; Uszkoreit, Julian; Sachsenberg, Timo; Pfeuffer, Julianus; Liang, Xiao; Hermjakob, Henning; Sanchez, Aniel; Eisenacher, Martin; Reinert, Knut; Tabb, David L; Kohlbacher, Oliver; Perez-Riverol, Yasset

2017-01-06

In mass spectrometry-based shotgun proteomics, protein identifications are usually the desired result. However, most of the analytical methods are based on the identification of reliable peptides and not the direct identification of intact proteins. Thus, assembling peptides identified from tandem mass spectra into a list of proteins, referred to as protein inference, is a critical step in proteomics research. Currently, different protein inference algorithms and tools are available for the proteomics community. Here, we evaluated five software tools for protein inference (PIA, ProteinProphet, Fido, ProteinLP, MSBayesPro) using three popular database search engines: Mascot, X!Tandem, and MS-GF+. All the algorithms were evaluated using a highly customizable KNIME workflow using four different public datasets with varying complexities (different sample preparation, species and analytical instruments). We defined a set of quality control metrics to evaluate the performance of each combination of search engines, protein inference algorithm, and parameters on each dataset. We show that the results for complex samples vary not only regarding the actual numbers of reported protein groups but also concerning the actual composition of groups. Furthermore, the robustness of reported proteins when using databases of differing complexities is strongly dependant on the applied inference algorithm. Finally, merging the identifications of multiple search engines does not necessarily increase the number of reported proteins, but does increase the number of peptides per protein and thus can generally be recommended. Protein inference is one of the major challenges in MS-based proteomics nowadays. Currently, there are a vast number of protein inference algorithms and implementations available for the proteomics community. Protein assembly impacts in the final results of the research, the quantitation values and the final claims in the research manuscript. Even though protein

Rationality in Society

NARCIS (Netherlands)

Flache, Andreas; Dijkstra, Jacob; Wright, James D.

2015-01-01

Contemporary theories of rational behavior in human society augment the orthodox model of rationality both by adding various forms of bounded rationality and relaxing the assumptions of self-interest and materialistic preferences. This entry discusses how these extensions of the theory of rational

Using Resurrected Ancestral Proviral Proteins to Engineer Virus Resistance

Directory of Open Access Journals (Sweden)

Asunción Delgado

2017-05-01

Full Text Available Proviral factors are host proteins hijacked by viruses for processes essential for virus propagation such as cellular entry and replication. Pathogens and their hosts co-evolve. It follows that replacing a proviral factor with a functional ancestral form of the same protein could prevent viral propagation without fatally compromising organismal fitness. Here, we provide proof of concept of this notion. Thioredoxins serve as general oxidoreductases in all known cells. We report that several laboratory resurrections of Precambrian thioredoxins display substantial levels of functionality within Escherichia coli. Unlike E. coli thioredoxin, however, these ancestral thioredoxins are not efficiently recruited by the bacteriophage T7 for its replisome and therefore prevent phage propagation in E. coli. These results suggest an approach to the engineering of virus resistance. Diseases caused by viruses may have a devastating effect in agriculture. We discuss how the suggested approach could be applied to the engineering of plant virus resistance.

Rational engineering of Geobacter sulfurreducens electron transfer components: a foundation for building improved Geobacter-based bioelectrochemical technologies

Directory of Open Access Journals (Sweden)

Joana M Dantas

2015-07-01

Full Text Available Multiheme cytochromes have been implicated in Geobacter sulfurreducens (Gs extracellular electron transfer (EET. These proteins are potential targets to improve EET and enhance bioremediation and electrical current production by Gs. However, the functional characterization of multiheme cytochromes is particularly complex due to the co-existence of several microstates in solution, connecting the fully reduced and fully oxidized states. Over the last decade, new strategies have been developed to characterize multiheme redox proteins functionally and structurally. These strategies were used to reveal the functional mechanism of Gs multiheme cytochromes and also to identify key residues in these proteins for EET. In previous studies, we set the foundations for enhancement of the EET abilities of Gs by characterizing a family of five triheme cytochromes (PpcA-E. These periplasmic cytochromes are implicated in electron transfer between the oxidative reactions of metabolism in the cytoplasm and the reduction of extracellular terminal electron acceptors at the cell’s outer surface. The results obtained suggested that PpcA can couple e-/H+ transfer, a property that might contribute to the proton electrochemical gradient across the cytoplasmic membrane for metabolic energy production. The structural and functional properties of PpcA were characterized in detail and used for rational design of a family of 23 single site PpcA mutants. In this review, we summarize the functional characterization of the native and mutant proteins. Mutants that retain the mechanistic features of PpcA and adopt preferential e-/H+ transfer pathways at lower reduction potential values compared to the wild-type protein were selected for in vivo studies as the best candidates to increase the electron transfer rate of Gs. For the first time Gs strains have been manipulated by the introduction of mutant forms of essential proteins with the aim to develop and improve

Preparation of scaffolds from human hair proteins for tissue-engineering applications

International Nuclear Information System (INIS)

Verma, Vipin; Verma, Poonam; Ray, Alok R; Ray, Pratima

2008-01-01

Human hair proteins were isolated and purified for the fabrication of tissue-engineering scaffolds. Their cellular compatibility was studied using NIH3T3 mice fibroblast cells. The proteins were characterized using FTIR spectroscopy, sodium dodecyl sulfate-polyacrylamide gel electrophoresis for molecular weights and two-dimensional polyacrylamide gel electrophoresis for their isoelectric points (pIs). The molecular weights of keratins were in the range of 40-60 kilo-Daltons (kDa) and of matrix proteins were in the range of 15-30 kDa. The pIs of keratins were found to be in the range of 4.5-5.3. Sponges of the proteins were formed by lyophilization. Scanning electron microscopy was performed to examine the surface. Swelling studies were carried out in phosphate buffer saline at physiological pH 7.4. The hydrophilic character of the protein surface was studied by determining an average contact angle, which came to be 37 0 . The wells of tissue culture plates were coated with these proteins for studying the attachment and morphology of the cells. The protein detachment study was done to ensure the adsorption of proteins on the wells until the completion of the experiments. The cellular growth on a protein-coated surface showed three-dimensional 'bulged' morphology due to cell-cell and cell-matrix contacts. The sponges of human hair proteins supported more cells for a longer period than control. The morphology and cell proliferation studies exhibited by NIH3T3 cells on these proteins have shown their potential to be used as tissue-engineering scaffolds with better cell-cell contacts and leucine-aspartic acid-valine (LDV)-mediated cell-matrix interactions

A Rational Approach to Rational Suicide.

Science.gov (United States)

Richman, Joseph

1992-01-01

Describes suicide as reaction to internal and external sources of stress and the impact of life events. Notes that, in the elderly, these situations are prevalent in many who are not suicidal. Contends that much more is written about rational suicide than its alternative (rational nonsuicide). Reviews reasons for this and suggests rational…

Fine-tuning of protein domain boundary by minimizing potential coiled coil regions

International Nuclear Information System (INIS)

Iwaya, Naoko; Goda, Natsuko; Unzai, Satoru; Fujiwara, Kenichiro; Tanaka, Toshiki; Tomii, Kentaro; Tochio, Hidehito; Shirakawa, Masahiro; Hiroaki, Hidekazu

2007-01-01

Structural determination of individual protein domains isolated from multidomain proteins is a common approach in the post-genomic era. Novel and thus uncharacterized domains liberated from intact proteins often self-associate due to incorrectly defined domain boundaries. Self-association results in missing signals, poor signal dispersion and a low signal-to-noise ratio in 1 H- 15 N HSQC spectra. We have found that a putative, non-canonical coiled coil region close to a domain boundary can cause transient hydrophobic self-association and monomer-dimer equilibrium in solution. Here we propose a rational method to predict putative coiled coil regions adjacent to the globular core domain using the program COILS. Except for the amino acid sequence, no preexisting knowledge concerning the domain is required. A small number of mutant proteins with a minimized coiled coil region have been rationally designed and tested. The engineered domains exhibit decreased self-association as assessed by 1 H- 15 N HSQC spectra with improved peak dispersion and sharper cross peaks. Two successful examples of isolating novel N-terminal domains from AAA-ATPases are demonstrated. Our method is useful for the experimental determination of domain boundaries suited for structural genomics studies

Fine-tuning of protein domain boundary by minimizing potential coiled coil regions.

Science.gov (United States)

Iwaya, Naoko; Goda, Natsuko; Unzai, Satoru; Fujiwara, Kenichiro; Tanaka, Toshiki; Tomii, Kentaro; Tochio, Hidehito; Shirakawa, Masahiro; Hiroaki, Hidekazu

2007-01-01

Structural determination of individual protein domains isolated from multidomain proteins is a common approach in the post-genomic era. Novel and thus uncharacterized domains liberated from intact proteins often self-associate due to incorrectly defined domain boundaries. Self-association results in missing signals, poor signal dispersion and a low signal-to-noise ratio in (1)H-(15)N HSQC spectra. We have found that a putative, non-canonical coiled coil region close to a domain boundary can cause transient hydrophobic self-association and monomer-dimer equilibrium in solution. Here we propose a rational method to predict putative coiled coil regions adjacent to the globular core domain using the program COILS. Except for the amino acid sequence, no preexisting knowledge concerning the domain is required. A small number of mutant proteins with a minimized coiled coil region have been rationally designed and tested. The engineered domains exhibit decreased self-association as assessed by (1)H-(15)N HSQC spectra with improved peak dispersion and sharper cross peaks. Two successful examples of isolating novel N-terminal domains from AAA-ATPases are demonstrated. Our method is useful for the experimental determination of domain boundaries suited for structural genomics studies.

Engineering Non-Heme Mono- and Dioxygenases for Biocatalysis

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Adi Dror

2012-09-01

Full Text Available Oxygenases are ubiquitous enzymes that catalyze the introduction of one or two oxygen atoms to unreactive chemical compounds. They require reduction equivalents from NADH or NADPH and comprise metal ions, metal ion complexes, or coenzymes in their active site. Thus, for industrial purposes, oxygenases are most commonly employed using whole cell catalysis, to alleviate the need for co-factor regeneration. Biotechnological applications include bioremediation, chiral synthesis, biosensors, fine chemicals, biofuels, pharmaceuticals, food ingredients and polymers. Controlling activity and selectivity of oxygenases is therefore of great importance and of growing interest to the scientific community. This review focuses on protein engineering of non-heme monooxygenases and dioxygenases for generating improved or novel functionalities. Rational mutagenesis based on x-ray structures and sequence alignment, as well as random methods such as directed evolution, have been utilized. It is concluded that knowledge-based protein engineering accompanied with targeted libraries, is most efficient for the design and tuning of biocatalysts towards novel substrates and enhanced catalytic activity while minimizing the screening efforts.

Molecular and macro-scale analysis of enzyme-crosslinked silk hydrogels for rational biomaterial design.

Science.gov (United States)

McGill, Meghan; Coburn, Jeannine M; Partlow, Benjamin P; Mu, Xuan; Kaplan, David L

2017-11-01

Silk fibroin-based hydrogels have exciting applications in tissue engineering and therapeutic molecule delivery; however, their utility is dependent on their diffusive properties. The present study describes a molecular and macro-scale investigation of enzymatically-crosslinked silk fibroin hydrogels, and demonstrates that these systems have tunable crosslink density and diffusivity. We developed a liquid chromatography tandem mass spectroscopy (LC-MS/MS) method to assess the quantity and order of covalent tyrosine crosslinks in the hydrogels. This analysis revealed between 28 and 56% conversion of tyrosine to dityrosine, which was dependent on the silk concentration and reactant concentration. The crosslink density was then correlated with storage modulus, revealing that both crosslinking and protein concentration influenced the mechanical properties of the hydrogels. The diffusive properties of the bulk material were studied by fluorescence recovery after photobleaching (FRAP), which revealed a non-linear relationship between silk concentration and diffusivity. As a result of this work, a model for synthesizing hydrogels with known crosslink densities and diffusive properties has been established, enabling the rational design of silk hydrogels for biomedical applications. Hydrogels from naturally-derived silk polymers offer versitile opportunities in the biomedical field, however, their design has largely been an empirical process. We present a fundamental study of the crosslink density, storage modulus, and diffusion behavior of enzymatically-crosslinked silk hydrogels to better inform scaffold design. These studies revealed unexpected non-linear trends in the crosslink density and diffusivity of silk hydrogels with respect to protein concentration and crosslink reagent concentration. This work demonstrates the tunable diffusivity and crosslinking in silk fibroin hydrogels, and enables the rational design of biomaterials. Further, the characterization methods

The influence of ration size on energetics and nitrogen retention in tilapia (Oreochromis niloticus)

DEFF Research Database (Denmark)

Skov, Peter Vilhelm; Duodu, Collins Prah; Adjei-Boateng, Daniel

2017-01-01

with ration size, feed conversion and protein retention were most efficient at ration sizes of 3%. Although the magnitude of the SDA response following feeding also increased with ration size, this was not proportionate to meal size. Therefore the metabolic cost of meal processing (SDA coefficient) was found......Proper nutrient management is essential for the environmental sustainability of aquaculture. While increasing daily rations generally may lead to improved growth rates, this does not necessarily mean that nutrients are utilized more efficiently. To investigate how ration size affects partitioning...... of dietary nutrient intake, the effects of meal size on growth and metabolism were examined in triplicate groups of adult Nile tilapia (Oreochromis niloticus) receiving daily rations corresponding to 1, 2, 3, or 4% of their biomass. While biomass gain and specific growth rates were positively correlated...

Ability of commercially available dairy ration programs to predict duodenal flows of protein and essential amino acids in dairy cows.

Science.gov (United States)

Pacheco, D; Patton, R A; Parys, C; Lapierre, H

2012-02-01

The objective of this analysis was to compare the rumen submodel predictions of 4 commonly used dairy ration programs to observed values of duodenal flows of crude protein (CP), protein fractions, and essential AA (EAA). The literature was searched and 40 studies, including 154 diets, were used to compare observed values with those predicted by AminoCow (AC), Agricultural Modeling and Training Systems (AMTS), Cornell-Penn-Miner (CPM), and National Research Council 2001 (NRC) models. The models were evaluated based on their ability to predict the mean, their root mean square prediction error (RMSPE), error bias, and adequacy of regression equations for each protein fraction. The models predicted the mean duodenal CP flow within 5%, with more than 90% of the variation due to random disturbance. The models also predicted within 5% the mean microbial CP flow except CPM, which overestimated it by 27%. Only NRC, however, predicted mean rumen-undegraded protein (RUP) flows within 5%, whereas AC and AMTS underpredicted it by 8 to 9% and CPM by 24%. Regarding duodenal flows of individual AA, across all diets, CPM predicted substantially greater (>10%) mean flows of Arg, His, Ile, Met, and Lys; AMTS predicted greater flow for Arg and Met, whereas AC and NRC estimations were, on average, within 10% of observed values. Overpredictions by the CPM model were mainly related to mean bias, whereas the NRC model had the highest proportion of bias in random disturbance for flows of EAA. Models tended to predict mean flows of EAA more accurately on corn silage and alfalfa diets than on grass-based diets, more accurately on corn grain-based diets than on non-corn-based diets, and finally more accurately in the mid range of diet types. The 4 models were accurate at predicting mean dry matter intake. The AC, AMTS, and NRC models were all sufficiently accurate to be used for balancing EAA in dairy rations under field conditions. Copyright © 2012 American Dairy Science Association

Cytocompatible and water stable ultrafine protein fibers for tissue engineering

Science.gov (United States)

Jiang, Qiuran

This dissertation proposal focuses on the development of cytocompatible and water stable protein ultrafine fibers for tissue engineering. The protein-based ultrafine fibers have the potential to be used for biomedicine, due to their biocompatibility, biodegradability, similarity to natural extracellular matrix (ECM) in physical structure and chemical composition, and superior adsorption properties due to their high surface to volume ratio. However, the current technologies to produce the protein-based ultrafine fibers for biomedical applications still have several problems. For instance, the current electrospinning and phase separation technologies generate scaffolds composed of densely compacted ultrafine fibers, and cells can spread just on the surface of the fiber bulk, and hardly penetrate into the inner sections of scaffolds. Thus, these scaffolds can merely emulate the ECM as a two dimensional basement membrane, but are difficult to mimic the three dimensional ECM stroma. Moreover, the protein-based ultrafine fibers do not possess sufficient water stability and strength for biomedical applications, and need modifications such as crosslinking. However, current crosslinking methods are either high in toxicity or low in crosslinking efficiency. To solve the problems mentioned above, zein, collagen, and gelatin were selected as the raw materials to represent plant proteins, animal proteins, and denatured proteins in this dissertation. A benign solvent system was developed specifically for the fabrication of collagen ultrafine fibers. In addition, the gelatin scaffolds with a loose fibrous structure, high cell-accessibility and cell viability were produced by a novel ultralow concentration phase separation method aiming to simulate the structure of three dimensional (3D) ECM stroma. Non-toxic crosslinking methods using citric acid as the crosslinker were also developed for electrospun or phase separated scaffolds from these three proteins, and proved to be

Optimization of fodder rations for intensive development of cattle-breeding in an radioactive contaminated zone

International Nuclear Information System (INIS)

Stolyarov, G.V.

1999-01-01

It has been calculated some variants of the optimal structure of milk cow herd's fodder rations in a radioactive contaminated zone in dependence of the contamination density. Rations were balanced in primary nutritive including digestible protein. It has been determined their costs and specific radioactivity of cesium-137. These fodder rations can be recommended to the farms of the Gomel Region suffered from the Chernobyl nuclear power station explosion

Engineering of the E. coli Outer Membrane Protein FhuA to overcome the Hydrophobic Mismatch in Thick Polymeric Membranes

Directory of Open Access Journals (Sweden)

Fioroni Marco

2011-03-01

Full Text Available Abstract Background Channel proteins like the engineered FhuA Δ1-159 often cannot insert into thick polymeric membranes due to a mismatch between the hydrophobic surface of the protein and the hydrophobic surface of the polymer membrane. To address this problem usually specific block copolymers are synthesized to facilitate protein insertion. Within this study in a reverse approach we match the protein to the polymer instead of matching the polymer to the protein. Results To increase the FhuA Δ1-159 hydrophobic surface by 1 nm, the last 5 amino acids of each of the 22 β-sheets, prior to the more regular periplasmatic β-turns, were doubled leading to an extended FhuA Δ1-159 (FhuA Δ1-159 Ext. The secondary structure prediction and CD spectroscopy indicate the β-barrel folding of FhuA Δ1-159 Ext. The FhuA Δ1-159 Ext insertion and functionality within a nanocontainer polymeric membrane based on the triblock copolymer PIB1000-PEG6000-PIB1000 (PIB = polyisobutylene, PEG = polyethyleneglycol has been proven by kinetic analysis using the HRP-TMB assay (HRP = Horse Radish Peroxidase, TMB = 3,3',5,5'-tetramethylbenzidine. Identical experiments with the unmodified FhuA Δ1-159 report no kinetics and presumably no insertion into the PIB1000-PEG6000-PIB1000 membrane. Furthermore labeling of the Lys-NH2 groups present in the FhuA Δ1-159 Ext channel, leads to controllability of in/out flux of substrates and products from the nanocontainer. Conclusion Using a simple "semi rational" approach the protein's hydrophobic transmembrane region was increased by 1 nm, leading to a predicted lower hydrophobic mismatch between the protein and polymer membrane, minimizing the insertion energy penalty. The strategy of adding amino acids to the FhuA Δ1-159 Ext hydrophobic part can be further expanded to increase the protein's hydrophobicity, promoting the efficient embedding into thicker/more hydrophobic block copolymer membranes.

Engineering of the E. coli outer membrane protein FhuA to overcome the hydrophobic mismatch in thick polymeric membranes.

Science.gov (United States)

Muhammad, Noor; Dworeck, Tamara; Fioroni, Marco; Schwaneberg, Ulrich

2011-03-17

Channel proteins like the engineered FhuA Δ1-159 often cannot insert into thick polymeric membranes due to a mismatch between the hydrophobic surface of the protein and the hydrophobic surface of the polymer membrane. To address this problem usually specific block copolymers are synthesized to facilitate protein insertion. Within this study in a reverse approach we match the protein to the polymer instead of matching the polymer to the protein. To increase the FhuA Δ1-159 hydrophobic surface by 1 nm, the last 5 amino acids of each of the 22 β-sheets, prior to the more regular periplasmatic β-turns, were doubled leading to an extended FhuA Δ1-159 (FhuA Δ1-159 Ext). The secondary structure prediction and CD spectroscopy indicate the β-barrel folding of FhuA Δ1-159 Ext. The FhuA Δ1-159 Ext insertion and functionality within a nanocontainer polymeric membrane based on the triblock copolymer PIB(1000)-PEG(6000)-PIB(1000) (PIB = polyisobutylene, PEG = polyethyleneglycol) has been proven by kinetic analysis using the HRP-TMB assay (HRP = Horse Radish Peroxidase, TMB = 3,3',5,5'-tetramethylbenzidine). Identical experiments with the unmodified FhuA Δ1-159 report no kinetics and presumably no insertion into the PIB(1000)-PEG(6000)-PIB(1000) membrane. Furthermore labeling of the Lys-NH(2) groups present in the FhuA Δ1-159 Ext channel, leads to controllability of in/out flux of substrates and products from the nanocontainer. Using a simple "semi rational" approach the protein's hydrophobic transmembrane region was increased by 1 nm, leading to a predicted lower hydrophobic mismatch between the protein and polymer membrane, minimizing the insertion energy penalty. The strategy of adding amino acids to the FhuA Δ1-159 Ext hydrophobic part can be further expanded to increase the protein's hydrophobicity, promoting the efficient embedding into thicker/more hydrophobic block copolymer membranes.

Structure-based, rational design of T cell receptors

Directory of Open Access Journals (Sweden)

Vincent eZoete

2013-09-01

Full Text Available Adoptive cell transfer using engineered T cells is emerging as a promising treatment for metastatic melanoma. Such an approach allows one to introduce TCR modifications that, while maintaining the specificity for the targeted antigen, can enhance the binding and kinetic parameters for the interaction pMHC. Using the well-characterized 2C TCR/SIYR/H-2K(b structure as a model system, we demonstrated that a binding free energy decomposition based on the MM-GBSA approach provides a detailed and reliable description of the TCR/pMHC interactions at the structural and thermodynamic levels. Starting from this result, we developed a new structure-based approach, to rationally design new TCR sequences, and applied it to the BC1 TCR targeting the HLA-A2 restricted NY-ESO-1157-165 cancer-testis epitope. 54% of the designed sequence replacements exhibited improved pMHC-binding as compared to the native TCR, with up to 150 fold increase in affinity, while preserving specificity. Genetically-engineered CD8+ T cells expressing these modified TCRs showed an improved functional activity compared to those expressing BC1 TCR. We measured maximum levels of activities for TCRs within the upper limit of natural affinity. Beyond the affinity threshold at KD < 1 μM we observed an attenuation in cellular function. We have also developed a homology modeling-based approach, TCRep 3D, to obtain accurate structural models of any TCR-pMHC complexes. We have complemented the approach with a simplified rigid method to predict the TCR orientation over pMHC. These methods potentially extend the use of our TCR engineering method to entire TCR repertoires for which no X-ray structure is available. We have also performed a steered molecular dynamics study of the unbinding of the TCR-pMHC complex to get a better understanding of how TCRs interact with pMHCs. This entire rational TCR design pipeline is now being used to produce rationally optimized TCRs for adoptive cell therapies of

Rational F-theory GUTs without exotics

International Nuclear Information System (INIS)

Krippendorf, Sven; Peña, Damián Kaloni Mayorga; Oehlmann, Paul-Konstantin; Ruehle, Fabian

2014-01-01

We construct F-theory GUT models without exotic matter, leading to the MSSM matter spectrum with potential singlet extensions. The interplay of engineering explicit geometric setups, absence of four-dimensional anomalies, and realistic phenomenology of the couplings places severe constraints on the allowed local models in a given geometry. In constructions based on the spectral cover we find no model satisfying all these requirements. We then provide a survey of models with additional U1 symmetries arising from rational sections of the elliptic fibration in toric constructions and obtain phenomenologically appealing models based on SU(5) tops. Furthermore we perform a bottom-up exploration beyond the toric section constructions discussed in the literature so far and identify benchmark models passing all our criteria, which can serve as a guideline for future geometric engineering.

Rational F-theory GUTs without exotics

Science.gov (United States)

Krippendorf, Sven; Peña, Damián Kaloni Mayorga; Oehlmann, Paul-Konstantin; Ruehle, Fabian

2014-07-01

We construct F-theory GUT models without exotic matter, leading to the MSSM matter spectrum with potential singlet extensions. The interplay of engineering explicit geometric setups, absence of four-dimensional anomalies, and realistic phenomenology of the couplings places severe constraints on the allowed local models in a given geometry. In constructions based on the spectral cover we find no model satisfying all these requirements. We then provide a survey of models with additional U(1) symmetries arising from rational sections of the elliptic fibration in toric constructions and obtain phenomenologically appealing models based on SU(5) tops. Furthermore we perform a bottom-up exploration beyond the toric section constructions discussed in the literature so far and identify benchmark models passing all our criteria, which can serve as a guideline for future geometric engineering.

Rational F-theory GUTs without exotics

International Nuclear Information System (INIS)

Krippendorf, Sven; Pena, Damian Kaloni Mayorga; Oehlmann, Paul-Konstantin

2014-01-01

We construct F-theory GUT models without exotic matter, leading to the MSSM matter spectrum with potential singlet extensions. The interplay of engineering explicit geometric setups, absence of four-dimensional anomalies, and realistic phenomenology of the couplings places severe constraints on the allowed local models in a given geometry. In constructions based on the spectral cover we find no model satisfying all these requirements. We then provide a survey of models with additional U(1) symmetries arising from rational sections of the elliptic fibration in toric constructions and obtain phenomenologically appealing models based on SU(5) tops. Furthermore we perform a bottom-up exploration beyond the toric section constructions discussed in the literature so far and identify benchmark models passing all our criteria, which can serve as a guideline for future geometric engineering.

Rational Molecular Engineering of Indoline-Based D-A-π-A Organic Sensitizers for Long-Wavelength-Responsive Dye-Sensitized Solar Cells.

Science.gov (United States)

Zhang, Weiwei; Wu, Yongzhen; Zhu, Haibo; Chai, Qipeng; Liu, Jingchuan; Li, Hui; Song, Xiongrong; Zhu, Wei-Hong

2015-12-09

Indoline-based D-A-π-A organic sensitizers are promising candidates for highly efficient and long-term stable dye-sensitized solar cells (DSSCs). In order to further broaden the spectral response of the known indoline dye WS-2, we rationally engineer the molecular structure through enhancing the electron donor and extending the π-bridge, resulting in two novel indoline-based D-A-π-A organic sensitizers WS-92 and WS-95. By replacing the 4-methylphenyl group on the indoline donor of WS-2 with a more electron-rich carbazole unit, the intramolecular charge transfer (ICT) absorption band of dye WS-92 is slightly red-shifted from 550 nm (WS-2) to 554 nm (WS-92). In comparison, the incorporation of a larger π-bridge of cyclopentadithiophene (CPDT) unit in dye WS-95 not only greatly bathochromatically tunes the absorption band to 574 nm but also largely enhances the molar extinction coefficients (ε), thus dramatically improving the light-harvesting capability. Under the standard global AM 1.5 solar light condition, the photovoltaic performances of both organic dyes have been evaluated in DSSCs on the basis of the iodide/triiodide electrolyte without any coadsorbent or cosensitizer. The DSSCs based on WS-95 display better device performance with power conversion efficiency (η) of 7.69%. The additional coadsorbent in the dye bath of WS-95 does not improve the photovoltaic performance, indicative of its negligible dye aggregation, which can be rationalized by the grafted dioctyl chains on the CPDT unit. The cosensitization of WS-95 with a short absorption wavelength dye S2 enhances the IPCE and improves the η to 9.18%. Our results indicate that extending the π-spacer is more rational than enhancing the electron donor in terms of broadening the spectral response of indoline-based D-A-π-A organic sensitizers.

On rationally supported surfaces

DEFF Research Database (Denmark)

Gravesen, Jens; Juttler, B.; Sir, Z.

2008-01-01

We analyze the class of surfaces which are equipped with rational support functions. Any rational support function can be decomposed into a symmetric (even) and an antisymmetric (odd) part. We analyze certain geometric properties of surfaces with odd and even rational support functions....... In particular it is shown that odd rational support functions correspond to those rational surfaces which can be equipped with a linear field of normal vectors, which were discussed by Sampoli et al. (Sampoli, M.L., Peternell, M., Juttler, B., 2006. Rational surfaces with linear normals and their convolutions...... with rational surfaces. Comput. Aided Geom. Design 23, 179-192). As shown recently, this class of surfaces includes non-developable quadratic triangular Bezier surface patches (Lavicka, M., Bastl, B., 2007. Rational hypersurfaces with rational convolutions. Comput. Aided Geom. Design 24, 410426; Peternell, M...

Rational design and application of responsive α-helical peptide hydrogels

Science.gov (United States)

Banwell, Eleanor F.; Abelardo, Edgardo S.; Adams, Dave J.; Birchall, Martin A.; Corrigan, Adam; Donald, Athene M.; Kirkland, Mark; Serpell, Louise C.; Butler, Michael F.; Woolfson, Derek N.

2009-01-01

Biocompatible hydrogels have a wide variety of potential applications in biotechnology and medicine, such as the controlled delivery and release of cells, cosmetics and drugs; and as supports for cell growth and tissue engineering1. Rational peptide design and engineering are emerging as promising new routes to such functional biomaterials2-4. Here we present the first examples of rationally designed and fully characterized self-assembling hydrogels based on standard linear peptides with purely α-helical structures, which we call hydrogelating self-assembling fibres (hSAFs). These form spanning networks of α-helical fibrils that interact to give self-supporting physical hydrogels of >99% water content. The peptide sequences can be engineered to alter the underlying mechanism of gelation and, consequently, the hydrogel properties. Interestingly, for example, those with hydrogen-bonded networks melt upon heating, whereas those formed via hydrophobic interactions strengthen when warmed. The hSAFs are dual-peptide systems that only gel on mixing, which gives tight control over assembly5. These properties raise possibilities for using the hSAFs as substrates in cell culture. We have tested this in comparison with the widely used Matrigel substrate, and demonstrate that, like Matrigel, hSAFs support both growth and differentiation of rat adrenal pheochromocytoma cells for sustained periods in culture. PMID:19543314

Roles of beta-turns in protein folding: from peptide models to protein engineering.

Science.gov (United States)

Marcelino, Anna Marie C; Gierasch, Lila M

2008-05-01

Reverse turns are a major class of protein secondary structure; they represent sites of chain reversal and thus sites where the globular character of a protein is created. It has been speculated for many years that turns may nucleate the formation of structure in protein folding, as their propensity to occur will favor the approximation of their flanking regions and their general tendency to be hydrophilic will favor their disposition at the solvent-accessible surface. Reverse turns are local features, and it is therefore not surprising that their structural properties have been extensively studied using peptide models. In this article, we review research on peptide models of turns to test the hypothesis that the propensities of turns to form in short peptides will relate to the roles of corresponding sequences in protein folding. Turns with significant stability as isolated entities should actively promote the folding of a protein, and by contrast, turn sequences that merely allow the chain to adopt conformations required for chain reversal are predicted to be passive in the folding mechanism. We discuss results of protein engineering studies of the roles of turn residues in folding mechanisms. Factors that correlate with the importance of turns in folding indeed include their intrinsic stability, as well as their topological context and their participation in hydrophobic networks within the protein's structure.

Using Resurrected Ancestral Proviral Proteins to Engineer Virus Resistance.

Science.gov (United States)

Delgado, Asunción; Arco, Rocio; Ibarra-Molero, Beatriz; Sanchez-Ruiz, Jose M

2017-05-09

Proviral factors are host proteins hijacked by viruses for processes essential for virus propagation such as cellular entry and replication. Pathogens and their hosts co-evolve. It follows that replacing a proviral factor with a functional ancestral form of the same protein could prevent viral propagation without fatally compromising organismal fitness. Here, we provide proof of concept of this notion. Thioredoxins serve as general oxidoreductases in all known cells. We report that several laboratory resurrections of Precambrian thioredoxins display substantial levels of functionality within Escherichia coli. Unlike E. coli thioredoxin, however, these ancestral thioredoxins are not efficiently recruited by the bacteriophage T7 for its replisome and therefore prevent phage propagation in E. coli. These results suggest an approach to the engineering of virus resistance. Diseases caused by viruses may have a devastating effect in agriculture. We discuss how the suggested approach could be applied to the engineering of plant virus resistance. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Supramolecular Architectures and Mimics of Complex Natural Folds Derived from Rationally Designed alpha-Helical Protein Structures

Science.gov (United States)

Tavenor, Nathan Albert

Protein-based supramolecular polymers (SMPs) are a class of biomaterials which draw inspiration from and expand upon the many examples of complex protein quaternary structures observed in nature: collagen, microtubules, viral capsids, etc. Designing synthetic supramolecular protein scaffolds both increases our understanding of natural superstructures and allows for the creation of novel materials. Similar to small-molecule SMPs, protein-based SMPs form due to self-assembly driven by intermolecular interactions between monomers, and monomer structure determines the properties of the overall material. Using protein-based monomers takes advantage of the self-assembly and highly specific molecular recognition properties encodable in polypeptide sequences to rationally design SMP architectures. The central hypothesis underlying our work is that alpha-helical coiled coils, a well-studied protein quaternary folding motif, are well-suited to SMP design through the addition of synthetic linkers at solvent-exposed sites. Through small changes in the structures of the cross-links and/or peptide sequence, we have been able to control both the nanoscale organization and the macroscopic properties of the SMPs. Changes to the linker and hydrophobic core of the peptide can be used to control polymer rigidity, stability, and dimensionality. The gaps in knowledge that this thesis sought to fill on this project were 1) the relationship between the molecular structure of the cross-linked polypeptides and the macroscopic properties of the SMPs and 2) a means of creating materials exhibiting multi-dimensional net or framework topologies. Separate from the above efforts on supramolecular architectures was work on improving backbone modification strategies for an alpha-helix in the context of a complex protein tertiary fold. Earlier work in our lab had successfully incorporated unnatural building blocks into every major secondary structure (beta-sheet, alpha-helix, loops and beta

Pragmatics & rationality.

OpenAIRE

Allott, N. E.

2007-01-01

This thesis is about the reconciliation of realistic views of rationality with inferential-intentional theories of communication. Grice (1957 1975) argued that working out what a speaker meant by an utterance is a matter of inferring the speaker's intentions on the presumption that she is acting rationally. This is abductive inference: inference to the best explanation for the utterance. Thus an utterance both rationalises and causes the interpretation the hearer constructs. Human rationality...

From Protein Engineering to Immobilization: Promising Strategies for the Upgrade of Industrial Enzymes

Science.gov (United States)

Singh, Raushan Kumar; Tiwari, Manish Kumar; Singh, Ranjitha; Lee, Jung-Kul

2013-01-01

Enzymes found in nature have been exploited in industry due to their inherent catalytic properties in complex chemical processes under mild experimental and environmental conditions. The desired industrial goal is often difficult to achieve using the native form of the enzyme. Recent developments in protein engineering have revolutionized the development of commercially available enzymes into better industrial catalysts. Protein engineering aims at modifying the sequence of a protein, and hence its structure, to create enzymes with improved functional properties such as stability, specific activity, inhibition by reaction products, and selectivity towards non-natural substrates. Soluble enzymes are often immobilized onto solid insoluble supports to be reused in continuous processes and to facilitate the economical recovery of the enzyme after the reaction without any significant loss to its biochemical properties. Immobilization confers considerable stability towards temperature variations and organic solvents. Multipoint and multisubunit covalent attachments of enzymes on appropriately functionalized supports via linkers provide rigidity to the immobilized enzyme structure, ultimately resulting in improved enzyme stability. Protein engineering and immobilization techniques are sequential and compatible approaches for the improvement of enzyme properties. The present review highlights and summarizes various studies that have aimed to improve the biochemical properties of industrially significant enzymes. PMID:23306150

From protein engineering to immobilization: promising strategies for the upgrade of industrial enzymes.

Science.gov (United States)

Singh, Raushan Kumar; Tiwari, Manish Kumar; Singh, Ranjitha; Lee, Jung-Kul

2013-01-10

Enzymes found in nature have been exploited in industry due to their inherent catalytic properties in complex chemical processes under mild experimental and environmental conditions. The desired industrial goal is often difficult to achieve using the native form of the enzyme. Recent developments in protein engineering have revolutionized the development of commercially available enzymes into better industrial catalysts. Protein engineering aims at modifying the sequence of a protein, and hence its structure, to create enzymes with improved functional properties such as stability, specific activity, inhibition by reaction products, and selectivity towards non-natural substrates. Soluble enzymes are often immobilized onto solid insoluble supports to be reused in continuous processes and to facilitate the economical recovery of the enzyme after the reaction without any significant loss to its biochemical properties. Immobilization confers considerable stability towards temperature variations and organic solvents. Multipoint and multisubunit covalent attachments of enzymes on appropriately functionalized supports via linkers provide rigidity to the immobilized enzyme structure, ultimately resulting in improved enzyme stability. Protein engineering and immobilization techniques are sequential and compatible approaches for the improvement of enzyme properties. The present review highlights and summarizes various studies that have aimed to improve the biochemical properties of industrially significant enzymes.

Sequence-engineered mRNA Without Chemical Nucleoside Modifications Enables an Effective Protein Therapy in Large Animals

Science.gov (United States)

Thess, Andreas; Grund, Stefanie; Mui, Barbara L; Hope, Michael J; Baumhof, Patrick; Fotin-Mleczek, Mariola; Schlake, Thomas

2015-01-01

Being a transient carrier of genetic information, mRNA could be a versatile, flexible, and safe means for protein therapies. While recent findings highlight the enormous therapeutic potential of mRNA, evidence that mRNA-based protein therapies are feasible beyond small animals such as mice is still lacking. Previous studies imply that mRNA therapeutics require chemical nucleoside modifications to obtain sufficient protein expression and avoid activation of the innate immune system. Here we show that chemically unmodified mRNA can achieve those goals as well by applying sequence-engineered molecules. Using erythropoietin (EPO) driven production of red blood cells as the biological model, engineered Epo mRNA elicited meaningful physiological responses from mice to nonhuman primates. Even in pigs of about 20 kg in weight, a single adequate dose of engineered mRNA encapsulated in lipid nanoparticles (LNPs) induced high systemic Epo levels and strong physiological effects. Our results demonstrate that sequence-engineered mRNA has the potential to revolutionize human protein therapies. PMID:26050989

Baseline rationing

DEFF Research Database (Denmark)

Hougaard, Jens Leth; Moreno-Ternero, Juan D.; Østerdal, Lars Peter Raahave

The standard problem of adjudicating conflicting claims describes a situation in which a given amount of a divisible good has to be allocated among agents who hold claims against it exceeding the available amount. This paper considers more general rationing problems in which, in addition to claims...... to international protocols for the reduction of greenhouse emissions, or water distribution in drought periods. We define a family of allocation methods for such general rationing problems - called baseline rationing rules - and provide an axiomatic characterization for it. Any baseline rationing rule within...... the family is associated with a standard rule and we show that if the latter obeys some properties reflecting principles of impartiality, priority and solidarity, the former obeys them too....

Rationing with baselines

DEFF Research Database (Denmark)

Hougaard, Jens Leth; Moreno-Ternero, Juan D.; Østerdal, Lars Peter Raahave

2013-01-01

We introduce a new operator for general rationing problems in which, besides conflicting claims, individual baselines play an important role in the rationing process. The operator builds onto ideas of composition, which are not only frequent in rationing, but also in related problems...... such as bargaining, choice, and queuing. We characterize the operator and show how it preserves some standard axioms in the literature on rationing. We also relate it to recent contributions in such literature....

Inaugurating Rationalization: Three Field Studies Find Increased Rationalization When Anticipated Realities Become Current.

Science.gov (United States)

Laurin, Kristin

2018-04-01

People will often rationalize the status quo, reconstruing it in an exaggeratedly positive light. They will even rationalize the status quo they anticipate, emphasizing the upsides and minimizing the downsides of sociopolitical realities they expect to take effect. Drawing on recent findings on the psychological triggers of rationalization, I present results from three field studies, one of which was preregistered, testing the hypothesis that an anticipated reality becoming current triggers an observable boost in people's rationalizations. San Franciscans rationalized a ban on plastic water bottles, Ontarians rationalized a targeted smoking ban, and Americans rationalized the presidency of Donald Trump, more in the days immediately after these realities became current compared with the days immediately before. Additional findings show evidence for a mechanism underlying these behaviors and rule out alternative accounts. These findings carry implications for scholarship on rationalization, for understanding protest behavior, and for policymakers.

Rapid Sampling of Hydrogen Bond Networks for Computational Protein Design.

Science.gov (United States)

Maguire, Jack B; Boyken, Scott E; Baker, David; Kuhlman, Brian

2018-05-08

Hydrogen bond networks play a critical role in determining the stability and specificity of biomolecular complexes, and the ability to design such networks is important for engineering novel structures, interactions, and enzymes. One key feature of hydrogen bond networks that makes them difficult to rationally engineer is that they are highly cooperative and are not energetically favorable until the hydrogen bonding potential has been satisfied for all buried polar groups in the network. Existing computational methods for protein design are ill-equipped for creating these highly cooperative networks because they rely on energy functions and sampling strategies that are focused on pairwise interactions. To enable the design of complex hydrogen bond networks, we have developed a new sampling protocol in the molecular modeling program Rosetta that explicitly searches for sets of amino acid mutations that can form self-contained hydrogen bond networks. For a given set of designable residues, the protocol often identifies many alternative sets of mutations/networks, and we show that it can readily be applied to large sets of residues at protein-protein interfaces or in the interior of proteins. The protocol builds on a recently developed method in Rosetta for designing hydrogen bond networks that has been experimentally validated for small symmetric systems but was not extensible to many larger protein structures and complexes. The sampling protocol we describe here not only recapitulates previously validated designs with performance improvements but also yields viable hydrogen bond networks for cases where the previous method fails, such as the design of large, asymmetric interfaces relevant to engineering protein-based therapeutics.

Crystallization and preliminary X-ray characterization of the genetically encoded fluorescent calcium indicator protein GCaMP2

International Nuclear Information System (INIS)

Rodríguez Guilbe, María M.; Alfaro Malavé, Elisa C.; Akerboom, Jasper; Marvin, Jonathan S.; Looger, Loren L.; Schreiter, Eric R.

2008-01-01

The genetically encoded fluorescent calcium-indicator protein GCaMP2 was crystallized in the calcium-saturated form. X-ray diffraction data were collected to 2.0 Å resolution and the structure was solved by molecular replacement. Fluorescent proteins and their engineered variants have played an important role in the study of biology. The genetically encoded calcium-indicator protein GCaMP2 comprises a circularly permuted fluorescent protein coupled to the calcium-binding protein calmodulin and a calmodulin target peptide, M13, derived from the intracellular calmodulin target myosin light-chain kinase and has been used to image calcium transients in vivo. To aid rational efforts to engineer improved variants of GCaMP2, this protein was crystallized in the calcium-saturated form. X-ray diffraction data were collected to 2.0 Å resolution. The crystals belong to space group C2, with unit-cell parameters a = 126.1, b = 47.1, c = 68.8 Å, β = 100.5° and one GCaMP2 molecule in the asymmetric unit. The structure was phased by molecular replacement and refinement is currently under way

Realization theory for rational systems: Minimal rational realizations

NARCIS (Netherlands)

J. Nemcová (Jana); J.H. van Schuppen (Jan)

2010-01-01

htmlabstractThe study of realizations of response maps is a topic of control and system theory. Realization theory is used in system identification and control synthesis. A minimal rational realization of a given response map p is a rational realization of p such that the dimension of its state

Many faces of rationality: Implications of the great rationality debate for clinical decision‐making

Science.gov (United States)

Elqayam, Shira

2017-01-01

Abstract Given that more than 30% of healthcare costs are wasted on inappropriate care, suboptimal care is increasingly connected to the quality of medical decisions. It has been argued that personal decisions are the leading cause of death, and 80% of healthcare expenditures result from physicians' decisions. Therefore, improving healthcare necessitates improving medical decisions, ie, making decisions (more) rational. Drawing on writings from The Great Rationality Debate from the fields of philosophy, economics, and psychology, we identify core ingredients of rationality commonly encountered across various theoretical models. Rationality is typically classified under umbrella of normative (addressing the question how people “should” or “ought to” make their decisions) and descriptive theories of decision‐making (which portray how people actually make their decisions). Normative theories of rational thought of relevance to medicine include epistemic theories that direct practice of evidence‐based medicine and expected utility theory, which provides the basis for widely used clinical decision analyses. Descriptive theories of rationality of direct relevance to medical decision‐making include bounded rationality, argumentative theory of reasoning, adaptive rationality, dual processing model of rationality, regret‐based rationality, pragmatic/substantive rationality, and meta‐rationality. For the first time, we provide a review of wide range of theories and models of rationality. We showed that what is “rational” behaviour under one rationality theory may be irrational under the other theory. We also showed that context is of paramount importance to rationality and that no one model of rationality can possibly fit all contexts. We suggest that in context‐poor situations, such as policy decision‐making, normative theories based on expected utility informed by best research evidence may provide the optimal approach to medical decision�

Advances in protease engineering for laundry detergents.

Science.gov (United States)

Vojcic, Ljubica; Pitzler, Christian; Körfer, Georgette; Jakob, Felix; Ronny Martinez; Maurer, Karl-Heinz; Schwaneberg, Ulrich

2015-12-25

Proteases are essential ingredients in modern laundry detergents. Over the past 30 years, subtilisin proteases employed in the laundry detergent industry have been engineered by directed evolution and rational design to tailor their properties towards industrial demands. This comprehensive review discusses recent success stories in subtilisin protease engineering. Advances in protease engineering for laundry detergents comprise simultaneous improvement of thermal resistance and activity at low temperatures, a rational strategy to modulate pH profiles, and a general hypothesis for how to increase promiscuous activity towards the production of peroxycarboxylic acids as mild bleaching agents. The three protease engineering campaigns presented provide in-depth analysis of protease properties and have identified principles that can be applied to improve or generate enzyme variants for industrial applications beyond laundry detergents. Copyright © 2015 Elsevier B.V. All rights reserved.

Selecting the Best: Evolutionary Engineering of Chemical Production in Microbes

DEFF Research Database (Denmark)

Shepelin, Denis; Hansen, Anne Sofie Lærke; Lennen, Rebecca

2018-01-01

, we focus primarily on a more challenging problem-the use of evolutionary engineering for improving the production of chemicals in microbes directly. We describe recent developments in evolutionary engineering strategies, in general, and discuss, in detail, case studies where production of a chemical......Microbial cell factories have proven to be an economical means of production for many bulk, specialty, and fine chemical products. However, we still lack both a holistic understanding of organism physiology and the ability to predictively tune enzyme activities in vivo, thus slowing down rational...... engineering of industrially relevant strains. An alternative concept to rational engineering is to use evolution as the driving force to select for desired changes, an approach often described as evolutionary engineering. In evolutionary engineering, in vivo selections for a desired phenotype are combined...

Many faces of rationality: Implications of the great rationality debate for clinical decision-making.

Science.gov (United States)

Djulbegovic, Benjamin; Elqayam, Shira

2017-10-01

Given that more than 30% of healthcare costs are wasted on inappropriate care, suboptimal care is increasingly connected to the quality of medical decisions. It has been argued that personal decisions are the leading cause of death, and 80% of healthcare expenditures result from physicians' decisions. Therefore, improving healthcare necessitates improving medical decisions, ie, making decisions (more) rational. Drawing on writings from The Great Rationality Debate from the fields of philosophy, economics, and psychology, we identify core ingredients of rationality commonly encountered across various theoretical models. Rationality is typically classified under umbrella of normative (addressing the question how people "should" or "ought to" make their decisions) and descriptive theories of decision-making (which portray how people actually make their decisions). Normative theories of rational thought of relevance to medicine include epistemic theories that direct practice of evidence-based medicine and expected utility theory, which provides the basis for widely used clinical decision analyses. Descriptive theories of rationality of direct relevance to medical decision-making include bounded rationality, argumentative theory of reasoning, adaptive rationality, dual processing model of rationality, regret-based rationality, pragmatic/substantive rationality, and meta-rationality. For the first time, we provide a review of wide range of theories and models of rationality. We showed that what is "rational" behaviour under one rationality theory may be irrational under the other theory. We also showed that context is of paramount importance to rationality and that no one model of rationality can possibly fit all contexts. We suggest that in context-poor situations, such as policy decision-making, normative theories based on expected utility informed by best research evidence may provide the optimal approach to medical decision-making, whereas in the context

Rational design and validation of an anti-protein kinase C active-state specific antibody based on conformational changes.

Science.gov (United States)

Pena, Darlene Aparecida; Andrade, Victor Piana de; Silva, Gabriela Ávila Fernandes; Neves, José Ivanildo; Oliveira, Paulo Sergio Lopes de; Alves, Maria Julia Manso; Devi, Lakshmi A; Schechtman, Deborah

2016-02-25

Protein kinase C (PKC) plays a regulatory role in key pathways in cancer. However, since phosphorylation is a step for classical PKC (cPKC) maturation and does not correlate with activation, there is a lack of tools to detect active PKC in tissue samples. Here, a structure-based rational approach was used to select a peptide to generate an antibody that distinguishes active from inactive cPKC. A peptide conserved in all cPKCs, C2Cat, was chosen since modeling studies based on a crystal structure of PKCβ showed that it is localized at the interface between the C2 and catalytic domains of cPKCs in an inactive kinase. Anti-C2Cat recognizes active cPKCs at least two-fold better than inactive kinase in ELISA and immunoprecipitation assays, and detects the temporal dynamics of cPKC activation upon receptor or phorbol stimulation. Furthermore, the antibody is able to detect active PKC in human tissue. Higher levels of active cPKC were observed in the more aggressive triple negative breast cancer tumors as compared to the less aggressive estrogen receptor positive tumors. Thus, this antibody represents a reliable, hitherto unavailable and a valuable tool to study PKC activation in cells and tissues. Similar structure-based rational design strategies can be broadly applied to obtain active-state specific antibodies for other signal transduction molecules.

Standby Gasoline Rationing Plan

Energy Technology Data Exchange (ETDEWEB)

None

1980-06-01

The final rules adopted by the President for a Standby Gasoline Rationing Plan are presented. The plan provides that eligibility for ration allotments will be determined primarily on the basis of motor vehicle registrations, taking into account historical differences in the use of gasoline among states. The regulations also provide authority for supplemental allotments to firms so that their allotment will equal a specified percentage of gasoline use during a base period. Priority classifications, i.e., agriculture, defense, etc., are established to assure adequate gasoline supplies for designated essential services. Ration rights must be provided by end-users to their suppliers for each gallon sold. DOE will regulate the distribution of gasoline at the wholesale level according to the transfer by suppliers of redeemed ration rights and the gasoline allocation regulations. Ration rights are transferable. A ration banking system is created to facilitate transfers of ration rights. Each state will be provided with a reserve of ration rights to provide for hardship needs and to alleviate inequities. (DC)

Proteins engineering

International Nuclear Information System (INIS)

2000-01-01

At the - Departement d'Ingenierie et d'etudes de proteines (Deip) of the CEA more than seventy researchers are working hard to understand the function of proteins. For that they use the molecular labelling technique (F.M.)

Rationalizing the chemical space of protein-protein interaction inhibitors.

Science.gov (United States)

Sperandio, Olivier; Reynès, Christelle H; Camproux, Anne-Claude; Villoutreix, Bruno O

2010-03-01

Protein-protein interactions (PPIs) are one of the next major classes of therapeutic targets, although they are too intricate to tackle with standard approaches. This is due, in part, to the inadequacy of today's chemical libraries. However, the emergence of a growing number of experimentally validated inhibitors of PPIs (i-PPIs) allows drug designers to use chemoinformatics and machine learning technologies to unravel the nature of the chemical space covered by the reported compounds. Key characteristics of i-PPIs can then be revealed and highlight the importance of specific shapes and/or aromatic bonds, enabling the design of i-PPI-enriched focused libraries and, therefore, of cost-effective screening strategies. 2009 Elsevier Ltd. All rights reserved.

Protein trans-splicing of multiple atypical split inteins engineered from natural inteins.

Directory of Open Access Journals (Sweden)

Ying Lin

Full Text Available Protein trans-splicing by split inteins has many uses in protein production and research. Splicing proteins with synthetic peptides, which employs atypical split inteins, is particularly useful for site-specific protein modifications and labeling, because the synthetic peptide can be made to contain a variety of unnatural amino acids and chemical modifications. For this purpose, atypical split inteins need to be engineered to have a small N-intein or C-intein fragment that can be more easily included in a synthetic peptide that also contains a small extein to be trans-spliced onto target proteins. Here we have successfully engineered multiple atypical split inteins capable of protein trans-splicing, by modifying and testing more than a dozen natural inteins. These included both S1 split inteins having a very small (11-12 aa N-intein fragment and S11 split inteins having a very small (6 aa C-intein fragment. Four of the new S1 and S11 split inteins showed high efficiencies (85-100% of protein trans-splicing both in E. coli cells and in vitro. Under in vitro conditions, they exhibited reaction rate constants ranging from ~1.7 × 10(-4 s(-1 to ~3.8 × 10(-4 s(-1, which are comparable to or higher than those of previously reported atypical split inteins. These findings should facilitate a more general use of trans-splicing between proteins and synthetic peptides, by expanding the availability of different atypical split inteins. They also have implications on understanding the structure-function relationship of atypical split inteins, particularly in terms of intein fragment complementation.

Tissue-engineered human bioartificial muscles expressing a foreign recombinant protein for gene therapy

Science.gov (United States)

Powell, C.; Shansky, J.; Del Tatto, M.; Forman, D. E.; Hennessey, J.; Sullivan, K.; Zielinski, B. A.; Vandenburgh, H. H.

1999-01-01

Murine skeletal muscle cells transduced with foreign genes and tissue engineered in vitro into bioartificial muscles (BAMs) are capable of long-term delivery of soluble growth factors when implanted into syngeneic mice (Vandenburgh et al., 1996b). With the goal of developing a therapeutic cell-based protein delivery system for humans, similar genetic tissue-engineering techniques were designed for human skeletal muscle stem cells. Stem cell myoblasts were isolated, cloned, and expanded in vitro from biopsied healthy adult (mean age, 42 +/- 2 years), and elderly congestive heart failure patient (mean age, 76 +/- 1 years) skeletal muscle. Total cell yield varied widely between biopsies (50 to 672 per 100 mg of tissue, N = 10), but was not significantly different between the two patient groups. Percent myoblasts per biopsy (73 +/- 6%), number of myoblast doublings prior to senescence in vitro (37 +/- 2), and myoblast doubling time (27 +/- 1 hr) were also not significantly different between the two patient groups. Fusion kinetics of the myoblasts were similar for the two groups after 20-22 doublings (74 +/- 2% myoblast fusion) when the biopsy samples had been expanded to 1 to 2 billion muscle cells, a number acceptable for human gene therapy use. The myoblasts from the two groups could be equally transduced ex vivo with replication-deficient retroviral expression vectors to secrete 0.5 to 2 microg of a foreign protein (recombinant human growth hormone, rhGH)/10(6) cells/day, and tissue engineered into human BAMs containing parallel arrays of differentiated, postmitotic myofibers. This work suggests that autologous human skeletal myoblasts from a potential patient population can be isolated, genetically modified to secrete foreign proteins, and tissue engineered into implantable living protein secretory devices for therapeutic use.

Adolescent rationality.

Science.gov (United States)

Moshman, David

2013-01-01

Adolescents are commonly seen as irrational, a position supported to varying degrees by many developmentalists, who often appeal to recent research on adolescent brains. Careful review of relevant evidence, however, shows that (1) adults are less rational than is generally assumed, (2) adolescents (and adults) are categorically different from children with respect to the attainment of advanced levels of rationality and psychological functioning, and (3) adolescents and adults do not differ categorically from each other with respect to any rational competencies, irrational tendencies, brain structures, or neurological functioning. Development often continues in adolescence and beyond but categorical claims about adolescents as distinct from adults cannot be justified. A review of U.S. Supreme Court decisions concerning intellectual freedom, reproductive freedom, and criminal responsibility shows ongoing ambivalence and confusion about the rationality of adolescents. Developmental theory and research suggest that adolescents should be conceptualized as young adults, not immature brains, with important implications for their roles, rights, and responsibilities.

Influence of energy concentration and source on the utilization of feed protein and NPN in lambs. 3. Allantoin excretion and microbial protein synthesis

Energy Technology Data Exchange (ETDEWEB)

Ulbrich, M; Geissler, C; Bassuny, S M; Borowiec, F; Hoffmann, M

1989-06-01

In an N balance experiment with male crossbreeding lambs at an age of 3-4 months four different rations were given differing in energy concentration (high > 700 EFU/sub cattle//kg DM and low < 650 EFU/sub cattle//kg DM) and in the energy source (sugar, starch or crude fibre) with crude protein intake being almost equal. The rations contained 2% urea. Microbial protein synthesis in the rumen was assessed according to Roth and Kichgessner (1978) (1), Rys et al. (1975) (2) and Bickel-Baumann and Landis (1986) (3) on the basis of allantoin excretion in urine. The highest ruminal protein synthesis quotas were 868-921 mg protein N per kg LW/sup 0.75/ in (2). In (3) 723-766 mg protein N/kg LW /sup 0.75/ were synthesized. From the /sup 15/N labelling of the supplemented urea and the excreted allantoin it could be calculated that 26-40% of the microbial protein resulted from the urea-N of the ration. Despite a high crude protein content of the ration of between 16 and 17% in the DM and a relation of NPN: pure protein of 0.95 the utilization of the NPN in the ration was relatively high but slightly lower than the utilization of pure protein. The variants with higher energy concentration showed as a tendency higher allantoin excretion in spite of slightly lower dry matter intake and a slightly higher NPN utilization than the variants with lower energy concentration. (author).

Love and rationality: on some possible rational effects of love

Directory of Open Access Journals (Sweden)

Gustavo Ortiz-Millán

Full Text Available In this paper I defend the idea that rather than disrupting rationality, as the common-sense conception has done it, love may actually help us to develop rational ways of thinking and acting. I make the case for romantic or erotic love, since this is the kind of love that is more frequently associated with irrationality in acting and thinking. I argue that this kind of love may make us develop epistemic and practical forms of rationality. Based on an analysis of its characteristic action tendencies, I argue that love may help us to develop an instrumental form of rationality in determining the best means to achieve the object of love. It may also narrow down the number of practical considerations that may help us to achieve our goals. Finally, love may generate rational ways of belief-formation by framing the parameters taken into account in perception and attention, and by bringing into light only a small portion of the epistemic information available. Love may make us perceive reality more acutely.

A de novo NADPH generation pathway for improving lysine production of Corynebacterium glutamicum by rational design of the coenzyme specificity of glyceraldehyde 3-phosphate dehydrogenase.

Science.gov (United States)

Bommareddy, Rajesh Reddy; Chen, Zhen; Rappert, Sugima; Zeng, An-Ping

2014-09-01

Engineering the cofactor availability is a common strategy of metabolic engineering to improve the production of many industrially important compounds. In this work, a de novo NADPH generation pathway is proposed by altering the coenzyme specificity of a native NAD-dependent glyceraldehyde 3-phosphate dehydrogenase (GAPDH) to NADP, which consequently has the potential to produce additional NADPH in the glycolytic pathway. Specifically, the coenzyme specificity of GAPDH of Corynebacterium glutamicum is systematically manipulated by rational protein design and the effect of the manipulation for cellular metabolism and lysine production is evaluated. By a combinatorial modification of four key residues within the coenzyme binding sites, different GAPDH mutants with varied coenzyme specificity were constructed. While increasing the catalytic efficiency of GAPDH towards NADP enhanced lysine production in all of the tested mutants, the most significant improvement of lysine production (~60%) was achieved with the mutant showing similar preference towards both NAD and NADP. Metabolic flux analysis with (13)C isotope studies confirmed that there was no significant change of flux towards the pentose phosphate pathway and the increased lysine yield was mainly attributed to the NADPH generated by the mutated GAPDH. The present study highlights the importance of protein engineering as a key strategy in de novo pathway design and overproduction of desired products. Copyright © 2014 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Adhesive protein interactions with chitosan: consequences for valve endothelial cell growth on tissue-engineering materials.

Science.gov (United States)

Cuy, Janet L; Beckstead, Benjamin L; Brown, Chad D; Hoffman, Allan S; Giachelli, Cecilia M

2003-11-01

Stable endothelialization of a tissue-engineered heart valve is essential for proper valve function, although adhesive characteristics of the native valve endothelial cell (VEC) have rarely been explored. This research evaluated VEC adhesive qualities and attempted to enhance VEC growth on the biopolymer chitosan, a novel tissue-engineering scaffold material with promising biological and chemical properties. Aortic VEC cultures were isolated and found to preferentially adhere to fibronectin, collagen types IV and I over laminin and osteopontin in a dose-dependent manner. Seeding of VEC onto comparison substrates revealed VEC growth and morphology to be preferential in the order: tissue culture polystyrene > gelatin, poly(DL-lactide-co-glycolide), chitosan > poly(hydroxy alkanoate). Adhesive protein precoating of chitosan did not significantly enhance VEC growth, despite equivalent protein adsorption as to polystyrene. Initial cell adhesion to protein-precoated chitosan, however, was higher than for polystyrene. Composite chitosan/collagen type IV films were investigated as an alternative to simple protein precoatings, and were shown to improve VEC growth and morphology over chitosan alone. These findings suggest potential manipulation of chitosan properties to improve amenability to valve tissue-engineering applications. Copyright 2003 Wiley Periodicals, Inc.

Engineering a novel multifunctional green fluorescent protein tag for a wide variety of protein research.

Directory of Open Access Journals (Sweden)

Takuya Kobayashi

Full Text Available BACKGROUND: Genetically encoded tag is a powerful tool for protein research. Various kinds of tags have been developed: fluorescent proteins for live-cell imaging, affinity tags for protein isolation, and epitope tags for immunological detections. One of the major problems concerning the protein tagging is that many constructs with different tags have to be made for different applications, which is time- and resource-consuming. METHODOLOGY/PRINCIPAL FINDINGS: Here we report a novel multifunctional green fluorescent protein (mfGFP tag which was engineered by inserting multiple peptide tags, i.e., octa-histidine (8xHis, streptavidin-binding peptide (SBP, and c-Myc tag, in tandem into a loop of GFP. When fused to various proteins, mfGFP monitored their localization in living cells. Streptavidin agarose column chromatography with the SBP tag successfully isolated the protein complexes in a native form with a high purity. Tandem affinity purification (TAP with 8xHis and SBP tags in mfGFP further purified the protein complexes. mfGFP was clearly detected by c-Myc-specific antibody both in immunofluorescence and immuno-electron microscopy (EM. These findings indicate that mfGFP works well as a multifunctional tag in mammalian cells. The tag insertion was also successful in other fluorescent protein, mCherry. CONCLUSIONS AND SIGNIFICANCE: The multifunctional fluorescent protein tag is a useful tool for a wide variety of protein research, and may have the advantage over other multiple tag systems in its higher expandability and compatibility with existing and future tag technologies.

Rational Design of a New Trypanosoma rangeli Trans-Sialidase for Efficient Sialylation of Glycans

DEFF Research Database (Denmark)

Jers, Carsten; Michalak, Malwina; Larsen, Dorte Møller

2014-01-01

This paper reports rational engineering of Trypanosoma rangeli sialidase to develop an effective enzyme for a potentially important type of reactivity: production of sialylated prebiotic glycans. The Trypanosoma cruzi trans-sialidase and the homologous T. rangeli sialidase has previously been use...

Ligand-regulated peptides: a general approach for modulating protein-peptide interactions with small molecules.

Science.gov (United States)

Binkowski, Brock F; Miller, Russell A; Belshaw, Peter J

2005-07-01

We engineered a novel ligand-regulated peptide (LiRP) system where the binding activity of intracellular peptides is controlled by a cell-permeable small molecule. In the absence of ligand, peptides expressed as fusions in an FKBP-peptide-FRB-GST LiRP scaffold protein are free to interact with target proteins. In the presence of the ligand rapamycin, or the nonimmunosuppressive rapamycin derivative AP23102, the scaffold protein undergoes a conformational change that prevents the interaction of the peptide with the target protein. The modular design of the scaffold enables the creation of LiRPs through rational design or selection from combinatorial peptide libraries. Using these methods, we identified LiRPs that interact with three independent targets: retinoblastoma protein, c-Src, and the AMP-activated protein kinase. The LiRP system should provide a general method to temporally and spatially regulate protein function in cells and organisms.

Rationally engineered nanoparticles target multiple myeloma cells, overcome cell-adhesion-mediated drug resistance, and show enhanced efficacy in vivo

International Nuclear Information System (INIS)

Kiziltepe, T; Ashley, J D; Stefanick, J F; Qi, Y M; Alves, N J; Handlogten, M W; Suckow, M A; Navari, R M; Bilgicer, B

2012-01-01

In the continuing search for effective cancer treatments, we report the rational engineering of a multifunctional nanoparticle that combines traditional chemotherapy with cell targeting and anti-adhesion functionalities. Very late antigen-4 (VLA-4) mediated adhesion of multiple myeloma (MM) cells to bone marrow stroma confers MM cells with cell-adhesion-mediated drug resistance (CAM-DR). In our design, we used micellar nanoparticles as dynamic self-assembling scaffolds to present VLA-4-antagonist peptides and doxorubicin (Dox) conjugates, simultaneously, to selectively target MM cells and to overcome CAM-DR. Dox was conjugated to the nanoparticles through an acid-sensitive hydrazone bond. VLA-4-antagonist peptides were conjugated via a multifaceted synthetic procedure for generating precisely controlled number of targeting functionalities. The nanoparticles were efficiently internalized by MM cells and induced cytotoxicity. Mechanistic studies revealed that nanoparticles induced DNA double-strand breaks and apoptosis in MM cells. Importantly, multifunctional nanoparticles overcame CAM-DR, and were more efficacious than Dox when MM cells were cultured on fibronectin-coated plates. Finally, in a MM xenograft model, nanoparticles preferentially homed to MM tumors with ∼10 fold more drug accumulation and demonstrated dramatic tumor growth inhibition with a reduced overall systemic toxicity. Altogether, we demonstrate the disease driven engineering of a nanoparticle-based drug delivery system, enabling the model of an integrative approach in the treatment of MM

Identifying and engineering promoters for high level and sustainable therapeutic recombinant protein production in cultured mammalian cells.

Science.gov (United States)

Ho, Steven C L; Yang, Yuansheng

2014-08-01

Promoters are essential on plasmid vectors to initiate transcription of the transgenes when generating therapeutic recombinant proteins expressing mammalian cell lines. High and sustained levels of gene expression are desired during therapeutic protein production while gene expression is useful for cell engineering. As many finely controlled promoters exhibit cell and product specificity, new promoters need to be identified, optimized and carefully evaluated before use. Suitable promoters can be identified using techniques ranging from simple molecular biology methods to modern high-throughput omics screenings. Promoter engineering is often required after identification to either obtain high and sustained expression or to provide a wider range of gene expression. This review discusses some of the available methods to identify and engineer promoters for therapeutic recombinant protein expression in mammalian cells.

CONTRIBUTIONS TO RATIONAL APPROXIMATION,

Science.gov (United States)

Some of the key results of linear Chebyshev approximation theory are extended to generalized rational functions. Prominent among these is Haar’s...linear theorem which yields necessary and sufficient conditions for uniqueness. Some new results in the classic field of rational function Chebyshev...Furthermore a Weierstrass type theorem is proven for rational Chebyshev approximation. A characterization theorem for rational trigonometric Chebyshev approximation in terms of sign alternation is developed. (Author)

Rational points, rational curves, and entire holomorphic curves on projective varieties

CERN Document Server

Gasbarri, Carlo; Roth, Mike; Tschinkel, Yuri

2015-01-01

This volume contains papers from the Short Thematic Program on Rational Points, Rational Curves, and Entire Holomorphic Curves and Algebraic Varieties, held from June 3-28, 2013, at the Centre de Recherches Mathématiques, Université de Montréal, Québec, Canada. The program was dedicated to the study of subtle interconnections between geometric and arithmetic properties of higher-dimensional algebraic varieties. The main areas of the program were, among others, proving density of rational points in Zariski or analytic topology on special varieties, understanding global geometric properties of rationally connected varieties, as well as connections between geometry and algebraic dynamics exploring new geometric techniques in Diophantine approximation.

Reversible S-nitrosylation in an engineered azurin

Energy Technology Data Exchange (ETDEWEB)

Tian, Shiliang; Liu, Jing; Cowley, Ryan E.; Hosseinzadeh, Parisa; Marshall, Nicholas M.; Yu, Yang; Robinson, Howard; Nilges, Mark J.; Blackburn, Ninian J.; Solomon, Edward I.; Lu, Yi

2016-04-25

S-Nitrosothiols are known as reagents for NO storage and transportation and as regulators in many physiological processes. Although the S-nitrosylation catalysed by haem proteins is well known, no direct evidence of S-nitrosylation in copper proteins has been reported. Here, we report reversible insertion of NO into a copper–thiolate bond in an engineered copper centre in Pseudomonas aeruginosa azurin by rational design of the primary coordination sphere and tuning its reduction potential by deleting a hydrogen bond in the secondary coordination sphere. The results not only provide the first direct evidence of S-nitrosylation of Cu(II)-bound cysteine in metalloproteins, but also shed light on the reaction mechanism and structural features responsible for stabilizing the elusive Cu(I)–S(Cys)NO species. The fast, efficient and reversible S-nitrosylation reaction is used to demonstrate its ability to prevent NO inhibition of cytochrome bo3 oxidase activity by competing for NO binding with the native enzyme under physiologically relevant conditions.

Rational Multiparty Computation

OpenAIRE

Wallrabenstein, John Ross

2014-01-01

The field of rational cryptography considers the design of cryptographic protocols in the presence of rational agents seeking to maximize local utility functions. This departs from the standard secure multiparty computation setting, where players are assumed to be either honest or malicious. ^ We detail the construction of both a two-party and a multiparty game theoretic framework for constructing rational cryptographic protocols. Our framework specifies the utility function assumptions neces...

Engineering central metabolism – a grand challenge for plant biologists

DEFF Research Database (Denmark)

Sweetlove, Lee J.; Nielsen, Jens; Fernie, Alisdair R.

2017-01-01

The goal of increasing crop productivity and nutrient-use efficiency is being addressed by a number of ambitious research projects seeking to re-engineer photosynthetic biochemistry. Many of these projects will require the engineering of substantial changes in fluxes of central metabolism. However......, as has been amply demonstrated in simpler systems such as microbes, central metabolism is extremely difficult to rationally engineer. This is because of multiple layers of regulation that operate to maintain metabolic steady state and because of the highly connected nature of central metabolism....... In this review we discuss new approaches for metabolic engineering that have the potential to address these problems and dramatically improve the success with which we can rationally engineer central metabolism in plants. In particular, we advocate the adoption of an iterative ‘design-build-test-learn’ cycle...

Rationally designed, heterologous S. cerevisiae transcripts expose novel expression determinants

Science.gov (United States)

Ben-Yehezkel, Tuval; Atar, Shimshi; Zur, Hadas; Diament, Alon; Goz, Eli; Marx, Tzipy; Cohen, Rafael; Dana, Alexandra; Feldman, Anna; Shapiro, Ehud; Tuller, Tamir

2015-01-01

Deducing generic causal relations between RNA transcript features and protein expression profiles from endogenous gene expression data remains a major unsolved problem in biology. The analysis of gene expression from heterologous genes contributes significantly to solving this problem, but has been heavily biased toward the study of the effect of 5′ transcript regions and to prokaryotes. Here, we employ a synthetic biology driven approach that systematically differentiates the effect of different regions of the transcript on gene expression up to 240 nucleotides into the ORF. This enabled us to discover new causal effects between features in previously unexplored regions of transcripts, and gene expression in natural regimes. We rationally designed, constructed, and analyzed 383 gene variants of the viral HRSVgp04 gene ORF, with multiple synonymous mutations at key positions along the transcript in the eukaryote S. cerevisiae. Our results show that a few silent mutations at the 5′UTR can have a dramatic effect of up to 15 fold change on protein levels, and that even synonymous mutations in positions more than 120 nucleotides downstream from the ORF 5′end can modulate protein levels up to 160%–300%. We demonstrate that the correlation between protein levels and folding energy increases with the significance of the level of selection of the latter in endogenous genes, reinforcing the notion that selection for folding strength in different parts of the ORF is related to translation regulation. Our measured protein abundance correlates notably(correlation up to r = 0.62 (p=0.0013)) with mean relative codon decoding times, based on ribosomal densities (Ribo-Seq) in endogenous genes, supporting the conjecture that translation elongation and adaptation to the tRNA pool can modify protein levels in a causal/direct manner. This report provides an improved understanding of transcript evolution, design principles of gene expression regulation, and suggests simple

Chromophore photophysics and dynamics in fluorescent proteins of the GFP family

International Nuclear Information System (INIS)

Nienhaus, Karin; Nienhaus, G Ulrich

2016-01-01

Proteins of the green fluorescent protein (GFP) family are indispensable for fluorescence imaging experiments in the life sciences, particularly of living specimens. Their essential role as genetically encoded fluorescence markers has motivated many researchers over the last 20 years to further advance and optimize these proteins by using protein engineering. Amino acids can be exchanged by site-specific mutagenesis, starting with naturally occurring proteins as templates. Optical properties of the fluorescent chromophore are strongly tuned by the surrounding protein environment, and a targeted modification of chromophore-protein interactions requires a profound knowledge of the underlying photophysics and photochemistry, which has by now been well established from a large number of structural and spectroscopic experiments and molecular-mechanical and quantum-mechanical computations on many variants of fluorescent proteins. Nevertheless, such rational engineering often does not meet with success and thus is complemented by random mutagenesis and selection based on the optical properties. In this topical review, we present an overview of the key structural and spectroscopic properties of fluorescent proteins. We address protein-chromophore interactions that govern ground state optical properties as well as processes occurring in the electronically excited state. Special emphasis is placed on photoactivation of fluorescent proteins. These light-induced reactions result in large structural changes that drastically alter the fluorescence properties of the protein, which enables some of the most exciting applications, including single particle tracking, pulse chase imaging and super-resolution imaging. We also present a few examples of fluorescent protein application in live-cell imaging experiments. (topical review)

Chromophore photophysics and dynamics in fluorescent proteins of the GFP family

Science.gov (United States)

Nienhaus, Karin; Nienhaus, G. Ulrich

2016-11-01

Proteins of the green fluorescent protein (GFP) family are indispensable for fluorescence imaging experiments in the life sciences, particularly of living specimens. Their essential role as genetically encoded fluorescence markers has motivated many researchers over the last 20 years to further advance and optimize these proteins by using protein engineering. Amino acids can be exchanged by site-specific mutagenesis, starting with naturally occurring proteins as templates. Optical properties of the fluorescent chromophore are strongly tuned by the surrounding protein environment, and a targeted modification of chromophore-protein interactions requires a profound knowledge of the underlying photophysics and photochemistry, which has by now been well established from a large number of structural and spectroscopic experiments and molecular-mechanical and quantum-mechanical computations on many variants of fluorescent proteins. Nevertheless, such rational engineering often does not meet with success and thus is complemented by random mutagenesis and selection based on the optical properties. In this topical review, we present an overview of the key structural and spectroscopic properties of fluorescent proteins. We address protein-chromophore interactions that govern ground state optical properties as well as processes occurring in the electronically excited state. Special emphasis is placed on photoactivation of fluorescent proteins. These light-induced reactions result in large structural changes that drastically alter the fluorescence properties of the protein, which enables some of the most exciting applications, including single particle tracking, pulse chase imaging and super-resolution imaging. We also present a few examples of fluorescent protein application in live-cell imaging experiments.

Rational Design of Rho Protein Inhibitors

National Research Council Canada - National Science Library

Rojas, Rafael J

2006-01-01

Rho GTPases are molecular switches that fluctuate between on and off states. When active, these proteins function to remodel the actin cytoskeleton by interacting with a number of downstream effector molecules...

Rational Design of Rho Protein Inhibitors

National Research Council Canada - National Science Library

Rojas, Rafael J

2005-01-01

Rho GTPases are molecular switches that fluctuate between on and off states. When active, these proteins function to remodel the actin cytoskeleton by interacting with a number of downstream effector molecules...

Effects of Bone Morphogenic Proteins on Engineered Cartilage

Science.gov (United States)

Gooch, Keith, J.; Blunk, Torsten; Courter, Donald L.; Sieminski, Alisha; Vunjak-Novakovic, Gordana; Freed, Lisa E.

2007-01-01

A report describes experiments on the effects of bone morphogenic proteins (BMPs) on engineered cartilage grown in vitro. In the experiments, bovine calf articular chondrocytes were seeded onto biodegradable polyglycolic acid scaffolds and cultured in, variously, a control medium or a medium supplemented with BMP-2, BMP-12, or BMP-13 in various concentrations. Under all conditions investigated, cell-polymer constructs cultivated for 4 weeks macroscopically and histologically resembled native cartilage. At a concentration of 100 ng/mL, BMP-2, BMP-12, or BMP-13 caused (1) total masses of the constructs to exceed those of the controls by 121, 80, or 62 percent, respectively; (2) weight percentages of glycosaminoglycans in the constructs to increase by 27, 18, or 15, respectively; and (3) total collagen contents of the constructs to decrease to 63, 89, or 83 percent of the control values, respectively. BMP-2, but not BMP-12 or BMP-13, promoted chondrocyte hypertrophy. These observations were interpreted as suggesting that the three BMPs increase the growth rates and modulate the compositions of engineered cartilage. It was also concluded that in vitro engineered cartilage is a suitable system for studying effects of BMPs on chondrogenesis in a well-defined environment.

Facile construction of a random protein domain insertion library using an engineered transposon.

Science.gov (United States)

Shah, Vandan; Pierre, Brennal; Kim, Jin Ryoun

2013-01-15

Insertional fusion between multiple protein domains represents a novel means of creating integrated functionalities. Currently, there is no robust guideline for selection of insertion sites ensuring the desired functional outcome of insertional fusion. Therefore, construction and testing of random domain insertion libraries, in which a host protein domain is randomly inserted into a guest protein domain, significantly benefit extensive exploration of sequence spaces for insertion sites. Short peptide residues are usually introduced between protein domains to alleviate structural conflicts, and the interdomain linker residues may affect the functional outcome of protein insertion complexes. Unfortunately, optimal control of interdomain linker residues is not always available in conventional methods used to construct random domain insertion libraries. Moreover, most conventional methods employ blunt-end rather than sticky-end ligation between host and guest DNA fragments, thus lowering library construction efficiency. Here, we report the facile construction of random domain insertion libraries using an engineered transposon. We show that random domain insertion with optimal control of interdomain linker residues was possible with our engineered transposon-based method. In addition, our method employs sticky-end rather than blunt-end ligation between host and guest DNA fragments, thus allowing for facile construction of relatively large sized libraries. Copyright © 2012 Elsevier Inc. All rights reserved.

Protein design and engineering of a de novo pathway for microbial production of 1,3-propanediol from glucose.

Science.gov (United States)

Chen, Zhen; Geng, Feng; Zeng, An-Ping

2015-02-01

Protein engineering to expand the substrate spectrum of native enzymes opens new possibilities for bioproduction of valuable chemicals from non-natural pathways. No natural microorganism can directly use sugars to produce 1,3-propanediol (PDO). Here, we present a de novo route for the biosynthesis of PDO from sugar, which may overcome the mentioned limitations by expanding the homoserine synthesis pathway. The accomplishment of pathway from homoserine to PDO is achieved by protein engineering of glutamate dehydrogenase (GDH) and pyruvate decarboxylase to sequentially convert homoserine to 4-hydroxy-2-ketobutyrate and 3-hydroxypropionaldehyde. The latter is finally converted to PDO by using a native alcohol dehydrogenase. In this work, we report on experimental accomplishment of this non-natural pathway, especially by protein engineering of GDH for the key step of converting homoserine to 4-hydroxy-2-ketobutyrate. These results show the feasibility and significance of protein engineering for de novo pathway design and overproduction of desired industrial products. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

CRISPR/Cas9-mediated genome engineering of CHO cell factories: application and perspectives

DEFF Research Database (Denmark)

Lee, Jae Seong; Grav, Lise Marie; Lewis, Nathan E.

2015-01-01

repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) system enables rapid,easy and efficient engineering of mammalian genomes. It has a wide range of applications frommodification of individual genes to genome-wide screening or regulation of genes. Facile genomeediting using CRISPR/Cas9 empowers...... researchers in the CHO community to elucidate the mechanisticbasis behind high level production of proteins and product quality attributes of interest. Inthis review, we describe the basis of CRISPR/Cas9-mediated genome editing and its applicationfor development of next generation CHO cell factories while...... highlighting both future perspectivesand challenges. As one of the main drivers for the CHO systems biology era, genome engineeringwith CRISPR/Cas9 will pave the way for rational design of CHO cell factories....

Irrational Rationality of Terrorism

Directory of Open Access Journals (Sweden)

Robert Nalbandov

2013-12-01

Full Text Available The present article deals with the ontological problem of applying the rational choice frameworks to the study of terrorism. It testing the application of the rational choice to the “old” (before the end of the Cold War and the “new” (after the end of the Cold War terrorisms. It starts with analyzing the fundamentals of rationality and applies it at two levels: the individual (actors and group (collective via two outlooks: tactical (short-term and strategic (long-term. The main argument of the article is that while the “old” terrorism can be explained by the rational choice theory its “new” version represents a substantial departure from rationality.

Artificial membrane-binding proteins stimulate oxygenation of stem cells during engineering of large cartilage tissue

Science.gov (United States)

Armstrong, James P. K.; Shakur, Rameen; Horne, Joseph P.; Dickinson, Sally C.; Armstrong, Craig T.; Lau, Katherine; Kadiwala, Juned; Lowe, Robert; Seddon, Annela; Mann, Stephen; Anderson, J. L. Ross; Perriman, Adam W.; Hollander, Anthony P.

2015-06-01

Restricted oxygen diffusion can result in central cell necrosis in engineered tissue, a problem that is exacerbated when engineering large tissue constructs for clinical application. Here we show that pre-treating human mesenchymal stem cells (hMSCs) with synthetic membrane-active myoglobin-polymer-surfactant complexes can provide a reservoir of oxygen capable of alleviating necrosis at the centre of hyaline cartilage. This is achieved through the development of a new cell functionalization methodology based on polymer-surfactant conjugation, which allows the delivery of functional proteins to the hMSC membrane. This new approach circumvents the need for cell surface engineering using protein chimerization or genetic transfection, and we demonstrate that the surface-modified hMSCs retain their ability to proliferate and to undergo multilineage differentiation. The functionalization technology is facile, versatile and non-disruptive, and in addition to tissue oxygenation, it should have far-reaching application in a host of tissue engineering and cell-based therapies.

Automatic and integrated micro-enzyme assay (AIμEA) platform for highly sensitive thrombin analysis via an engineered fluorescence protein-functionalized monolithic capillary column.

Science.gov (United States)

Lin, Lihua; Liu, Shengquan; Nie, Zhou; Chen, Yingzhuang; Lei, Chunyang; Wang, Zhen; Yin, Chao; Hu, Huiping; Huang, Yan; Yao, Shouzhuo

2015-04-21

Nowadays, large-scale screening for enzyme discovery, engineering, and drug discovery processes require simple, fast, and sensitive enzyme activity assay platforms with high integration and potential for high-throughput detection. Herein, a novel automatic and integrated micro-enzyme assay (AIμEA) platform was proposed based on a unique microreaction system fabricated by a engineered green fluorescence protein (GFP)-functionalized monolithic capillary column, with thrombin as an example. The recombinant GFP probe was rationally engineered to possess a His-tag and a substrate sequence of thrombin, which enable it to be immobilized on the monolith via metal affinity binding, and to be released after thrombin digestion. Combined with capillary electrophoresis-laser-induced fluorescence (CE-LIF), all the procedures, including thrombin injection, online enzymatic digestion in the microreaction system, and label-free detection of the released GFP, were integrated in a single electrophoretic process. By taking advantage of the ultrahigh loading capacity of the AIμEA platform and the CE automatic programming setup, one microreaction column was sufficient for many times digestion without replacement. The novel microreaction system showed significantly enhanced catalytic efficiency, about 30 fold higher than that of the equivalent bulk reaction. Accordingly, the AIμEA platform was highly sensitive with a limit of detection down to 1 pM of thrombin. Moreover, the AIμEA platform was robust and reliable to detect thrombin in human serum samples and its inhibition by hirudin. Hence, this AIμEA platform exhibits great potential for high-throughput analysis in future biological application, disease diagnostics, and drug screening.

Lying for Strategic Advantage: Rational and Boundedly Rational Misrepresentation of Intentions

OpenAIRE

Crawford, Vincent P.

2001-01-01

Starting from Hendricks and McAfee's (2000) example of the Allies' decision to feint at Calais and attack at Normandy on D-Day, this paper models misrepresentation of intentions to competitors or enemies. Allowing for the possibility of bounded strategic rationality and rational players' responses to it yields a sensible account of lying via costless, noiseless messages. In many cases the model has generically unique pure-strategy sequential equilibria, in which rational players exploit bound...

Getting the big beast to work--systems biotechnology of Bacillus megaterium for novel high-value proteins.

Science.gov (United States)

Korneli, Claudia; David, Florian; Biedendieck, Rebekka; Jahn, Dieter; Wittmann, Christoph

2013-01-20

The high industrial relevance of the soil bacterium Bacillus megaterium as host for recombinant proteins is driving systems-wide analyses of its metabolic and regulatory networks. The present review highlights novel systems biology tools available to unravel the various cellular components on the level of metabolic and regulatory networks. These provide a rational platform for systems metabolic engineering of B. megaterium. In line, a number of interesting studies have particularly focused on studying recombinant B. megaterium in its industrial bioprocess environment thus integrating systems metabolic engineering with systems biotechnology and providing the full picture toward optimal processes. Copyright © 2012 Elsevier B.V. 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.

COMPARATIVE EFFICIENCY OF CALF STARTER AND CONVENTIONAL RATIONS IN BUFFALO SUCKLING CALVES

Directory of Open Access Journals (Sweden)

F. Ahmad, M. A. Jabbar1, I. Ahmad2 , M. Rafique and I. Ahmad3

2004-10-01

Full Text Available Twenty-four buffalo calves, having similar age and initial body weight, were divided into two groups with equal number of calves of both sexes in each group to study the effect of calf starter ration on feed intake and weight gain. Calf starter and conventional dairy rations with crude protein 18% and total digestible nutrients 75–80% along with green fodder were offered ad libitum to calves of respective groups for a period of 113 days. The average daily feed intakes were 0.95 and 0.57 kg, average daily weight gains were 0.47 and 0.34 kg and feed conversion ratio averaged 2.00 and 1.70 in calf starter and conventional groups respectively. On the overall performance, calf starter group was found better than the conventional ration.

Determinants of Actor Rationality

DEFF Research Database (Denmark)

Ellegaard, Chris

Industrial companies must exercise influence on their suppliers (or supplier actors). Actor rationality is a central theme connected to this management task. In this article, relevant literature is studied with the purpose of shedding light on determinants of actor rationality. Two buyer-supplier...... relations are investigated in a multiple case study, leading to the proposal of various additional factors that determine and shape actor rationality. Moreover a conceptual model of rationality determinants in the buyer-supplier relation is proposed, a model that may help supply managers analyse...

Two Concepts of Rationality

Directory of Open Access Journals (Sweden)

Danny Frederick

2010-02-01

Full Text Available The dominant tradition in Western philosophy sees rationality as dictating. Thus rationality may require that we believe the best explanation and simple conceptual truths and that we infer in accordance with evident rules of inference. I argue that, given what we know about the growth of knowledge, this authoritarian concept of rationality leads to absurdities and should be abandoned. I then outline a libertarian concept of rationality, derived from Popper, which eschews the dictates and which sees a rational agent as one who questions, criticises, conjectures and experiments. I argue that, while the libertarian approach escapes the absurdities of the authoritarian, it requires two significant developments and an important clarification to be made fully consistent with itself.

Rationality and institutions : an inquiry into the normative implications of rational choice theory

OpenAIRE

Engelen, Bart

2007-01-01

I aim to analyze in this dissertation what a desirable basic institutional structure looks like from the perspective of rationality. While the main topic is thus normative in nature, I start by clarifying in the first part what the notion of rationality exactly entails. I do so by focusing explicitly on the economic conception of rationality, according to which a rational individual is motivated to serve his self-interest on the basis of cost-benefit calculations. Such a Homo Economicus is ch...

Engineering nanomaterials with a combined electrochemical and molecular biomimetic approach

Science.gov (United States)

Dai, Haixia

high affinity of the engineered TraI for Cu2O over other related copper compounds led to the formation of Cu2O nanoparticles from a cuprous chloride complex (Cu2Cln1-n, n = 2 or 3) electrolyte under conditions where the mineral atacamite (CuCl(OH) 3) is thermodynamically preferred. The nonequilibrium Cu 2O nanoparticles consisted of 2--3 nm Cu2O cores and functional protein shells that enabled predictable meso-scale assembly on DNA templates. In short, we have rationally designed a protein-based scheme for forming and organizing inorganic materials that Nature has not previous worked with.

Rationing medical education.

African Journals Online (AJOL)

This paper discussed the pros and cons of the application of rationing to medical education and the different ... Different types of rationing exist in healthcare professional education. ... state-of-the-art resources, technology and tutors con-.

Development of Genome Engineering Tools from Plant-Specific PPR Proteins Using Animal Cultured Cells.

Science.gov (United States)

Kobayashi, Takehito; Yagi, Yusuke; Nakamura, Takahiro

2016-01-01

The pentatricopeptide repeat (PPR) motif is a sequence-specific RNA/DNA-binding module. Elucidation of the RNA/DNA recognition mechanism has enabled engineering of PPR motifs as new RNA/DNA manipulation tools in living cells, including for genome editing. However, the biochemical characteristics of PPR proteins remain unknown, mostly due to the instability and/or unfolding propensities of PPR proteins in heterologous expression systems such as bacteria and yeast. To overcome this issue, we constructed reporter systems using animal cultured cells. The cell-based system has highly attractive features for PPR engineering: robust eukaryotic gene expression; availability of various vectors, reagents, and antibodies; highly efficient DNA delivery ratio (>80 %); and rapid, high-throughput data production. In this chapter, we introduce an example of such reporter systems: a PPR-based sequence-specific translational activation system. The cell-based reporter system can be applied to characterize plant genes of interested and to PPR engineering.

Rationality problem for algebraic tori

CERN Document Server

Hoshi, Akinari

2017-01-01

The authors give the complete stably rational classification of algebraic tori of dimensions 4 and 5 over a field k. In particular, the stably rational classification of norm one tori whose Chevalley modules are of rank 4 and 5 is given. The authors show that there exist exactly 487 (resp. 7, resp. 216) stably rational (resp. not stably but retract rational, resp. not retract rational) algebraic tori of dimension 4, and there exist exactly 3051 (resp. 25, resp. 3003) stably rational (resp. not stably but retract rational, resp. not retract rational) algebraic tori of dimension 5. The authors make a procedure to compute a flabby resolution of a G-lattice effectively by using the computer algebra system GAP. Some algorithms may determine whether the flabby class of a G-lattice is invertible (resp. zero) or not. Using the algorithms, the suthors determine all the flabby and coflabby G-lattices of rank up to 6 and verify that they are stably permutation. The authors also show that the Krull-Schmidt theorem for G-...

Rational Approximations to Rational Models: Alternative Algorithms for Category Learning

Science.gov (United States)

Sanborn, Adam N.; Griffiths, Thomas L.; Navarro, Daniel J.

2010-01-01

Rational models of cognition typically consider the abstract computational problems posed by the environment, assuming that people are capable of optimally solving those problems. This differs from more traditional formal models of cognition, which focus on the psychological processes responsible for behavior. A basic challenge for rational models…

Design, Engineering and Application of an Amyloidogenic Protein, SBAFP-m1, for use in Nanotechnological Applications

Science.gov (United States)

Peralta, Maria del Refugio

Nanotechnology relies on collaborations across scientific disciplines such as physics, engineering, chemistry and biology. In nanotechnology, researchers manipulate molecules on the nanometer scale for various applications, ranging from tissue engineering, nanowire synthesis, and alternative energy devices. By utilizing various biological scaffolds, namely amyloid fibrils, the work of nanometer molecular control can be achieved through the use of self-assembly systems. Here, a systematic design scheme was developed to engineer protein based amyloid fibrils and was successfully applied to the design of two, unique self-assembled monomers, SBAFP-m1 and RGAFP-m1, from naturally occurring ice binding proteins found in insects and plants. A highly idealized, in-register dimer interface was designed and experimentally synthesized and demonstrated to form micron long amyloid fibrils (Chapter 2). The strength and resistance of the designer amyloid fibrils formed by SBAFP-m1 were probed in Chapter 3. Most notably, the ultimate tensile strength of SBAFP-m1 fibrils was experimentally determined to be 2.1 +/- 1.7 GPa, on par with that of naturally occurring amyloid fibrils in literature and steel. The fibrils were found to maintain their beta-sheet structure over a wide range of temperatures, from - 80 °C to 90 °C. Fibrils were resistant to common protein denaturants like 8M urea, 2.5 M guanidine hydrochloride, 2.5 M NaCl, organic solvents (methanol, ethanol, isopropanol and acetone), and across the pH range two to 11. SBAFP-m1 was mutated to add a 5x cysteine tag to the N-terminus, allowing for gold nanoparticle conjugation along the fibril axis (Chapter 4). The gold-conjugated fibrils were then enhanced with silver to produce nanowires. Various attempts to selectively synthesize heterogeneous fibrils from SBAFP-m1 mutants were attempted in Chapter 5. An attempt to de-stabilize the homogeneous fibril assembly through unfavorable homogeneous protein interactions was not

Exploring G Protein-Coupled Receptors (GPCRs) Ligand Space via Cheminformatics Approaches: Impact on Rational Drug Design

Science.gov (United States)

Basith, Shaherin; Cui, Minghua; Macalino, Stephani J. Y.; Park, Jongmi; Clavio, Nina A. B.; Kang, Soosung; Choi, Sun

2018-01-01

The primary goal of rational drug discovery is the identification of selective ligands which act on single or multiple drug targets to achieve the desired clinical outcome through the exploration of total chemical space. To identify such desired compounds, computational approaches are necessary in predicting their drug-like properties. G Protein-Coupled Receptors (GPCRs) represent one of the largest and most important integral membrane protein families. These receptors serve as increasingly attractive drug targets due to their relevance in the treatment of various diseases, such as inflammatory disorders, metabolic imbalances, cardiac disorders, cancer, monogenic disorders, etc. In the last decade, multitudes of three-dimensional (3D) structures were solved for diverse GPCRs, thus referring to this period as the “golden age for GPCR structural biology.” Moreover, accumulation of data about the chemical properties of GPCR ligands has garnered much interest toward the exploration of GPCR chemical space. Due to the steady increase in the structural, ligand, and functional data of GPCRs, several cheminformatics approaches have been implemented in its drug discovery pipeline. In this review, we mainly focus on the cheminformatics-based paradigms in GPCR drug discovery. We provide a comprehensive view on the ligand– and structure-based cheminformatics approaches which are best illustrated via GPCR case studies. Furthermore, an appropriate combination of ligand-based knowledge with structure-based ones, i.e., integrated approach, which is emerging as a promising strategy for cheminformatics-based GPCR drug design is also discussed. PMID:29593527

Reprogramming cells with synthetic proteins

Directory of Open Access Journals (Sweden)

Xiaoxiao Yang

2015-06-01

Full Text Available Conversion of one cell type into another cell type by forcibly expressing specific cocktails of transcription factors (TFs has demonstrated that cell fates are not fixed and that cellular differentiation can be a two-way street with many intersections. These experiments also illustrated the sweeping potential of TFs to "read" genetically hardwired regulatory information even in cells where they are not normally expressed and to access and open up tightly packed chromatin to execute gene expression programs. Cellular reprogramming enables the modeling of diseases in a dish, to test the efficacy and toxicity of drugs in patient-derived cells and ultimately, could enable cell-based therapies to cure degenerative diseases. Yet, producing terminally differentiated cells that fully resemble their in vivocounterparts in sufficient quantities is still an unmet clinical need. While efforts are being made to reprogram cells nongenetically by using drug-like molecules, defined TF cocktails still dominate reprogramming protocols. Therefore, the optimization of TFs by protein engineering has emerged as a strategy to enhance reprogramming to produce functional, stable and safe cells for regenerative biomedicine. Engineering approaches focused on Oct4, MyoD, Sox17, Nanog and Mef2c and range from chimeric TFs with added transactivation domains, designer transcription activator-like effectors to activate endogenous TFs to reprogramming TFs with rationally engineered DNA recognition principles. Possibly, applying the complete toolkit of protein design to cellular reprogramming can help to remove the hurdles that, thus far, impeded the clinical use of cells derived from reprogramming technologies.

Reprogramming cells with synthetic proteins.

Science.gov (United States)

Yang, Xiaoxiao; Malik, Vikas; Jauch, Ralf

2015-01-01

Conversion of one cell type into another cell type by forcibly expressing specific cocktails of transcription factors (TFs) has demonstrated that cell fates are not fixed and that cellular differentiation can be a two-way street with many intersections. These experiments also illustrated the sweeping potential of TFs to "read" genetically hardwired regulatory information even in cells where they are not normally expressed and to access and open up tightly packed chromatin to execute gene expression programs. Cellular reprogramming enables the modeling of diseases in a dish, to test the efficacy and toxicity of drugs in patient-derived cells and ultimately, could enable cell-based therapies to cure degenerative diseases. Yet, producing terminally differentiated cells that fully resemble their in vivocounterparts in sufficient quantities is still an unmet clinical need. While efforts are being made to reprogram cells nongenetically by using drug-like molecules, defined TF cocktails still dominate reprogramming protocols. Therefore, the optimization of TFs by protein engineering has emerged as a strategy to enhance reprogramming to produce functional, stable and safe cells for regenerative biomedicine. Engineering approaches focused on Oct4, MyoD, Sox17, Nanog and Mef2c and range from chimeric TFs with added transactivation domains, designer transcription activator-like effectors to activate endogenous TFs to reprogramming TFs with rationally engineered DNA recognition principles. Possibly, applying the complete toolkit of protein design to cellular reprogramming can help to remove the hurdles that, thus far, impeded the clinical use of cells derived from reprogramming technologies.

Reprogramming cells with synthetic proteins

Science.gov (United States)

Yang, Xiaoxiao; Malik, Vikas; Jauch, Ralf

2015-01-01

Conversion of one cell type into another cell type by forcibly expressing specific cocktails of transcription factors (TFs) has demonstrated that cell fates are not fixed and that cellular differentiation can be a two-way street with many intersections. These experiments also illustrated the sweeping potential of TFs to “read” genetically hardwired regulatory information even in cells where they are not normally expressed and to access and open up tightly packed chromatin to execute gene expression programs. Cellular reprogramming enables the modeling of diseases in a dish, to test the efficacy and toxicity of drugs in patient-derived cells and ultimately, could enable cell-based therapies to cure degenerative diseases. Yet, producing terminally differentiated cells that fully resemble their in vivo counterparts in sufficient quantities is still an unmet clinical need. While efforts are being made to reprogram cells nongenetically by using drug-like molecules, defined TF cocktails still dominate reprogramming protocols. Therefore, the optimization of TFs by protein engineering has emerged as a strategy to enhance reprogramming to produce functional, stable and safe cells for regenerative biomedicine. Engineering approaches focused on Oct4, MyoD, Sox17, Nanog and Mef2c and range from chimeric TFs with added transactivation domains, designer transcription activator-like effectors to activate endogenous TFs to reprogramming TFs with rationally engineered DNA recognition principles. Possibly, applying the complete toolkit of protein design to cellular reprogramming can help to remove the hurdles that, thus far, impeded the clinical use of cells derived from reprogramming technologies. PMID:25652623

Overview on zein protein: a promising pharmaceutical excipient in drug delivery systems and tissue engineering.

Science.gov (United States)

Labib, Gihan

2018-01-01

Natural pharmaceutical excipients have been applied extensively in the past decades owing to their safety and biocompatibility. Zein, a natural protein of plant origin offers great benefit over other synthetic polymers used in controlled drug and biomedical delivery systems. It was used in a variety of medical fields including pharmaceutical and biomedical drug targeting, vaccine, tissue engineering, and gene delivery. Being biodegradable and biocompatible, the current review focuses on the history and the medical application of zein as an attractive still promising biopolymer. Areas covered: The current review gives a broadscope on zein as a still promising protein excipient in different fields. Zein- based drug and biomedical delivery systems are discussed with special focus on current and potential application in controlled drug delivery systems, and tissue engineering. Expert opinion: Zein as a protein of natural origin can still be considered a promising polymer in the field of drug delivery systems as well as in tissue engineering. Although different researchers spotted light on zein application in different industrial fields extensively, the feasibility of its use in the field of drug delivery replenished by investigators in recent years has not yet been fully approached.

Optimal public rationing and price response.

Science.gov (United States)

Grassi, Simona; Ma, Ching-To Albert

2011-12-01

We study optimal public health care rationing and private sector price responses. Consumers differ in their wealth and illness severity (defined as treatment cost). Due to a limited budget, some consumers must be rationed. Rationed consumers may purchase from a monopolistic private market. We consider two information regimes. In the first, the public supplier rations consumers according to their wealth information (means testing). In equilibrium, the public supplier must ration both rich and poor consumers. Rationing some poor consumers implements price reduction in the private market. In the second information regime, the public supplier rations consumers according to consumers' wealth and cost information. In equilibrium, consumers are allocated the good if and only if their costs are below a threshold (cost effectiveness). Rationing based on cost results in higher equilibrium consumer surplus than rationing based on wealth. Copyright © 2011 Elsevier B.V. All rights reserved.

Exploring rationality in schizophrenia

DEFF Research Database (Denmark)

Revsbech, Rasmus; Mortensen, Erik Lykke; Owen, Gareth

2015-01-01

Background Empirical studies of rationality (syllogisms) in patients with schizophrenia have obtained different results. One study found that patients reason more logically if the syllogism is presented through an unusual content. Aims To explore syllogism-based rationality in schizophrenia. Meth...... differences became non-significant. Conclusions When taking intelligence and neuropsychological performance into account, patients with schizophrenia and controls perform similarly on syllogism tests of rationality.......Background Empirical studies of rationality (syllogisms) in patients with schizophrenia have obtained different results. One study found that patients reason more logically if the syllogism is presented through an unusual content. Aims To explore syllogism-based rationality in schizophrenia. Method...... Thirty-eight first-admitted patients with schizophrenia and 38 healthy controls solved 29 syllogisms that varied in presentation content (ordinary v. unusual) and validity (valid v. invalid). Statistical tests were made of unadjusted and adjusted group differences in models adjusting for intelligence...

Respect for rational autonomy.

Science.gov (United States)

Walker, Rebecca L

2009-12-01

The standard notion of autonomy in medical ethics does not require that autonomous choices not be irrational. The paper gives three examples of seemingly irrational patient choices and discusses how a rational autonomy analysis differs from the standard view. It then considers whether a switch to the rational autonomy view would lead to overriding more patient decisions but concludes that this should not be the case. Rather, a determination of whether individual patient decisions are autonomous is much less relevant than usually considered in determining whether health care providers must abide by these decisions. Furthermore, respect for rational autonomy entails strong positive requirements of respect for the autonomy of the person as a rational decision maker. The rationality view of autonomy is conceptually stronger than the standard view, allows for a more nuanced understanding of the practical moral calculus involved in respecting patient autonomy, and promotes positive respect for patient autonomy.

Embodying rationality

OpenAIRE

Mastrogiorgio, Antonio; Petracca, Enrico

2016-01-01

The current notions of bounded rationality in economics share distinctive features with Simon’s original notion, which still influences the theoretical and experimental research in the fields of choice, judgment, decision making, problem solving, and social cognition. All these notions of bounded rationality are in fact equally rooted in the information-processing approach to human cognition, expressing the view that reasoning is disembodied and that it can be reduced to the processing of abs...

A rational approach to heavy-atom derivative screening

International Nuclear Information System (INIS)

Joyce, M. Gordon; Radaev, Sergei; Sun, Peter D.

2010-01-01

In order to overcome the difficulties associated with the ‘classical’ heavy-atom derivatization procedure, an attempt has been made to develop a rational crystal-free heavy-atom-derivative screening method and a quick-soak derivatization procedure which allows heavy-atom compound identification. Despite the development in recent times of a range of techniques for phasing macromolecules, the conventional heavy-atom derivatization method still plays a significant role in protein structure determination. However, this method has become less popular in modern high-throughput oriented crystallography, mostly owing to its trial-and-error nature, which often results in lengthy empirical searches requiring large numbers of well diffracting crystals. In addition, the phasing power of heavy-atom derivatives is often compromised by lack of isomorphism or even loss of diffraction. In order to overcome the difficulties associated with the ‘classical’ heavy-atom derivatization procedure, an attempt has been made to develop a rational crystal-free heavy-atom derivative-screening method and a quick-soak derivatization procedure which allows heavy-atom compound identification. The method includes three basic steps: (i) the selection of likely reactive compounds for a given protein and specific crystallization conditions based on pre-defined heavy-atom compound reactivity profiles, (ii) screening of the chosen heavy-atom compounds for their ability to form protein adducts using mass spectrometry and (iii) derivatization of crystals with selected heavy-metal compounds using the quick-soak method to maximize diffraction quality and minimize non-isomorphism. Overall, this system streamlines the process of heavy-atom compound identification and minimizes the problem of non-isomorphism in phasing

A rational approach to heavy-atom derivative screening

Energy Technology Data Exchange (ETDEWEB)

Joyce, M. Gordon; Radaev, Sergei; Sun, Peter D., E-mail: psun@nih.gov [Structural Immunology Section, Laboratory of Immunogenetics, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 12441 Parklawn Drive, Rockville, Maryland 20852 (United States)

2010-04-01

In order to overcome the difficulties associated with the ‘classical’ heavy-atom derivatization procedure, an attempt has been made to develop a rational crystal-free heavy-atom-derivative screening method and a quick-soak derivatization procedure which allows heavy-atom compound identification. Despite the development in recent times of a range of techniques for phasing macromolecules, the conventional heavy-atom derivatization method still plays a significant role in protein structure determination. However, this method has become less popular in modern high-throughput oriented crystallography, mostly owing to its trial-and-error nature, which often results in lengthy empirical searches requiring large numbers of well diffracting crystals. In addition, the phasing power of heavy-atom derivatives is often compromised by lack of isomorphism or even loss of diffraction. In order to overcome the difficulties associated with the ‘classical’ heavy-atom derivatization procedure, an attempt has been made to develop a rational crystal-free heavy-atom derivative-screening method and a quick-soak derivatization procedure which allows heavy-atom compound identification. The method includes three basic steps: (i) the selection of likely reactive compounds for a given protein and specific crystallization conditions based on pre-defined heavy-atom compound reactivity profiles, (ii) screening of the chosen heavy-atom compounds for their ability to form protein adducts using mass spectrometry and (iii) derivatization of crystals with selected heavy-metal compounds using the quick-soak method to maximize diffraction quality and minimize non-isomorphism. Overall, this system streamlines the process of heavy-atom compound identification and minimizes the problem of non-isomorphism in phasing.

Selecting the Best: Evolutionary Engineering of Chemical Production in Microbes.

Science.gov (United States)

Shepelin, Denis; Hansen, Anne Sofie Lærke; Lennen, Rebecca; Luo, Hao; Herrgård, Markus J

2018-05-11

Microbial cell factories have proven to be an economical means of production for many bulk, specialty, and fine chemical products. However, we still lack both a holistic understanding of organism physiology and the ability to predictively tune enzyme activities in vivo, thus slowing down rational engineering of industrially relevant strains. An alternative concept to rational engineering is to use evolution as the driving force to select for desired changes, an approach often described as evolutionary engineering. In evolutionary engineering, in vivo selections for a desired phenotype are combined with either generation of spontaneous mutations or some form of targeted or random mutagenesis. Evolutionary engineering has been used to successfully engineer easily selectable phenotypes, such as utilization of a suboptimal nutrient source or tolerance to inhibitory substrates or products. In this review, we focus primarily on a more challenging problem-the use of evolutionary engineering for improving the production of chemicals in microbes directly. We describe recent developments in evolutionary engineering strategies, in general, and discuss, in detail, case studies where production of a chemical has been successfully achieved through evolutionary engineering by coupling production to cellular growth.

Capital Requirements and Credit Rationing

OpenAIRE

Itai Agur

2010-01-01

This paper analyzes the trade-off between financial stability and credit rationing that arises when increasing capital requirements. It extends the Stiglitz-Weiss model of credit rationing to allow for bank default. Bank capital structure then matters for lending incentives. With default and rationing endogenous, optimal capital requirements can be analyzed. Introducing bank financiers, the paper also shows that uninsured funding raises the sensitivity of rationing to capital requirements. In...

Protein engineering of subtilisins to improve stability in detergent formulations.

Science.gov (United States)

von der Osten, C; Branner, S; Hastrup, S; Hedegaard, L; Rasmussen, M D; Bisgård-Frantzen, H; Carlsen, S; Mikkelsen, J M

1993-03-01

Microbial proteases are used extensively in a large number of industrial processes and most importantly in detergent formulations facilitating the removal of proteinaceous stains. Site-directed mutagenesis has been employed in the construction of subtilisin variants with improved storage and oxidation stabilities. It is shown that in spite of significant structural homology between subtilisins subjected to protein engineering the effects of specific mutations can be quite different. Mutations that stabilize one subtilisin may destabilize another.

In vivo and in vitro protein ligation by naturally occurring and engineered split DnaE inteins.

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A Sesilja Aranko

Full Text Available BACKGROUND: Protein trans-splicing by naturally occurring split DnaE inteins is used for protein ligation of foreign peptide fragments. In order to widen biotechnological applications of protein trans-splicing, it is highly desirable to have split inteins with shorter C-terminal fragments, which can be chemically synthesized. PRINCIPAL FINDINGS: We report the identification of new functional split sites in DnaE inteins from Synechocystis sp. PCC6803 and from Nostoc punctiforme. One of the newly engineered split intein bearing C-terminal 15 residues showed more robust protein trans-splicing activity than naturally occurring split DnaE inteins in a foreign context. During the course of our experiments, we found that protein ligation by protein trans-splicing depended not only on the splicing junction sequences, but also on the foreign extein sequences. Furthermore, we could classify the protein trans-splicing reactions in foreign contexts with a simple kinetic model into three groups according to their kinetic parameters in the presence of various reducing agents. CONCLUSION: The shorter C-intein of the newly engineered split intein could be a useful tool for biotechnological applications including protein modification, incorporation of chemical probes, and segmental isotopic labelling. Based on kinetic analysis of the protein splicing reactions, we propose a general strategy to improve ligation yields by protein trans-splicing, which could significantly enhance the applications of protein ligation by protein trans-splicing.

[Rationalization and rationing at the bedside. A normative and empirical status quo analysis].

Science.gov (United States)

Strech, D

2014-02-01

The topic of bedside rationing is increasingly discussed in Germany. Further need for clarification exists for the question how bedside rationing (e.g., in the area of overcare) can be justified despite coexistent inefficiencies. This paper outlines and analyses the relationship of waste avoidance and rationing from an ethical perspective. Empirical findings regarding the status quo of bedside rationing and rationalization are presented. These normative and empirical explorations will then be further specified regarding opportunities for future physician-driven activities to tackle overuse. The self-government partners in Germany should communicate more explicitly within their communities and to the public how and with which benchmarks they aim to reduce inefficient health care (overuse) in an appropriate manner. Physician-driven activities such as the "Choosing Wisely®" initiative in the USA could provide a first step to raise the awareness for overuse among physicians as well as in the public.

Electric engineering summary

International Nuclear Information System (INIS)

Kang, Sing Eun; Park, Seong Taek; Lim, Yong Un

1975-03-01

This book is made up six parts, which deals with circuit theory about sinusoidal alternating current, basic current circuit, wave power, distorted wave, two terminal network, distributed circuit, laplace transformation and transfer function, power engineering on line, failure analysis transmission of line, substation and protection device and hydroelectric power plant, electricity machine like DC machine, electric transformer, induction machine and rectifier, electromagnetic on dielectric substance current, electromagnetic, electricity application like lighting engineering, heat transfer and electricity chemistry, industry, industry math with integer, rational number, factorization, matrix and differential.

PIPE: a protein-protein interaction prediction engine based on the re-occurring short polypeptide sequences between known interacting protein pairs

Directory of Open Access Journals (Sweden)

Greenblatt Jack

2006-07-01

Full Text Available Abstract Background Identification of protein interaction networks has received considerable attention in the post-genomic era. The currently available biochemical approaches used to detect protein-protein interactions are all time and labour intensive. Consequently there is a growing need for the development of computational tools that are capable of effectively identifying such interactions. Results Here we explain the development and implementation of a novel Protein-Protein Interaction Prediction Engine termed PIPE. This tool is capable of predicting protein-protein interactions for any target pair of the yeast Saccharomyces cerevisiae proteins from their primary structure and without the need for any additional information or predictions about the proteins. PIPE showed a sensitivity of 61% for detecting any yeast protein interaction with 89% specificity and an overall accuracy of 75%. This rate of success is comparable to those associated with the most commonly used biochemical techniques. Using PIPE, we identified a novel interaction between YGL227W (vid30 and YMR135C (gid8 yeast proteins. This lead us to the identification of a novel yeast complex that here we term vid30 complex (vid30c. The observed interaction was confirmed by tandem affinity purification (TAP tag, verifying the ability of PIPE to predict novel protein-protein interactions. We then used PIPE analysis to investigate the internal architecture of vid30c. It appeared from PIPE analysis that vid30c may consist of a core and a secondary component. Generation of yeast gene deletion strains combined with TAP tagging analysis indicated that the deletion of a member of the core component interfered with the formation of vid30c, however, deletion of a member of the secondary component had little effect (if any on the formation of vid30c. Also, PIPE can be used to analyse yeast proteins for which TAP tagging fails, thereby allowing us to predict protein interactions that are not

Revealing Atomic-Level Mechanisms of Protein Allostery with Molecular Dynamics Simulations.

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Samuel Hertig

2016-06-01

Full Text Available Molecular dynamics (MD simulations have become a powerful and popular method for the study of protein allostery, the widespread phenomenon in which a stimulus at one site on a protein influences the properties of another site on the protein. By capturing the motions of a protein's constituent atoms, simulations can enable the discovery of allosteric binding sites and the determination of the mechanistic basis for allostery. These results can provide a foundation for applications including rational drug design and protein engineering. Here, we provide an introduction to the investigation of protein allostery using molecular dynamics simulation. We emphasize the importance of designing simulations that include appropriate perturbations to the molecular system, such as the addition or removal of ligands or the application of mechanical force. We also demonstrate how the bidirectional nature of allostery-the fact that the two sites involved influence one another in a symmetrical manner-can facilitate such investigations. Through a series of case studies, we illustrate how these concepts have been used to reveal the structural basis for allostery in several proteins and protein complexes of biological and pharmaceutical interest.

Rational BRDF.

Science.gov (United States)

Pacanowski, Romain; Salazar Celis, Oliver; Schlick, Christophe; Granier, Xavier; Poulin, Pierre; Cuyt, Annie

2012-11-01

Over the last two decades, much effort has been devoted to accurately measuring Bidirectional Reflectance Distribution Functions (BRDFs) of real-world materials and to use efficiently the resulting data for rendering. Because of their large size, it is difficult to use directly measured BRDFs for real-time applications, and fitting the most sophisticated analytical BRDF models is still a complex task. In this paper, we introduce Rational BRDF, a general-purpose and efficient representation for arbitrary BRDFs, based on Rational Functions (RFs). Using an adapted parametrization, we demonstrate how Rational BRDFs offer 1) a more compact and efficient representation using low-degree RFs, 2) an accurate fitting of measured materials with guaranteed control of the residual error, and 3) efficient importance sampling by applying the same fitting process to determine the inverse of the Cumulative Distribution Function (CDF) generated from the BRDF for use in Monte-Carlo rendering.

Self-Assembly of Protein Monolayers Engineered for Improved Monoclonal Immunoglobulin G Binding

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Jeremy H. Lakey

2011-08-01

Full Text Available Bacterial outer membrane proteins, along with a filling lipid molecule can be modified to form stable self-assembled monolayers on gold. The transmembrane domain of Escherichia coli outer membrane protein A has been engineered to create a scaffold protein to which functional motifs can be fused. In earlier work we described the assembly and structure of an antibody-binding array where the Z domain of Staphylococcus aureus protein A was fused to the scaffold protein. Whilst the binding of rabbit polyclonal immunoglobulin G (IgG to the array is very strong, mouse monoclonal IgG dissociates from the array easily. This is a problem since many immunodiagnostic tests rely upon the use of mouse monoclonal antibodies. Here we describe a strategy to develop an antibody-binding array that will bind mouse monoclonal IgG with lowered dissociation from the array. A novel protein consisting of the scaffold protein fused to two pairs of Z domains separated by a long flexible linker was manufactured. Using surface plasmon resonance the self-assembly of the new protein on gold and the improved binding of mouse monoclonal IgG were demonstrated.

Microfluidic screening and whole-genome sequencing identifies mutations associated with improved protein secretion by yeast

DEFF Research Database (Denmark)

Huang, Mingtao; Bai, Yunpeng; Sjostrom, Staffan L.

2015-01-01

There is an increasing demand for biotech-based production of recombinant proteins for use as pharmaceuticals in the food and feed industry and in industrial applications. Yeast Saccharomyces cerevisiae is among preferred cell factories for recombinant protein production, and there is increasing...... interest in improving its protein secretion capacity. Due to the complexity of the secretory machinery in eukaryotic cells, it is difficult to apply rational engineering for construction of improved strains. Here we used high-throughput microfluidics for the screening of yeast libraries, generated by UV...... mutagenesis. Several screening and sorting rounds resulted in the selection of eight yeast clones with significantly improved secretion of recombinant a-amylase. Efficient secretion was genetically stable in the selected clones. We performed whole-genome sequencing of the eight clones and identified 330...

Is It Rational to Assume that Infants Imitate Rationally? A Theoretical Analysis and Critique

Science.gov (United States)

Paulus, Markus

2012-01-01

It has been suggested that preverbal infants evaluate the efficiency of others' actions (by applying a "principle of rational action") and that they imitate others' actions rationally. The present contribution presents a conceptual analysis of the claim that preverbal infants imitate rationally. It shows that this ability rests on at least three…

Mis-translation of a Computationally Designed Protein Yields an Exceptionally Stable Homodimer: Implications for Protein Engineering and Evolution.

Energy Technology Data Exchange (ETDEWEB)

Dantas, Gautam; Watters, Alexander L.; Lunde, Bradley; Eletr, Ziad; Isern, Nancy G.; Roseman, Toby; Lipfert, Jan; Doniach, Sebastian; Tompa, Martin; Kuhlman, Brian; Stoddard, Barry L.; Varani, Gabriele; Baker, David

2006-10-06

We recently used computational protein design to create an extremely stable, globular protein, Top7, with a sequence and fold not observed previously in nature. Since Top7 was created in the absence of genetic selection, it provides a rare opportunity to investigate aspects of the cellular protein production and surveillance machinery that are subject to natural selection. Here we show that a portion of the Top7 protein corresponding to the final 49 C-terminal residues is efficiently mistranslated and accumulates at high levels in E. coli. We used circular dichroism spectroscopy, size-exclusion chromatography, small-angle x-ray scattering, analytical ultra-centrifugation, and NMR spectroscopy to show that the resulting CFr protein adopts a compact, extremely-stable, obligate, symmetric, homo-dimeric structure. Based on the solution structure, we engineered an even more stable variant of CFr by disulfide-induced covalent circularisation that should be an excellent platform for design of novel functions. The accumulation of high levels of CFr exposes the high error rate of the protein translation machinery, and the rarity of correspondingly stable fragments in natural proteins implies a stringent evolutionary pressure against protein sub-fragments that can independently fold into stable structures. The symmetric self-association between two identical mistranslated CFr sub-units to generate an extremely stable structure parallels a mechanism for natural protein-fold evolution by modular recombination of stable protein sub-structures.

Translation system engineering in Escherichia coli enhances non-canonical amino acid incorporation into proteins.

Science.gov (United States)

Gan, Rui; Perez, Jessica G; Carlson, Erik D; Ntai, Ioanna; Isaacs, Farren J; Kelleher, Neil L; Jewett, Michael C

2017-05-01

The ability to site-specifically incorporate non-canonical amino acids (ncAAs) into proteins has made possible the study of protein structure and function in fundamentally new ways, as well as the bio synthesis of unnatural polymers. However, the task of site-specifically incorporating multiple ncAAs into proteins with high purity and yield continues to present a challenge. At the heart of this challenge lies the lower efficiency of engineered orthogonal translation system components compared to their natural counterparts (e.g., translation elements that specifically use a ncAA and do not interact with the cell's natural translation apparatus). Here, we show that evolving and tuning expression levels of multiple components of an engineered translation system together as a whole enhances ncAA incorporation efficiency. Specifically, we increase protein yield when incorporating multiple p-azido-phenylalanine(pAzF) residues into proteins by (i) evolving the Methanocaldococcus jannaschii p-azido-phenylalanyl-tRNA synthetase anti-codon binding domain, (ii) evolving the elongation factor Tu amino acid-binding pocket, and (iii) tuning the expression of evolved translation machinery components in a single vector. Use of the evolved translation machinery in a genomically recoded organism lacking release factor one enabled enhanced multi-site ncAA incorporation into proteins. We anticipate that our approach to orthogonal translation system development will accelerate and expand our ability to site-specifically incorporate multiple ncAAs into proteins and biopolymers, advancing new horizons for synthetic and chemical biotechnology. Biotechnol. Bioeng. 2017;114: 1074-1086. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

The Effect of Used of Soy Sauce Waste in the Ration on Yolk Chemical protein, fat and calcium of Mojosari Laying Duck

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R. D. Jayanti

2017-10-01

Full Text Available The aim of this study was to examine the effect of soy sauce waste in the ration on protein, fat and calcium content in the yolk of Mojosari laying duck. The animals used were 240 Mojosari laying ducks at 20 weeks old with the average of body weight were 1,385.0 ± 130.85 g (CV = 9.44%. Feed materials used were corn, rice bran, soybean meal, fish meal, wheat pollard, premix and soy sauce waste. The experimental design used was a completely randomized design (CRD with 4 treatment and 6 replications so that there were 24 experimental units and each consists 10 ducks. The treatments were soy sauce waste at level T0 : 0%, T1: 5%, T2: 7,5% and T3: 10%. The data were analyzed using analysis of variance (ANOVA and Significant differences were denoted as P<0.05. There was no significant effect of soy sauce waste protein, fat, and calcium content. The conclusion of this study that soy sauce waste can be used up to 10% without adverse the content of protein, fat, and calcium in Mojosari laying ducks.

Bounded Rationality and Budgeting

OpenAIRE

Ibrahim, Mukdad

2016-01-01

This article discusses the theory of bounded rationality which had been introduced by Herbert Simon in the 1950s. Simon introduced the notion of bounded rationality stating that while decision-makers strive for rationality, they are limited by the effect of the environment, their information process capacity and by the constraints on their information storage and retrieval capabilities. Moreover, this article tries to specifically blend this notion into budgeting, using the foundations of inc...

Applications of Engineered DNA-Binding Molecules Such as TAL Proteins and the CRISPR/Cas System in Biology Research

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Toshitsugu Fujita

2015-09-01

Full Text Available Engineered DNA-binding molecules such as transcription activator-like effector (TAL or TALE proteins and the clustered regularly interspaced short palindromic repeats (CRISPR and CRISPR-associated proteins (Cas (CRISPR/Cas system have been used extensively for genome editing in cells of various types and species. The sequence-specific DNA-binding activities of these engineered DNA-binding molecules can also be utilized for other purposes, such as transcriptional activation, transcriptional repression, chromatin modification, visualization of genomic regions, and isolation of chromatin in a locus-specific manner. In this review, we describe applications of these engineered DNA-binding molecules for biological purposes other than genome editing.

Apatite nano-crystalline surface modification of poly(lactide-co-glycolide) sintered microsphere scaffolds for bone tissue engineering: implications for protein adsorption.

Science.gov (United States)

Jabbarzadeh, Ehsan; Nair, Lakshmi S; Khan, Yusuf M; Deng, Meng; Laurencin, Cato T

2007-01-01

A number of bone tissue engineering approaches are aimed at (i) increasing the osteconductivity and osteoinductivity of matrices, and (ii) incorporating bioactive molecules within the scaffolds. In this study we examined the growth of a nano-crystalline mineral layer on poly(lactide-co-glycolide) (PLAGA) sintered microsphere scaffolds for tissue engineering. In addition, the influence of the mineral precipitate layer on protein adsorption on the scaffolds was studied. Scaffolds were mineralized by incubation in simulated body fluid (SBF). Scanning electron microscopy (SEM) analysis revealed that mineralized scaffolds possess a rough surface with a plate-like nanostructure covering the surface of microspheres. The results of protein adsorption and release studies showed that while the protein release pattern was similar for PLAGA and mineralized PLAGA scaffolds, precipitation of the mineral layer on PLAGA led to enhanced protein adsorption and slower protein release. Mineralization of tissue-engineered surfaces provides a method for both imparting bioactivity and controlling levels of protein adsorption and release.

Digestion and ruminal fermentation of cocoa pod silage based ration enriched by gliricidia and calliandra leaves on goats

Directory of Open Access Journals (Sweden)

Puastuti W

2015-03-01

Full Text Available In term of availability, cacao pod is potential for ruminant feed. According to its nutrients content, cacao pod can be used as feed fiber source. Protein sources materials must be added when cacao pod was ensilaged due to low protein content of this material. The aim of this study was to investigate digestibility value and end products of rumen fermentation of goat fed grass or cacao pod based ration. Randomized block design and 20 heads of lambs (16.95±2.36 kg to evaluated 5 type of rations: R (50% grass + 50% concentrate; S (50% cacao pod silage + 50% concentrate; SG (50% cacao pod-gliricidia silage + 50% concentrate; SK (50% cacao pod-calliandra silage + 50% concentrate dan SC (50% cacao pod-mixture of gliricidia-calliandra silage + 50% concentrate. Feeding trial was conducted for over 15 weeks. Measurements were taken on feed digestibility and rumen-fermentation end-products after 3 weeks of treatments. Results shows that nutrients digestibility was different significantly among the groups of treatments (P<0.05. Digestibillity of organic matter, NDF and energy of R ration was those of higher significantly (P<0.05 than those of other groups. N-ammonia of rumen from goat feed R ration was higher (P<0.05 than other groups. Total VFA and each component were different among the groups (P<0.05, however the value was similar among the groups of cacao pod silage rations. It is concluded that cacao pod silaged based rations enriched by Gliricidia and Calliandra leaves did not produce similar digestibility value and end products of rumen fermentation with grass based ration.

Rational valuations

Directory of Open Access Journals (Sweden)

Georg Spielthenner

2007-01-01

Full Text Available Valuations are ubiquitous. We may be for or against genetically modified food; we find some politicians irresponsible; we prefer Beethoven to rock ‘n’ roll or vice versa; some enjoy bird-watching while others find it boring; and we may think that we have to tighten up on green-house gas emissions. Valuing is pervasive and often we are not even aware that we are valuing. However, many of ourvaluations are ill grounded and rationally defective. They are frequently based on misinformation, sloppy thinking, prejudice, and are biased in many ways as psychological research shows. For this reason there is widespread agreement among phi-losophers that we need an account of substantive valuational rationality, both for the theory of practical reasoning and for ethics as well. My main objectin this paper is to outline such an account and to present a principle that allows a non-technical rational criticism of valuations

Rational emotions.

Science.gov (United States)

Meshulam, Meir; Winter, Eyal; Ben-Shakhar, Gershon; Aharon, Itzhak

2012-01-01

We present here the concept of rational emotions: Emotions may be directly controlled and utilized in a conscious, analytic fashion, enabling an individual to size up a situation, to determine that a certain "mental state" is strategically advantageous and adjust accordingly. Building on the growing body of literature recognizing the vital role of emotions in determining decisions, we explore the complementary role of rational choice in choosing emotional states. Participants played the role of "recipient" in the dictator game, in which an anonymous "dictator" decides how to split an amount of money between himself and the recipient. A subset of recipients was given a monetary incentive to be angry at low-split offers. That subset demonstrated increased physiological arousal at low offers relative to high offers as well as more anger than other participants. These results provide a fresh outlook on human decision-making and contribute to the continuing effort to build more complete models of rational behavior.

Models for Rational Number Bases

Science.gov (United States)

Pedersen, Jean J.; Armbruster, Frank O.

1975-01-01

This article extends number bases to negative integers, then to positive rationals and finally to negative rationals. Methods and rules for operations in positive and negative rational bases greater than one or less than negative one are summarized in tables. Sample problems are explained and illustrated. (KM)

Are security analysts rational? a literature review

OpenAIRE

Peixinho, Rúben; Coelho, Luís; Taffler, Richard J.

2005-01-01

Rational choice theory and bounded rationality constitute the basis for the discussion in several areas regarding human rationality. In finance, this discussion has been made between traditional finance and behavioural finance approach, which have different perspectives concerning market agents’ rationality. This paper reviews several studies addressing rationality among security analysts. The analysis shows that analysts’systematic optimism seems to be inconsistent with rationality....

Homology modeling and protein engineering of alkane monooxygenase in Burkholderia thailandensis MSMB121: in silico insights.

Science.gov (United States)

Jain, Chakresh Kumar; Gupta, Money; Prasad, Yamuna; Wadhwa, Gulshan; Sharma, Sanjeev Kumar

2014-07-01

The degradation of hydrocarbons plays an important role in the eco-balancing of petroleum products, pesticides and other toxic products in the environment. The degradation of hydrocarbons by microbes such as Geobacillus thermodenitrificans, Burkhulderia, Gordonia sp. and Acinetobacter sp. has been studied intensively in the literature. The present study focused on the in silico protein engineering of alkane monooxygenase (ladA)-a protein involved in the alkane degradation pathway. We demonstrated the improvement in substrate binding energy with engineered ladA in Burkholderia thailandensis MSMB121. We identified an ortholog of ladA monooxygenase found in B. thailandensis MSMB121, and showed it to be an enzyme involved in an alkane degradation pathway studied extensively in Geobacillus thermodenitrificans. Homology modeling of the three-dimensional structure of ladA was performed with a crystal structure (protein databank ID: 3B9N) as a template in MODELLER 9v11, and further validated using PROCHECK, VERIFY-3D and WHATIF tools. Specific amino acids were substituted in the region corresponding to amino acids 305-370 of ladA protein, resulting in an enhancement of binding energy in different alkane chain molecules as compared to wild protein structures in the docking experiments. The substrate binding energy with the protein was calculated using Vina (Implemented in VEGAZZ). Molecular dynamics simulations were performed to study the dynamics of different alkane chain molecules inside the binding pockets of wild and mutated ladA. Here, we hypothesize an improvement in binding energies and accessibility of substrates towards engineered ladA enzyme, which could be further facilitated for wet laboratory-based experiments for validation of the alkane degradation pathway in this organism.

The influence of nitrogen supplementation on microbial protein synthesis on water-buffaloes

International Nuclear Information System (INIS)

Abidin, Zainal; Hendratno, C.; Suharjono; Rustam, B.

1982-01-01

This work was carried out to observe the effects of nitrogen supplementation from urea and soybean meal on microbial protein synthesis, and other parameters of rumen functions of the waterbuffalo. Four rations were given to four water-buffaloes assigned in 4x4 latin square design. Ration A consisted of local grass+0% urea, ration B local grass+0.7% urea, ration C local grass+1.4% urea and ration D local grass+8.5% soybean meal. The result indicated that microbial protein synthesis was significantly affected (P/0.05) by the supplementation of urea, and the utilization of N in ration B was more efficient compared to the other rations. The ammonia concentration in the rumen fluid also increased (P/0.05) as a result of urea supplementation. However, no changes were found in the total volatile fatty acids production and total protozoal counts. An increased (P/0.05) of pH in the rumen fluid was also observed in the rations B and C. (author)

Engineer medium and feed for modulating N-glycosylation of recombinant protein production in CHO cell culture

DEFF Research Database (Denmark)

Fan, Yuzhou; Kildegaard, Helene Faustrup; Andersen, Mikael Rørdam

2017-01-01

Chinese hamster ovary (CHO) cells have become the primary expression system for the production of complex recombinant proteins due to their long-term success in industrial scale production and generating appropriate protein N-glycans similar to that of humans. Control and optimization of protein N......-glycosylation is crucial, as the structure of N-glycans can largely influence both biological and physicochemical properties of recombinant proteins. Protein N-glycosylation in CHO cell culture can be controlled and tuned by engineering medium, feed, culture process, as well as genetic elements of the cell...

A fluorescent cassette-based strategy for engineering multiple domain fusion proteins

Directory of Open Access Journals (Sweden)

Khorchid Ahmad

2003-07-01

Full Text Available Abstract Background The engineering of fusion proteins has become increasingly important and most recently has formed the basis of many biosensors, protein purification systems, and classes of new drugs. Currently, most fusion proteins consist of three or fewer domains, however, more sophisticated designs could easily involve three or more domains. Using traditional subcloning strategies, this requires micromanagement of restriction enzymes sites that results in complex workaround solutions, if any at all. Results Therefore, to aid in the efficient construction of fusion proteins involving multiple domains, we have created a new expression vector that allows us to rapidly generate a library of cassettes. Cassettes have a standard vector structure based on four specific restriction endonuclease sites and using a subtle property of blunt or compatible cohesive end restriction enzymes, they can be fused in any order and number of times. Furthermore, the insertion of PCR products into our expression vector or the recombination of cassettes can be dramatically simplified by screening for the presence or absence of fluorescence. Conclusions Finally, the utility of this new strategy was demonstrated by the creation of basic cassettes for protein targeting to subcellular organelles and for protein purification using multiple affinity tags.

Programming molecular self-assembly of intrinsically disordered proteins containing sequences of low complexity

Science.gov (United States)

Simon, Joseph R.; Carroll, Nick J.; Rubinstein, Michael; Chilkoti, Ashutosh; López, Gabriel P.

2017-06-01

Dynamic protein-rich intracellular structures that contain phase-separated intrinsically disordered proteins (IDPs) composed of sequences of low complexity (SLC) have been shown to serve a variety of important cellular functions, which include signalling, compartmentalization and stabilization. However, our understanding of these structures and our ability to synthesize models of them have been limited. We present design rules for IDPs possessing SLCs that phase separate into diverse assemblies within droplet microenvironments. Using theoretical analyses, we interpret the phase behaviour of archetypal IDP sequences and demonstrate the rational design of a vast library of multicomponent protein-rich structures that ranges from uniform nano-, meso- and microscale puncta (distinct protein droplets) to multilayered orthogonally phase-separated granular structures. The ability to predict and program IDP-rich assemblies in this fashion offers new insights into (1) genetic-to-molecular-to-macroscale relationships that encode hierarchical IDP assemblies, (2) design rules of such assemblies in cell biology and (3) molecular-level engineering of self-assembled recombinant IDP-rich materials.

A model for evaluating beef cattle rations considering effects of ruminal fiber mass

OpenAIRE

Henrique,Douglas Sampaio; Lana,Rogério de Paula; Vieira,Ricardo Augusto Mendonça; Fontes,Carlos Augusto de Alencar; Botelho,Mosar Faria

2011-01-01

A mathematical model based on Cornell Net Carbohydrate and Protein System (CNCPS) was developed and adapted in order to evaluate beef cattle rations at tropical climate conditions. The presented system differs from CNCPS in the modeling of insoluble particles' digestion and passage kinetics, which enabled the estimation of fiber mass in rumen and its effects on animal performance. The equations used to estimate metabolizable protein and net energy requirements for gain, net energy requirement...

P185-M Protein Identification and Validation of Results in Workflows that Integrate over Various Instruments, Datasets, Search Engines

Science.gov (United States)

Hufnagel, P.; Glandorf, J.; Körting, G.; Jabs, W.; Schweiger-Hufnagel, U.; Hahner, S.; Lubeck, M.; Suckau, D.

2007-01-01

Analysis of complex proteomes often results in long protein lists, but falls short in measuring the validity of identification and quantification results on a greater number of proteins. Biological and technical replicates are mandatory, as is the combination of the MS data from various workflows (gels, 1D-LC, 2D-LC), instruments (TOF/TOF, trap, qTOF or FTMS), and search engines. We describe a database-driven study that combines two workflows, two mass spectrometers, and four search engines with protein identification following a decoy database strategy. The sample was a tryptically digested lysate (10,000 cells) of a human colorectal cancer cell line. Data from two LC-MALDI-TOF/TOF runs and a 2D-LC-ESI-trap run using capillary and nano-LC columns were submitted to the proteomics software platform ProteinScape. The combined MALDI data and the ESI data were searched using Mascot (Matrix Science), Phenyx (GeneBio), ProteinSolver (Bruker and Protagen), and Sequest (Thermo) against a decoy database generated from IPI-human in order to obtain one protein list across all workflows and search engines at a defined maximum false-positive rate of 5%. ProteinScape combined the data to one LC-MALDI and one LC-ESI dataset. The initial separate searches from the two combined datasets generated eight independent peptide lists. These were compiled into an integrated protein list using the ProteinExtractor algorithm. An initial evaluation of the generated data led to the identification of approximately 1200 proteins. Result integration on a peptide level allowed discrimination of protein isoforms that would not have been possible with a mere combination of protein lists.

Combining rational and random strategies in β-glucosidase Zm-p60.1 protein library construction.

Directory of Open Access Journals (Sweden)

Dušan Turek

Full Text Available Saturation mutagenesis is a cornerstone technique in protein engineering because of its utility (in conjunction with appropriate analytical techniques for assessing effects of varying residues at selected positions on proteins' structures and functions. Site-directed mutagenesis with degenerate primers is the simplest and most rapid saturation mutagenesis technique. Thus, it is highly appropriate for assessing whether or not variation at certain sites is permissible, but not necessarily the most time- and cost-effective technique for detailed assessment of variations' effects. Thus, in the presented study we applied the technique to randomize position W373 in β-glucosidase Zm-p60.1, which is highly conserved among β-glucosidases. Unexpectedly, β-glucosidase activity screening of the generated variants showed that most variants were active, although they generally had significantly lower activity than the wild type enzyme. Further characterization of the library led us to conclude that a carefully selected combination of randomized codon-based saturation mutagenesis and site-directed mutagenesis may be most efficient, particularly when constructing and investigating randomized libraries with high fractions of positive hits.

Combining rational and random strategies in β-glucosidase Zm-p60.1 protein library construction.

Science.gov (United States)

Turek, Dušan; Klimeš, Pavel; Mazura, Pavel; Brzobohatý, Břetislav

2014-01-01

Saturation mutagenesis is a cornerstone technique in protein engineering because of its utility (in conjunction with appropriate analytical techniques) for assessing effects of varying residues at selected positions on proteins' structures and functions. Site-directed mutagenesis with degenerate primers is the simplest and most rapid saturation mutagenesis technique. Thus, it is highly appropriate for assessing whether or not variation at certain sites is permissible, but not necessarily the most time- and cost-effective technique for detailed assessment of variations' effects. Thus, in the presented study we applied the technique to randomize position W373 in β-glucosidase Zm-p60.1, which is highly conserved among β-glucosidases. Unexpectedly, β-glucosidase activity screening of the generated variants showed that most variants were active, although they generally had significantly lower activity than the wild type enzyme. Further characterization of the library led us to conclude that a carefully selected combination of randomized codon-based saturation mutagenesis and site-directed mutagenesis may be most efficient, particularly when constructing and investigating randomized libraries with high fractions of positive hits.

Limited rationality and strategic interaction

DEFF Research Database (Denmark)

Fehr, Ernst; Tyran, Jean-Robert

2008-01-01

Much evidence suggests that people are heterogeneous with regard to their abilities to make rational, forward-looking decisions. This raises the question as to when the rational types are decisive for aggregate outcomes and when the boundedly rational types shape aggregate results. We examine...... this question in the context of a long-standing and important economic problem: the adjustment of nominal prices after an anticipated monetary shock. Our experiments suggest that two types of bounded rationality-money illusion and anchoring-are important behavioral forces behind nominal inertia. However......, depending on the strategic environment, bounded rationality has vastly different effects on aggregate price adjustment. If agents' actions are strategic substitutes, adjustment to the new equilibrium is extremely quick, whereas under strategic complementarity, adjustment is both very slow and associated...

De Novo Metabolic Engineering and the Promise of Synthetic DNA

Science.gov (United States)

Klein-Marcuschamer, Daniel; Yadav, Vikramaditya G.; Ghaderi, Adel; Stephanopoulos, Gregory N.

The uncertain price and tight supply of crude oil and the ever-increasing demand for clean energy have prompted heightened attention to the development of sustainable fuel technologies that ensure continued economic development while maintaining stewardship of the environment. In the face of these enormous challenges, biomass has emerged as a viable alternative to petroleum for the production of energy, chemicals, and materials owing to its abundance, inexpensiveness, and carbon-neutrality. Moreover, the immense ease and efficiency of biological systems at converting biomass-derived feedstocks into fuels, chemicals, and materials has generated renewed interest in biotechnology as a replacement for traditional chemical processes. Aided by the ever-expanding repertoire of microbial genetics and plant biotechnology, improved understanding of gene regulation and cellular metabolism, and incessantly accumulating gene and protein data, scientists are now contemplating engineering microbial cell factories to produce fuels, chemical feedstocks, polymers and pharmaceuticals in an economically and environmentally sustainable way. This goal resonates with that of metabolic engineering - the improvement of cellular properties through the intelligent design, rational modification, or directed evolution of biochemical pathways, and arguably, metabolic engineering seems best positioned to achieve the concomittant goals of environmental stewardship and economic prolificity.

THE EFFECTS OF SYNCHRONIZATION OF CARBOHYDRATE AND PROTEIN SUPPLY IN SUGARCANE BAGASSE BASED RATION ON BODY COMPOSITION OF SHEEP

Directory of Open Access Journals (Sweden)

N. E. Wati

2016-03-01

Full Text Available The objective of this research was to study the effects of synchronization of carbohydrate and protein supply in sugarcane bagasse based ration on the body composition of sheep. The study was consisted of two steps of experiment. The first step of experiment used two rumen cannulated adult rams to create formulation of three diets with different synchronization index, namely 0.37; 0.50 and 0.63 respectively. The experimental diets were designed to be iso-energy, iso-nitrogenous and iso-neutral detergent fibre (iso-NDF. The second step of experiment was to determine the body composition of sheep fed the experimental diets, which were created in the first experiment. The body composition of fifteen rams were determined on week 0; 4; and 8 of experimental period, these were accomplished using the technique of urea dilution. The alteration of synchronization index did not affect on feed intake, ratio of ruminal acetate to propionate and serum glucose concentration, but dry matter (DM digestibility was affected (P<0.05 by the treatment of synchronization index in the diet. The alteration of synchronization index in the diet did not affect on the percentage of body protein, fat and water significantly, though body weight of sheep gained slightly during the experimental period.

Interpolation of rational matrix functions

CERN Document Server

Ball, Joseph A; Rodman, Leiba

1990-01-01

This book aims to present the theory of interpolation for rational matrix functions as a recently matured independent mathematical subject with its own problems, methods and applications. The authors decided to start working on this book during the regional CBMS conference in Lincoln, Nebraska organized by F. Gilfeather and D. Larson. The principal lecturer, J. William Helton, presented ten lectures on operator and systems theory and the interplay between them. The conference was very stimulating and helped us to decide that the time was ripe for a book on interpolation for matrix valued functions (both rational and non-rational). When the work started and the first partial draft of the book was ready it became clear that the topic is vast and that the rational case by itself with its applications is already enough material for an interesting book. In the process of writing the book, methods for the rational case were developed and refined. As a result we are now able to present the rational case as an indepe...

Effect of graded levels and sources of protein on scrotal ...

African Journals Online (AJOL)

Iso caloric rations (10.50 MJ/kg DM ME) were formulated using non-conventional protein source (maize offal and dry layer litter) to contain 12.11% CP, 14.96% CP, and 17.94% CP and fed to groups A, B and C respectively. Another ration was formulated using conventional protein source (maize, wheat bran, groundnut ...

History of Economic Rationalities

DEFF Research Database (Denmark)

This book concentrates upon how economic rationalities have been embedded into particular historical practices, cultures, and moral systems. Through multiple case-studies, situated in different historical contexts of the modern West, the book shows that the development of economic rationalities...... takes place in the meeting with other regimes of thought, values, and moral discourses. The book offers new and refreshing insights, ranging from the development of early economic thinking to economic aspects and concepts in the works of classical thinkers such as Thomas Hobbes, John Locke and Karl Marx......, to the role of economic reasoning in contemporary policies of art and health care. With economic rationalities as the read thread, the reader is offered a unique chance of historical self-awareness and recollection of how economic rationality became the powerful ideological and moral force that it is today....

A genetic replacement system for selection-based engineering of essential proteins

Science.gov (United States)

2012-01-01

Background Essential genes represent the core of biological functions required for viability. Molecular understanding of essentiality as well as design of synthetic cellular systems includes the engineering of essential proteins. An impediment to this effort is the lack of growth-based selection systems suitable for directed evolution approaches. Results We established a simple strategy for genetic replacement of an essential gene by a (library of) variant(s) during a transformation. The system was validated using three different essential genes and plasmid combinations and it reproducibly shows transformation efficiencies on the order of 107 transformants per microgram of DNA without any identifiable false positives. This allowed for reliable recovery of functional variants out of at least a 105-fold excess of non-functional variants. This outperformed selection in conventional bleach-out strains by at least two orders of magnitude, where recombination between functional and non-functional variants interfered with reliable recovery even in recA negative strains. Conclusions We propose that this selection system is extremely suitable for evaluating large libraries of engineered essential proteins resulting in the reliable isolation of functional variants in a clean strain background which can readily be used for in vivo applications as well as expression and purification for use in in vitro studies. PMID:22898007

Tissue-engineered matrices as functional delivery systems: adsorption and release of bioactive proteins from degradable composite scaffolds.

Science.gov (United States)

Cushnie, Emily K; Khan, Yusuf M; Laurencin, Cato T

2010-08-01

A tissue-engineered bone graft should imitate the ideal autograft in both form and function. However, biomaterials that have appropriate chemical and mechanical properties for grafting applications often lack biological components that may enhance regeneration. The concept of adding proteins such as growth factors to scaffolds has therefore emerged as a possible solution to improve overall graft design. In this study, we investigated this concept by loading porous hydroxyapatite-poly(lactide-co-glycolide) (HA-PLAGA) scaffolds with a model protein, cytochrome c, and then studying its release in a phosphate-buffered saline solution. The HA-PLAGA scaffold has previously been shown to be bioactive, osteoconductive, and to have appropriate physical properties for tissue engineering applications. The loading experiments demonstrated that the HA-PLAGA scaffold could also function effectively as a substrate for protein adsorption and release. Scaffold protein adsorptive loading (as opposed to physical entrapment within the matrix) was directly related to levels of scaffold HA-content. The HA phase of the scaffold facilitated protein retention in the matrix following incubation in aqueous buffer for periods up to 8 weeks. Greater levels of protein retention time may improve the protein's effective activity by increasing the probability for protein-cell interactions. The ability to control protein loading and delivery simply via composition of the HA-PLAGA scaffold offers the potential of forming robust functionalized bone grafts. (c) 2010 Wiley Periodicals, Inc.

Rational design of FRET-based sensor proteins

NARCIS (Netherlands)

Merkx, M.

2008-01-01

Real-time imaging of molecular events inside living cells is important for understanding the basis of physiological processes and diseases. Genetically encoded sensors that use fluorescence resonance energy transfer (FRET) between two fluorescent proteins are attractive in this respect because they

Psychology and the Rationality of Emotion.

Science.gov (United States)

Clore, Gerald L

2011-04-01

Questions addressed by recent psychological research on emotion include questions about how thought shapes emotion and how emotion, in turn, shapes thought. Research on emotion and cognition paints a somewhat different picture than that seen in traditional discussions of passion and reason. This article reviews several aspects of this research, concentrating specifically on three views of rationality: Rationality as Process, Rationality as Product, and Rationality as Outcome.

Psychology and the Rationality of Emotion*

OpenAIRE

Clore, Gerald L.

2011-01-01

Questions addressed by recent psychological research on emotion include questions about how thought shapes emotion and how emotion, in turn, shapes thought. Research on emotion and cognition paints a somewhat different picture than that seen in traditional discussions of passion and reason. This article reviews several aspects of this research, concentrating specifically on three views of rationality: Rationality as Process, Rationality as Product, and Rationality as Outcome.

Protein- protein interaction detection system using fluorescent protein microdomains

Science.gov (United States)

Waldo, Geoffrey S.; Cabantous, Stephanie

2010-02-23

The invention provides a protein labeling and interaction detection system based on engineered fragments of fluorescent and chromophoric proteins that require fused interacting polypeptides to drive the association of the fragments, and further are soluble and stable, and do not change the solubility of polypeptides to which they are fused. In one embodiment, a test protein X is fused to a sixteen amino acid fragment of GFP (.beta.-strand 10, amino acids 198-214), engineered to not perturb fusion protein solubility. A second test protein Y is fused to a sixteen amino acid fragment of GFP (.beta.-strand 11, amino acids 215-230), engineered to not perturb fusion protein solubility. When X and Y interact, they bring the GFP strands into proximity, and are detected by complementation with a third GFP fragment consisting of GFP amino acids 1-198 (strands 1-9). When GFP strands 10 and 11 are held together by interaction of protein X and Y, they spontaneous association with GFP strands 1-9, resulting in structural complementation, folding, and concomitant GFP fluorescence.

Testing bounded rationality against full rationality in job changing behavior

OpenAIRE

Contini, Bruno; Morini, Matteo

2007-01-01

In this paper we question the hypothesis of full rationality in the context of job changing behaviour, via simple econometric explorations on microdata drawn from WHIP (Worker Histories Italian Panel). Workers' performance is compared at the end of a three-year time window that starts when choices are expressed, under the accepted notion that the main driving forces of job change are future real wages and expected job quality. Bounded rationality suggests that individuals will search for new ...

Testing Bounded Rationality Against Full Rationality in Job Changing Behavior

OpenAIRE

Bruno Contini

2008-01-01

In this paper I question the hypothesis of full rationality in the context of job changing behaviour, via simple econometric explorations on microdata drawn from WHIP (Worker Histories Italian Panel). Workers’ performance is compared at the end of a three-year time window that starts when choices are expressed, under the accepted notion that the main driving forces of job change are future real wages and expected job quality. Bounded rationality suggests that individuals will search for new o...

Engineering and Application of Zinc Finger Proteins and TALEs for Biomedical Research.

Science.gov (United States)

Kim, Moon-Soo; Kini, Anu Ganesh

2017-08-01

Engineered DNA-binding domains provide a powerful technology for numerous biomedical studies due to their ability to recognize specific DNA sequences. Zinc fingers (ZF) are one of the most common DNA-binding domains and have been extensively studied for a variety of applications, such as gene regulation, genome engineering and diagnostics. Another novel DNA-binding domain known as a transcriptional activator-like effector (TALE) has been more recently discovered, which has a previously undescribed DNA-binding mode. Due to their modular architecture and flexibility, TALEs have been rapidly developed into artificial gene targeting reagents. Here, we describe the methods used to design these DNA-binding proteins and their key applications in biomedical research.

Psychology and the Rationality of Emotion*

Science.gov (United States)

Clore, Gerald L.

2014-01-01

Questions addressed by recent psychological research on emotion include questions about how thought shapes emotion and how emotion, in turn, shapes thought. Research on emotion and cognition paints a somewhat different picture than that seen in traditional discussions of passion and reason. This article reviews several aspects of this research, concentrating specifically on three views of rationality: Rationality as Process, Rationality as Product, and Rationality as Outcome. PMID:25125770

DISTRIBUTED RC NETWORKS WITH RATIONAL TRANSFER FUNCTIONS,

Science.gov (United States)

A distributed RC circuit analogous to a continuously tapped transmission line can be made to have a rational short-circuit transfer admittance and...one rational shortcircuit driving-point admittance. A subcircuit of the same structure has a rational open circuit transfer impedance and one rational ...open circuit driving-point impedance. Hence, rational transfer functions may be obtained while considering either generator impedance or load

Max Weber's Types of Rationality: Cornerstones for the Analysis of Rationalization Processes in History.

Science.gov (United States)

Kalberg, Stephen

1980-01-01

Explores rationality in Max Weber's works and identifies four types of rationality which play major roles in his writing--practical, theoretical, substantive, and formal. Implications for society and education are discussed. (DB)

Cell biology of sarcomeric protein engineering: disease modeling and therapeutic potential.

Science.gov (United States)

Thompson, Brian R; Metzger, Joseph M

2014-09-01

The cardiac sarcomere is the functional unit for myocyte contraction. Ordered arrays of sarcomeric proteins, held in stoichiometric balance with each other, respond to calcium to coordinate contraction and relaxation of the heart. Altered sarcomeric structure-function underlies the primary basis of disease in multiple acquired and inherited heart disease states. Hypertrophic and restrictive cardiomyopathies are caused by inherited mutations in sarcomeric genes and result in altered contractility. Ischemia-mediated acidosis directly alters sarcomere function resulting in decreased contractility. In this review, we highlight the use of acute genetic engineering of adult cardiac myocytes through stoichiometric replacement of sarcomeric proteins in these disease states with particular focus on cardiac troponin I. Stoichiometric replacement of disease causing mutations has been instrumental in defining the molecular mechanisms of hypertrophic and restrictive cardiomyopathy in a cellular context. In addition, taking advantage of stoichiometric replacement through gene therapy is discussed, highlighting the ischemia-resistant histidine-button, A164H cTnI. Stoichiometric replacement of sarcomeric proteins offers a potential gene therapy avenue to replace mutant proteins, alter sarcomeric responses to pathophysiologic insults, or neutralize altered sarcomeric function in disease. © 2014 Wiley Periodicals, Inc.

The concept of rational suicide.

Science.gov (United States)

Mayo, D J

1986-05-01

Suicide has been condemned in our culture in one way or another since Augustine offered theological arguments against it in the sixth century. More recently, theological condemnation has given way to the view that suicidal behavior must always be symptomatic of emotional disturbance and mental illness. However, suicide has not always been viewed so negatively. In other times and cultures, it has been held that circumstances might befall a person in which suicide would be a perfectly rational course of action, in the same sense that any other course of action could be rational: that it could be sensible, i.e., defensible by good reasons, or that it could be in keeping with the agent's fundamental interests. Indiscriminate use of modern life-sustaining technologies has renewed interest in the possibility of rational suicide. Today proponents of rational suicide tend to equate the rationality of suicide with the competence of the decision to commit suicide.

Efficient myogenic differentiation of human adipose-derived stem cells by the transduction of engineered MyoD protein

Energy Technology Data Exchange (ETDEWEB)

Sung, Min Sun [Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon 305-806 (Korea, Republic of); Biosystems and Bioengineering Program, University of Science and Technology (UST), Daejeon 305-350 (Korea, Republic of); Mun, Ji-Young [Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon 305-806 (Korea, Republic of); Kwon, Ohsuk [Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon 305-806 (Korea, Republic of); Biosystems and Bioengineering Program, University of Science and Technology (UST), Daejeon 305-350 (Korea, Republic of); Kwon, Ki-Sun [Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon 305-806 (Korea, Republic of); Oh, Doo-Byoung, E-mail: dboh@kribb.re.kr [Korea Research Institute of Bioscience and Biotechnology (KRIBB), 125 Gwahak-ro, Yuseong-gu, Daejeon 305-806 (Korea, Republic of); Biosystems and Bioengineering Program, University of Science and Technology (UST), Daejeon 305-350 (Korea, Republic of)

2013-07-19

Highlights: •MyoD was engineered to contain protein transduction domain and endosome-disruptive INF7 peptide. •The engineered MyoD-IT showed efficient nuclear targeting through an endosomal escape by INF7 peptide. •By applying MyoD-IT, human adipose-derived stem cells (hASCs) were differentiated into myogenic cells. •hASCs differentiated by applying MyoD-IT fused to myotubes through co-culturing with mouse myoblasts. •Myogenic differentiation using MyoD-IT is a safe method without the concern of altering the genome. -- Abstract: Human adipose-derived stem cells (hASCs) have great potential as cell sources for the treatment of muscle disorders. To provide a safe method for the myogenic differentiation of hASCs, we engineered the MyoD protein, a key transcription factor for myogenesis. The engineered MyoD (MyoD-IT) was designed to contain the TAT protein transduction domain for cell penetration and the membrane-disrupting INF7 peptide, which is an improved version of the HA2 peptide derived from influenza. MyoD-IT showed greatly improved nuclear targeting ability through an efficient endosomal escape induced by the pH-sensitive membrane disruption of the INF7 peptide. By applying MyoD-IT to a culture, hASCs were efficiently differentiated into long spindle-shaped myogenic cells expressing myosin heavy chains. Moreover, these cells differentiated by an application of MyoD-IT fused to myotubes with high efficiency through co-culturing with mouse C2C12 myoblasts. Because internalized proteins can be degraded in cells without altering the genome, the myogenic differentiation of hASCs using MyoD-IT would be a safe and clinically applicable method.

Efficient myogenic differentiation of human adipose-derived stem cells by the transduction of engineered MyoD protein

International Nuclear Information System (INIS)

Sung, Min Sun; Mun, Ji-Young; Kwon, Ohsuk; Kwon, Ki-Sun; Oh, Doo-Byoung

2013-01-01

Highlights: •MyoD was engineered to contain protein transduction domain and endosome-disruptive INF7 peptide. •The engineered MyoD-IT showed efficient nuclear targeting through an endosomal escape by INF7 peptide. •By applying MyoD-IT, human adipose-derived stem cells (hASCs) were differentiated into myogenic cells. •hASCs differentiated by applying MyoD-IT fused to myotubes through co-culturing with mouse myoblasts. •Myogenic differentiation using MyoD-IT is a safe method without the concern of altering the genome. -- Abstract: Human adipose-derived stem cells (hASCs) have great potential as cell sources for the treatment of muscle disorders. To provide a safe method for the myogenic differentiation of hASCs, we engineered the MyoD protein, a key transcription factor for myogenesis. The engineered MyoD (MyoD-IT) was designed to contain the TAT protein transduction domain for cell penetration and the membrane-disrupting INF7 peptide, which is an improved version of the HA2 peptide derived from influenza. MyoD-IT showed greatly improved nuclear targeting ability through an efficient endosomal escape induced by the pH-sensitive membrane disruption of the INF7 peptide. By applying MyoD-IT to a culture, hASCs were efficiently differentiated into long spindle-shaped myogenic cells expressing myosin heavy chains. Moreover, these cells differentiated by an application of MyoD-IT fused to myotubes with high efficiency through co-culturing with mouse C2C12 myoblasts. Because internalized proteins can be degraded in cells without altering the genome, the myogenic differentiation of hASCs using MyoD-IT would be a safe and clinically applicable method

RATGRAPH: Computer Graphing of Rational Functions.

Science.gov (United States)

Minch, Bradley A.

1987-01-01

Presents an easy-to-use Applesoft BASIC program that graphs rational functions and any asymptotes that the functions might have. Discusses the nature of rational functions, graphing them manually, employing a computer to graph rational functions, and describes how the program works. (TW)

F-theory and all things rational: surveying U(1) symmetries with rational sections

International Nuclear Information System (INIS)

Lawrie, Craig; Schäfer-Nameki, Sakura; Wong, Jin-Mann

2015-01-01

We study elliptic fibrations for F-theory compactifications realizing 4d and 6d supersymmetric gauge theories with abelian gauge factors. In the fibration these U(1) symmetries are realized in terms of additional rational section. We obtain a universal characterization of all the possible U(1) charges of matter fields by determining the corresponding codimension two fibers with rational sections. In view of modelling supersymmetric Grand Unified Theories, one of the main examples that we analyze are U(1) symmetries for SU(5) gauge theories with 5̄ and 10 matter. We use a combination of constraints on the normal bundle of rational curves in Calabi-Yau three- and four-folds, as well as the splitting of rational curves in the fibers in codimension two, to determine the possible configurations of smooth rational sections. This analysis straightforwardly generalizes to multiple U(1)s. We study the flops of such fibers, as well as some of the Yukawa couplings in codimension three. Furthermore, we carry out a universal study of the U(1)-charged GUT singlets, including their KK-charges, and determine all realizations of singlet fibers. By giving vacuum expectation values to these singlets, we propose a systematic way to analyze the Higgsing of U(1)s to discrete gauge symmetries in F-theory.

"Leaky" Rationality: How Research on Behavioral Decision Making Challenges Normative Standards of Rationality.

Science.gov (United States)

Keys, Daniel J; Schwartz, Barry

2007-06-01

For more than 30 years, decision-making research has documented that people often violate various principles of rationality, some of which are so fundamental that theorists of rationality rarely bother to state them. We take these characteristics of decision making as a given but argue that it is problematic to conclude that they typically represent departures from rationality. The very psychological processes that lead to "irrational" decisions (e.g., framing, mental accounting) continue to exert their influence when one experiences the results of the decisions. That is, psychological processes that affect decisions may be said also to "leak" into one's experience. The implication is that formal principles of rationality do not provide a good enough normative standard against which to assess decision making. Instead, what is needed is a substantive theory of rationality-one that takes subjective experience seriously, considers both direct and indirect consequences of particular decisions, considers how particular decisions fit into life as a whole, and considers the effects of decisions on others. Formal principles may play a role as approximations of the substantive theory that can be used by theorists and decision makers in cases in which the formal principles can capture most of the relevant considerations and leakage into experience is negligible. © 2007 Association for Psychological Science.

Popper, Rationality and the Possibility of Social Science

Directory of Open Access Journals (Sweden)

Danny Frederick

2013-02-01

Full Text Available Social science employs teleological explanations which depend upon the rationality principle, according to which people exhibit instrumental rationality. Popper points out that people also exhibit critical rationality, the tendency to stand back from, and to question or criticise, their views. I explain how our critical rationality impugns the explanatory value of the rationality principle and thereby threatens the very possibility of social science. I discuss the relationship between instrumental and critical rationality and show how we can reconcile our critical rationality with the possibility of social science if we invoke Popper’s conception of limited rationality and his indeterminism.

Rational and evolutionary engineering approaches uncover a small set of genetic changes efficient for rapid xylose fermentation in Saccharomyces cerevisiae.

Directory of Open Access Journals (Sweden)

Soo Rin Kim

Full Text Available Economic bioconversion of plant cell wall hydrolysates into fuels and chemicals has been hampered mainly due to the inability of microorganisms to efficiently co-ferment pentose and hexose sugars, especially glucose and xylose, which are the most abundant sugars in cellulosic hydrolysates. Saccharomyces cerevisiae cannot metabolize xylose due to a lack of xylose-metabolizing enzymes. We developed a rapid and efficient xylose-fermenting S. cerevisiae through rational and inverse metabolic engineering strategies, comprising the optimization of a heterologous xylose-assimilating pathway and evolutionary engineering. Strong and balanced expression levels of the XYL1, XYL2, and XYL3 genes constituting the xylose-assimilating pathway increased ethanol yields and the xylose consumption rates from a mixture of glucose and xylose with little xylitol accumulation. The engineered strain, however, still exhibited a long lag time when metabolizing xylose above 10 g/l as a sole carbon source, defined here as xylose toxicity. Through serial-subcultures on xylose, we isolated evolved strains which exhibited a shorter lag time and improved xylose-fermenting capabilities than the parental strain. Genome sequencing of the evolved strains revealed that mutations in PHO13 causing loss of the Pho13p function are associated with the improved phenotypes of the evolved strains. Crude extracts of a PHO13-overexpressing strain showed a higher phosphatase activity on xylulose-5-phosphate (X-5-P, suggesting that the dephosphorylation of X-5-P by Pho13p might generate a futile cycle with xylulokinase overexpression. While xylose consumption rates by the evolved strains improved substantially as compared to the parental strain, xylose metabolism was interrupted by accumulated acetate. Deletion of ALD6 coding for acetaldehyde dehydrogenase not only prevented acetate accumulation, but also enabled complete and efficient fermentation of xylose as well as a mixture of glucose and

Ethics of rationing of nursing care.

Science.gov (United States)

Rooddehghan, Zahra; Yekta, Zohreh Parsa; Nasrabadi, Alireza N

2016-09-21

Rationing of various needed services, for example, nursing care, is inevitable due to unlimited needs and limited resources. Rationing of nursing care is considered an ethical issue since it requires judgment about potential conflicts between personal and professional values. The present research sought to explore aspects of rationing nursing care in Iran. This study applied qualitative content analysis, a method to explore people's perceptions of everyday life phenomena and interpret the subjective content of text data. Data collection was performed through in-depth, unstructured, face-to-face interviews with open-ended questions. The study population included Iranian nurses of all nursing positions, from clinical nurses to nurse managers. Purposive sampling was employed to select 15 female and 3 male nurses (11 clinical nurses, 3 supervisors, 1 matron, 1 nurse, and 2 members of the Nursing Council) working in hospitals of three cities in Iran. The study protocol was approved by Tehran University of Medical Sciences (91D1302870). Written informed consent was also obtained from all participants. According to the participants, rationing of nursing care consisted of two categories, that is, causes of rationing and consequences of rationing. The first category comprised three subcategories, namely, patient needs and demands, routinism, and VIP patients. The three subcategories forming the second category were missed nursing care, patient dissatisfaction, and nurses' feeling of guilt. Levels at which healthcare practices are rationed and clarity of the rationing are important structural considerations in the development of an equal, appropriate, and ethical healthcare system. Moreover, the procedure of rationing is critical as it not only influences people's lives but also reflects the values that dominate in the society. Therefore, in order to minimize the negative consequences of rationing of nursing care, further studies on the ethical dimensions of this phenomenon

Casebook on rationalization of power use

International Nuclear Information System (INIS)

1986-06-01

This book introduces the cases on rationalization of power use, which is divided into four parts. The first part refers the goal of rational use of energy and the result. The second part deals with the excellent cases on rationalization of domestic power use, which list the name of the company, hotel and factory according to the field such as building, textile and food. The third part contains the outstanding cases on rationalization of foreign power use, which were listed by the specific way to reduce electricity use each section. The fourth part is comprised of two chapters, which deals with the cases of domestic technical data and foreign technical data for rational use of energy.

Arming Technology in Yeast-Novel Strategy for Whole-cell Biocatalyst and Protein Engineering.

Science.gov (United States)

Kuroda, Kouichi; Ueda, Mitsuyoshi

2013-09-09

Cell surface display of proteins/peptides, in contrast to the conventional intracellular expression, has many attractive features. This arming technology is especially effective when yeasts are used as a host, because eukaryotic modifications that are often required for functional use can be added to the surface-displayed proteins/peptides. A part of various cell wall or plasma membrane proteins can be genetically fused to the proteins/peptides of interest to be displayed. This technology, leading to the generation of so-called "arming technology", can be employed for basic and applied research purposes. In this article, we describe various strategies for the construction of arming yeasts, and outline the diverse applications of this technology to industrial processes such as biofuel and chemical productions, pollutant removal, and health-related processes, including oral vaccines. In addition, arming technology is suitable for protein engineering and directed evolution through high-throughput screening that is made possible by the feature that proteins/peptides displayed on cell surface can be directly analyzed using intact cells without concentration and purification. Actually, novel proteins/peptides with improved or developed functions have been created, and development of diagnostic/therapeutic antibodies are likely to benefit from this powerful approach.

Single-Point Mutation with a Rotamer Library Toolkit: Toward Protein Engineering.

Science.gov (United States)

Pottel, Joshua; Moitessier, Nicolas

2015-12-28

Protein engineers have long been hard at work to harness biocatalysts as a natural source of regio-, stereo-, and chemoselectivity in order to carry out chemistry (reactions and/or substrates) not previously achieved with these enzymes. The extreme labor demands and exponential number of mutation combinations have induced computational advances in this domain. The first step in our virtual approach is to predict the correct conformations upon mutation of residues (i.e., rebuilding side chains). For this purpose, we opted for a combination of molecular mechanics and statistical data. In this work, we have developed automated computational tools to extract protein structural information and created conformational libraries for each amino acid dependent on a variable number of parameters (e.g., resolution, flexibility, secondary structure). We have also developed the necessary tool to apply the mutation and optimize the conformation accordingly. For side-chain conformation prediction, we obtained overall average root-mean-square deviations (RMSDs) of 0.91 and 1.01 Å for the 18 flexible natural amino acids within two distinct sets of over 3000 and 1500 side-chain residues, respectively. The commonly used dihedral angle differences were also evaluated and performed worse than the state of the art. These two metrics are also compared. Furthermore, we generated a family-specific library for kinases that produced an average 2% lower RMSD upon side-chain reconstruction and a residue-specific library that yielded a 17% improvement. Ultimately, since our protein engineering outlook involves using our docking software, Fitted/Impacts, we applied our mutation protocol to a benchmarked data set for self- and cross-docking. Our side-chain reconstruction does not hinder our docking software, demonstrating differences in pose prediction accuracy of approximately 2% (RMSD cutoff metric) for a set of over 200 protein/ligand structures. Similarly, when docking to a set of over 100

Metabolizable protein systems in ruminant nutrition: A review

Directory of Open Access Journals (Sweden)

Lalatendu Keshary Das

2014-08-01

Full Text Available Protein available to ruminants is supplied by both microbial and dietary sources. Metabolizable protein (MP is the true protein which is absorbed by the intestine and supplied by both microbial protein and protein which escapes degradation in the rumen; the protein which is available to the animal for maintenance, growth, fetal growth during gestation, and milk production. Thus, the concept of balancing ruminant rations basing on only dietary crude protein (CP content seems erroneous. In India, ruminant rations are still balanced for digestible CP and total digestible nutrients for protein and energy requirements, respectively. Traditional feed analysis methods such as proximate analysis and detergent analysis consider feed protein as a single unit and do not take into account of the degradation processes that occur in rumen and passage rates of feed fractions from rumen to intestine. Therefore, the protein requirement of ruminants should include not only the dietary protein source, but also the microbial CP from rumen. The MP systems consider both the factors, thus predict the protein availability more accurately and precisely. This system is aptly designed to represent the extent of protein degradation in the rumen and the synthesis of microbial protein as variable functions. Feed protein fractions, i.e., rumen degradable protein and rumen undegradable protein play vital roles in meeting protein requirements of rumen microbes and host animal, respectively. With the advent of sophisticated nutrition models such as Cornell net carbohydrate and protein system, National Research Council, Agricultural Research Council, Cornell Penn Miner Dairy and Amino Cow; ration formulation has moved from balancing diets from CP to MP, a concept that describes the protein requirements of ruminantsat intestinal level, and which is available to animals for useful purposes.

Geometric Rationalization for Freeform Architecture

KAUST Repository

Jiang, Caigui

2016-06-20

The emergence of freeform architecture provides interesting geometric challenges with regards to the design and manufacturing of large-scale structures. To design these architectural structures, we have to consider two types of constraints. First, aesthetic constraints are important because the buildings have to be visually impressive. Sec- ond, functional constraints are important for the performance of a building and its e cient construction. This thesis contributes to the area of architectural geometry. Specifically, we are interested in the geometric rationalization of freeform architec- ture with the goal of combining aesthetic and functional constraints and construction requirements. Aesthetic requirements typically come from designers and architects. To obtain visually pleasing structures, they favor smoothness of the building shape, but also smoothness of the visible patterns on the surface. Functional requirements typically come from the engineers involved in the construction process. For exam- ple, covering freeform structures using planar panels is much cheaper than using non-planar ones. Further, constructed buildings have to be stable and should not collapse. In this thesis, we explore the geometric rationalization of freeform archi- tecture using four specific example problems inspired by real life applications. We achieve our results by developing optimization algorithms and a theoretical study of the underlying geometrical structure of the problems. The four example problems are the following: (1) The design of shading and lighting systems which are torsion-free structures with planar beams based on quad meshes. They satisfy the functionality requirements of preventing light from going inside a building as shad- ing systems or reflecting light into a building as lighting systems. (2) The Design of freeform honeycomb structures that are constructed based on hex-dominant meshes with a planar beam mounted along each edge. The beams intersect without

Generalized Information Architecture for Managing Requirements in IBM?s Rational DOORS(r) Application.

Energy Technology Data Exchange (ETDEWEB)

Aragon, Kathryn M.; Eaton, Shelley M.; McCornack, Marjorie Turner; Shannon, Sharon A.

2014-12-01

When a requirements engineering effort fails to meet expectations, often times the requirements management tool is blamed. Working with numerous project teams at Sandia National Laboratories over the last fifteen years has shown us that the tool is rarely the culprit; usually it is the lack of a viable information architecture with well- designed processes to support requirements engineering. This document illustrates design concepts with rationale, as well as a proven information architecture to structure and manage information in support of requirements engineering activities for any size or type of project. This generalized information architecture is specific to IBM's Rational DOORS (Dynamic Object Oriented Requirements System) software application, which is the requirements management tool in Sandia's CEE (Common Engineering Environment). This generalized information architecture can be used as presented or as a foundation for designing a tailored information architecture for project-specific needs. It may also be tailored for another software tool. Version 1.0 4 November 201

From rational creatures to the rational Creator: ancient and patristic analogies of the “fine-tuning” argument

Directory of Open Access Journals (Sweden)

Alexey Fokin

2017-12-01

Full Text Available In this article the author deals with the proofs of the existence of God, based on the fact of the presence of rational beings in the world. These proofs can be found in Greek and Roman Classical philosophy and early Patristics, and can be viewed as an analogue of the modern “fine-tuning argument”. The author considers the origins and development of this argument in the Greek and Roman philosophy: in Socrates, Plato, Cicero, Sextus Empiricus, and especially the Stoics, who gave to the argument a logical form, based on the relationship between “the parts and the whole”: if the world as a whole produces and contains rational beings as its parts, it should also be rational or contain a rational principle that generates all its parts and governs them, just as human soul governs human body. The correction of this argument was proposed by Sextus Empiricus, who introduced the concept of "spermatic logoi", or rational principles, to explain the generation of the rational beings by the world, so that if the world contains the spermatic logoi of rational and living beings, then the world as a whole is also rational. The author also investigates the reception of the argument based on the existence of rational beings in the world in Latin Patristics: in Minucius Felix, Tertullian and especially Lactantius, in whom this argument has reached the climax of its development. At the same time it was shown, that the ancient argument has undergone a transformation from the postulation of the rationality of the world as a whole or its equation with God himself to a genuine proof of the existence of the one God who is the Creator both of the world and human beings, superior to the both as the all-powerful and transcendent Mind.

Engineered, highly reactive substrates of microbial transglutaminase enable protein labeling within various secondary structure elements.

Science.gov (United States)

Rachel, Natalie M; Quaglia, Daniela; Lévesque, Éric; Charette, André B; Pelletier, Joelle N

2017-11-01

Microbial transglutaminase (MTG) is a practical tool to enzymatically form isopeptide bonds between peptide or protein substrates. This natural approach to crosslinking the side-chains of reactive glutamine and lysine residues is solidly rooted in food and textile processing. More recently, MTG's tolerance for various primary amines in lieu of lysine have revealed its potential for site-specific protein labeling with aminated compounds, including fluorophores. Importantly, MTG can label glutamines at accessible positions in the body of a target protein, setting it apart from most labeling enzymes that react exclusively at protein termini. To expand its applicability as a labeling tool, we engineered the B1 domain of Protein G (GB1) to probe the selectivity and enhance the reactivity of MTG toward its glutamine substrate. We built a GB1 library where each variant contained a single glutamine at positions covering all secondary structure elements. The most reactive and selective variants displayed a >100-fold increase in incorporation of a recently developed aminated benzo[a]imidazo[2,1,5-cd]indolizine-type fluorophore, relative to native GB1. None of the variants were destabilized. Our results demonstrate that MTG can react readily with glutamines in α-helical, β-sheet, and unstructured loop elements and does not favor one type of secondary structure. Introducing point mutations within MTG's active site further increased reactivity toward the most reactive substrate variant, I6Q-GB1, enhancing MTG's capacity to fluorescently label an engineered, highly reactive glutamine substrate. This work demonstrates that MTG-reactive glutamines can be readily introduced into a protein domain for fluorescent labeling. © 2017 The Protein Society.

Rationing medical education | Walsh | African Health Sciences

African Journals Online (AJOL)

Even though some stakeholders in medical education might be taken aback at the prospect of rationing, the truth is that rationing has always occurred in one form or another in medical education and in healthcare more broadly. Different types of rationing exist in healthcare professional education. For example rationing may ...

Rational Design of Adjuvant for Skin Delivery: Conjugation of Synthetic β-Glucan Dectin-1 Agonist to Protein Antigen.

Science.gov (United States)

Donadei, Agnese; Gallorini, Simona; Berti, Francesco; O'Hagan, Derek T; Adamo, Roberto; Baudner, Barbara C

2015-05-04

The potential benefits of skin delivery of vaccines derive from the presence of a densely connected network of antigen presenting cells in the skin layer, most significantly represented by Langerhans cells and dermal dendritic cells. Targeting these cells by adjuvant conjugated to an antigen should result in enhanced immunogenicity of a vaccine. Since one of the most widely used adjuvants is an insoluble salt of aluminum (aluminum hydroxide) that cannot be used for skin delivery due to reactogenicity, we focused our attention on agonists of receptors present on skin dendritic cells, including the Dectin-1 receptor. β-(1-3)-glucans, which are the most abundant components of the fungal surface, are known to activate the innate immune response by interaction with the C-type lectin-like Dectin-1 receptor. In this work we identified by rational design a well-defined synthetic β-(1-3)-glucan hexasaccharide as a Dectin-1 agonist and chemically conjugated it to the genetically detoxified diphtheria toxin (CRM197) protein antigen, as a means to increase the binding to Dectin-1 receptor and to target to skin dendritic cells. We demonstrated that the in vitro activation of the receptor was significantly impacted by the presentation of the glucan on the protein carrier. In vivo results in mice showed that the conjugation of the synthetic β-(1-3)-glucan when delivered intradermally resulted in higher antibody titers in comparison to intramuscular (i.m.) immunization and was not different from subcutaneous (s.c.) delivery. These findings suggest that weak receptor binders can be turned into more potent agonists by the multivalent presentation of many ligands covalently conjugated to the protein core. Moreover, this approach is particularly valuable to increase the immunogenicity of antigens administered via skin delivery.

Effects of different sources of protein on digestive characteristics, microbial efficiency, and nutrient flow in dairy goats

Directory of Open Access Journals (Sweden)

Nivea Regina de Oliveira Felisberto

2011-10-01

Full Text Available Diets formulated with protein sources presenting different resistance to ruminal degradation were compared by evaluating ruminal parameters, production and microbial efficiency and nutrients flow to the omasum in goats. Eight rumen cannulated non-lactating, non-pregnant goats were distributed in a 4 × 4 Latin square design with two replicates. Treatments consisted of four diets where different sources of plant protein accounted for the major protein source named soybean meal, source of higher ruminal degradability, and three other sources of higher resistance of degradation: roasted soybean, corn gluten meal, and cottonseed cake. Amounts of rumen protein were similar among rations; however, flows of dry matter, protein and non-fiber carbohydrate to omasum were higher for diets with protein source with reduced rumen degradation rate. Higher values of rumen ammonia were obtained by using ration with soybean meal as major source of protein. Higher values of pH were obtained for rations with roasted soybean e cottonseed cake. Regarding kinetic of transit, similar values were found among rations. Diets with protein sources presenting reduced ruminal degradation increase nutrients flow to the omasum in goats and alter digestive parameters such as pH and ammonia without compromising bacteria growth and efficiency, which grants their use for dairy goats with similar efficiency to rations using more degradable sources of protein.

Emotional Engineers : Toward Morally Responsible Design

NARCIS (Netherlands)

Roeser, S.

2010-01-01

Engineers are normally seen as the archetype of people who make decisions in a rational and quantitative way. However, technological design is not value neutral. The way a technology is designed determines its possibilities, which can, for better or for worse, have consequences for human wellbeing.

Effects of Rational-Emotive Therapy, Rational Role Reversal, and Rational-Emotive Imagery on the Emotional Adjustment of Community Mental Health Center Patients.

Science.gov (United States)

Lipsky, Marc J.; And Others

1980-01-01

Results showed that rational-emotive therapy (RET), with the addition of rational role reversal, produced significantly better results than did relaxation training and support or no contact. This was the first study to demonstrate the efficacy of RET with multisymptomatic applicants to a community mental health center. (Author/BEF)

Argumentation, rationality, and psychology of reasoning

Directory of Open Access Journals (Sweden)

David Godden

2015-05-01

Full Text Available This paper explicates an account of argumentative rationality by articulating the common, basic idea of its nature, and then identifying a collection of assumptions inherent in it. Argumentative rationality is then contrasted with dual-process theories of reasoning and rationality prevalent in the psychology of reasoning. It is argued that argumentative rationality properly corresponds only with system-2 reasoning in dual-process theories. This result challenges the prescriptive force of argumentative norms derives if they derive at all from their descriptive accuracy of our cognitive capacities. In response, I propose an activity-based account of reasoning which retains the assumptions of argumentative rationality while recontextualizing the relationship between reasoning as a justificatory activity and the psychological states and processes underlying that activity.

Accelerated and Rational Design of Improved CHO Cell Factories

DEFF Research Database (Denmark)

Grav, Lise Marie

Recombinant production of therapeutic proteins provides huge benefits to human health and promises solutions to some of the most devastating and currently untreatable diseases in healthcare. Key to the development of new therapeutic proteins is to optimize and engineer living cells, namely cell...... of a number of novel tools is reported that aim to accelerate the construction of production cell lines for therapeutic proteins with optimal phenotypic attributes for industrial processes. Chinese hamster ovary (CHO) cells are the predominant production host for therapeutic proteins, and are the cell factory...... of interest in this thesis. The core of the thesis is revolved around the development and application of genome editing techniques that enable us to precisely engineer the genome of CHO cells by either rendering specific-targeted genes unfunctional or inserting new genes in precise genomic locations...

Evolutionary engineering for industrial microbiology.

Science.gov (United States)

Vanee, Niti; Fisher, Adam B; Fong, Stephen S

2012-01-01

Superficially, evolutionary engineering is a paradoxical field that balances competing interests. In natural settings, evolution iteratively selects and enriches subpopulations that are best adapted to a particular ecological niche using random processes such as genetic mutation. In engineering desired approaches utilize rational prospective design to address targeted problems. When considering details of evolutionary and engineering processes, more commonality can be found. Engineering relies on detailed knowledge of the problem parameters and design properties in order to predict design outcomes that would be an optimized solution. When detailed knowledge of a system is lacking, engineers often employ algorithmic search strategies to identify empirical solutions. Evolution epitomizes this iterative optimization by continuously diversifying design options from a parental design, and then selecting the progeny designs that represent satisfactory solutions. In this chapter, the technique of applying the natural principles of evolution to engineer microbes for industrial applications is discussed to highlight the challenges and principles of evolutionary engineering.

Comparative analysis of field ration for military personnel of the ukrainian army and armies of other countries worldwide

Directory of Open Access Journals (Sweden)

M. Mardar

2017-04-01

Full Text Available For the purpose of improvement of the Ukrainian nutritional standards this Article provides comparative analysis of field rations of different countries worldwide to make a proposal on improvement of food-stuff assortment in food ration for military personnel in the Armed Forces of Ukraine, Army of USA, the British Army, Army of Germany, Army of Italy, Army of Canada, Army of France, Army of Belarus, Army of Armenia. In accordance with the comparative analysis it was established that ration composition used for the Armed Forces of Ukraine military personnel lags behind developed countries of the world both in nutrition arrangement and in nutrient composition, especially in relation to assortment and variety of ration food-stuff. Moreover, a field ration is strictly unified and doesn’t consider individual needs of military personnel in calories, proteins, fats, carbohydrates, food fibers. Selection of individual field ration takes to account only age of military personnel, i. e. individual needs related to nutrition composition such as physical abilities, level of physical activity, gender, type of occupation before military conscription and etc. are not consideredThe obtained results confirms practicability of assortment products assortment included to field rations for the purpose to correct nutrition rations towards optimal balance for military efficiency of army, adaptation of military personnel to physical and psychological loads.

The Effects of 60 Days of Tray Ration Consumption in Marine Combat Engineers While Deployed on Great Inagua Island, Bahamas.

Science.gov (United States)

2000-01-01

Turkey Eggs Fruit Sausage Vegetables Spaghetti and Meatballs Potatoes BBQ Ribs Chocolate Cake Strawberry Oatmeal Western Omlet Beef Stew Chow Mein...Stir Fry Juice 2 Potatoes Rice 2 Hamburgers Eggs With Sausage 2 Eggs in General Meatballs Table 10.10. T Ration items Marines tired of at breakfast...the three test periods. RESULTS Ratinas of T and B Breakfasts Significant main effect acceptability ratings were observed between eggs (T Ration

Rational Thinking in School-Based Practice

Science.gov (United States)

Clark, Mary Kristen; Flynn, Perry

2011-01-01

Purpose: We reflect on Alan Kamhi's (2011) prologue on balancing certainty and uncertainty as it pertains to school-based practice. Method: In schools, rational thinking depends on effective team processes, much like professional learning communities. We consider the conditions that are required for rational thinking and how rational team dialogue…

Rationality in Machiavelli and in Kant

Directory of Open Access Journals (Sweden)

Vadim Chaly

2016-11-01

Full Text Available The paper contains interpretation and comparative analysis of Machiavelli’s and Kant’s conceptions on rationality as two prime examples of “realist” and “idealist” modes of agency. Kantian model of rationality is viewed as an augmentation of the Machiavellian one, not an opposition to it. To elaborate the point, Robert Aumann’s model of act-rationality and rulerationality is applied to the two philosophical models. Kantian practical reason is interpreted as an addition to Aumann’s instrumental rationality, providing rules for rules, or “rule-rule-rationality”.

Integrated engineering system for nuclear facilities building

International Nuclear Information System (INIS)

Tomura, H.; Miyamoto, A.; Futami, F.; Yasuda, S.; Ohtomo, T.

1995-01-01

In the construction of buildings for nuclear facilities in Japan, construction companies are generally in charge of the building engineering work, coordinating with plant engineering. An integrated system for buildings (PROMOTE: PROductive MOdeling system for Total nuclear Engineering) described here is a building engineering system including the entire life cycle of buildings for nuclear facilities. A Three-dimensional (3D) building model (PRO-model) is to be in the core of the system (PROMOTE). Data sharing in the PROMOTE is also done with plant engineering systems. By providing these basic technical foundations, PROMOTE is oriented toward offering rational, highquality engineering for the projects. The aim of the system is to provide a technical foundation in building engineering. This paper discusses the characteristics of buildings for nuclear facilities and the outline of the PROMOTE. (author)

Effective Entrepreneurial Choice: The Role of Rationality and Non-Rationality in Three Entrepreneurs Success Stories

OpenAIRE

Chong, Anne Michele, Siang Yoon

2007-01-01

Good entrepreneurship is important for economic growth and productivity in any modern economy. The purpose of this dissertation is to research how good entrepreneurial decisions are made. In theory, optimal or rational decision making means choosing the best alternative in response to the problem. However, in reality, people do not act rationally because they often cannot make rational choices. The reason is that people do not have enough brain power, time or resources to process the compl...

Handheld highly selective plasmonic chem/biosensor using engineered binding proteins for extreme conformational changes

Science.gov (United States)

Kosciolek, Derek J.; Sonar, Ajay; Lepak, Lori A.; Schnatz, Peter; Bendoym, Igor; Brown, Mia C.; Koder, Ronald L.; Crouse, David T.

2017-08-01

In this project we develop a handheld, portable, highly selective and sensitive chem/biosensor that has potential applications in both airborne and water-based environmental sensing. The device relies on a plasmonic chip of subwavelength-scale periodic gold rods engineered to resonate in the near infrared. The chip is functionalized with a novel class of proteins that exhibit large conformational changes upon binding to a specific target analyte. The subsequent change in local refractive index near the surface of the gold is one to two orders of magnitude greater than current conventional methods, which produces a readily measurable 5 to 10 percent difference in light transmission. This allows us to forgo traditional, bulky tabletop setups in favor of a compact form factor. Using commercially available optics to construct a transmission-based optical train, measured changes in bulk refractive index are presented here. While synthesis of binding protein efforts are focused on heme as analyte for proof of concept validation, the functionalized protein can be engineered to pair with a wide variety of analytes with minimal alterations to the plasmonic chip or device design. Such flexibility allows for this device to potentially meet the needs of first responders and health care professionals in a multitude of scenarios.

Engineered P450 biocatalysts show improved activity and regio-promiscuity in aromatic nitration.

Science.gov (United States)

Zuo, Ran; Zhang, Yi; Jiang, Chao; Hackett, John C; Loria, Rosemary; Bruner, Steven D; Ding, Yousong

2017-04-12

Nitroaromatics are among the most important and commonly used chemicals but their production often suffers from multiple unsolved challenges. We have previously described the development of biocatalytic nitration processes driven by an engineered P450 TxtE fusion construct. Herein we report the creation of improved nitration biocatalysts through constructing and characterizing fusion proteins of TxtE with the reductase domain of CYP102A1 (P450BM3, BM3R). The majority of constructs contained variable linker length while one was rationally designed for optimizing protein-protein interactions. Detailed biochemical characterization identified multiple active chimeras that showed improved nitration activity, increased coupling efficiency and higher total turnover numbers compared with TxtE. Substrate promiscuity of the most active chimera was further assessed with a substrate library. Finally, a biocatalytic nitration process was developed to nitrate 4-Me-DL-Trp. The production of both 4-Me-5-NO 2 -L-Trp and 4-Me-7-NO 2 -L-Trp uncovered remarkable regio-promiscuity of nitration biocatalysts.

Experimental investigation of protein folding and misfolding.

Science.gov (United States)

Dobson, Christopher M

2004-09-01

Newly synthesised proteins need to fold, often to intricate and close-packed structures, in order to function. The underlying mechanism by which this complex process takes place both in vitro and in vivo is now becoming understood, at least in general terms, as a result of the application of a wide range of biophysical and computational methods used in combination with the techniques of biochemistry and protein engineering. It is increasingly apparent, however, that folding is not only crucial for generating biological activity, but that it is also coupled to a wide range of processes within the cell, ranging from the trafficking of proteins to specific organelles to the regulation of cell growth and differentiation. Not surprisingly, therefore, the failure of proteins to fold appropriately, or to remain correctly folded, is associated with a large number of cellular malfunctions that give rise to disease. Misfolding, and its consequences such as aggregation, can be investigated by extending the types of techniques used to study the normal folding process. Application of these techniques is enabling the development of a unified description of the interconversion and regulation of the different conformational states available to proteins in living systems. Such a description proves a generic basis for understanding the fundamental links between protein misfolding and its associated clinical disorders, such as Alzheimer's disease and Type II diabetes, and for exploring novel therapeutic strategies directed at their prevention and treatment on a rational basis.

GREEN: A program package for docking studies in rational drug design

Science.gov (United States)

Tomioka, Nobuo; Itai, Akiko

1994-08-01

A program package, GREEN, has been developed that enables docking studies between ligand molecules and a protein molecule. Based on the structure of the protein molecule, the physical and chemical environment of the ligand-binding site is expressed as three-dimensional grid-point data. The grid-point data are used for the real-time evaluation of the protein-ligand interaction energy, as well as for the graphical representation of the binding-site environment. The interactive docking operation is facilitated by various built-in functions, such as energy minimization, energy contribution analysis and logging of the manipulation trajectory. Interactive modeling functions are incorporated for designing new ligand molecules while considering the binding-site environment and the protein-ligand interaction. As an example of the application of GREEN, a docking study is presented on the complex between trypsin and a synthetic trypsin inhibitor. The program package will be useful for rational drug design, based on the 3D structure of the target protein.

Resilin-like polypeptide-poly(ethylene gylcol) hybrid hydrogels for mechanically-demanding tissue engineering applications

Science.gov (United States)

McGann, Christopher Leland

Technological progress in the life sciences and engineering has combined with important insights in the fields of biology and material science to make possible the development of biological substitutes which aim to restore function to damaged tissue. Numerous biomimetic hydrogels have been developed with the purpose of harnessing the regenerative capacity of cells and tissue through the rational deployment of biological signals. Aided by recombinant DNA technology and protein engineering methods, a new class of hydrogel precursor, the biosynthetic protein polymer, has demonstrated great promise towards the development of highly functional tissue engineering materials. In particular, protein polymers based upon resilin, a natural protein elastomer, have demonstrated outstanding mechanical properties that would have great value in soft tissue applications. This dissertation introduces hybrid hydrogels composed of recombinant resilin-like polypeptides (RLPs) cross-linked with multi-arm PEG macromers. Two different chemical strategies were employed to form RLP-PEG hydrogels: one utilized a Michael-type addition reaction between the thiols of cysteine residues present within the RLP and vinyl sulfone moieties functionalized on a multi-arm PEG macromer; the second system cross-links a norbornene-functionalized RLP with a thiol-functionalized multi-arm PEG macromer via a photoinitiated thiol-ene step polymerization. Oscillatory rheology and tensile testing confirmed the formation of elastic, resilient hydrogels in the RLP-PEG system cross-linked via Michael-type addition. These hydrogels supported the encapsulation and culture of both human aortic adventitial fibroblasts and human mesenchymal stem cells. Additionally, these RLP-PEG hydrogels exhibited phase separation behavior during cross-linking that led to the formation of a heterogeneous microstructure. Degradation could be triggered through incubation with matrix metalloproteinase. Photocross-linking was conferred to

Guiding rational reservoir flood operation using penalty-type genetic algorithm

Science.gov (United States)

Chang, Li-Chiu

2008-06-01

SummaryReal-time flood control of a multi-purpose reservoir should consider decreasing the flood peak stage downstream and storing floodwaters for future usage during typhoon seasons. This study proposes a reservoir flood control optimization model with linguistic description of requirements and existing regulations for rational operating decisions. The approach involves formulating reservoir flood operation as an optimization problem and using the genetic algorithm (GA) as a search engine. The optimizing formulation is expressed not only by mathematical forms of objective function and constraints, but also by no analytic expression in terms of parameters. GA is used to search a global optimum of a mixture of mathematical and nonmathematical formulations. Due to the great number of constraints and flood control requirements, it is difficult to reach a solution without violating constraints. To tackle this bottleneck, the proper penalty strategy for each parameter is proposed to guide the GA searching process. The proposed approach is applied to the Shihmen reservoir in North Taiwan for finding the rational release and desired storage as a case study. The hourly historical data sets of 29 typhoon events that have hit the area in last thirty years are investigated bye the proposed method. To demonstrate the effectiveness of the proposed approach, the simplex method was performed. The results demonstrated that a penalty-type genetic algorithm could effectively provide rational hydrographs to reduce flood damage during the flood operation and to increase final storage for future usages.

IdentiPy: an extensible search engine for protein identification in shotgun proteomics.

Science.gov (United States)

Levitsky, Lev I; Ivanov, Mark V; Lobas, Anna A; Bubis, Julia A; Tarasova, Irina A; Solovyeva, Elizaveta M; Pridatchenko, Marina L; Gorshkov, Mikhail V

2018-04-23

We present an open-source, extensible search engine for shotgun proteomics. Implemented in Python programming language, IdentiPy shows competitive processing speed and sensitivity compared with the state-of-the-art search engines. It is equipped with a user-friendly web interface, IdentiPy Server, enabling the use of a single server installation accessed from multiple workstations. Using a simplified version of X!Tandem scoring algorithm and its novel ``auto-tune'' feature, IdentiPy outperforms the popular alternatives on high-resolution data sets. Auto-tune adjusts the search parameters for the particular data set, resulting in improved search efficiency and simplifying the user experience. IdentiPy with the auto-tune feature shows higher sensitivity compared with the evaluated search engines. IdentiPy Server has built-in post-processing and protein inference procedures and provides graphic visualization of the statistical properties of the data set and the search results. It is open-source and can be freely extended to use third-party scoring functions or processing algorithms, and allows customization of the search workflow for specialized applications.

Discrete Choice and Rational Inattention

DEFF Research Database (Denmark)

Fosgerau, Mogens; Melo, Emerson; de Palma, André

2017-01-01

This paper establishes a general equivalence between discrete choice and rational inattention models. Matejka and McKay (2015, AER) showed that when information costs are modelled using the Shannon entropy, the result- ing choice probabilities in the rational inattention model take the multinomial...... logit form. We show that when information costs are modelled using a class of generalized entropies, then the choice probabilities in any rational inattention model are observationally equivalent to some additive random utility discrete choice model and vice versa. This equivalence arises from convex...

Rational top and its classical r-matrix

International Nuclear Information System (INIS)

Aminov, G; Arthamonov, S; Smirnov, A; Zotov, A

2014-01-01

We construct a rational integrable system (the rational top) on a co-adjoint orbit of SL N Lie group. It is described by the Lax operator with spectral parameter and classical non-dynamical skew-symmetric r-matrix. In the case of the orbit of minimal dimension the model is gauge equivalent to the rational Calogero–Moser (CM) system. To obtain the results we represent the Lax operator of the CM model in two different factorized forms—without spectral parameter (related to the spinless case) and another one with the spectral parameter. The latter gives rise to the rational top while the first one is related to generalized Cremmer–Gervais r-matrices. The gauge transformation relating the rational top and CM model provides the classical rational version of the IRF-Vertex correspondence. From the geometrical point of view it describes the modification of SL(N,C)-bundles over degenerated elliptic curve. In view of the Symplectic Hecke Correspondence the rational top is related to the rational spin CM model. Possible applications and generalizations of the suggested construction are discussed. In particular, the obtained r-matrix defines a class of KZB equations. (paper)

Fiber sources for complete calf starter rations.

Science.gov (United States)

Murdock, F R; Wallenius, R W

1980-11-01

Complete calf starter rations containing either 1) alfalfa hay, 2) cottonseed hulls, or 3) alfalfa-beet pulp as sources of fiber were fed to Holstein heifer calves at two locations on a limited milk program from 3 days to 12 wk of age. Rations were isonitrogenous and similar in content of crude fiber and acid detergent fiber. Although growth and development were normal on all rations, calves fed the cottonseed hull ration consumed more starter and gained more body weight than calves fed the other sources of fiber. The similarity of feed efficiencies, rumen pH, and molar ratios of volatile fatty acids between rations indicated no appreciable differences in rumen development or function. The growth response of calves fed the cottonseed hull ration appeared to be a result of better ration acceptability for which no reason was evident. Calves raised at Puyallup gained more body weight than calves at Pullman, and these gains were made more efficiently. These location effects may be related to seasonal differences and greater demands for production of body heat. Although the incidence of scours was less for calves fed alfalfa hay starter, the incidence and severity of bloat were higher for that ration.

Arming Technology in Yeast—Novel Strategy for Whole-cell Biocatalyst and Protein Engineering

Directory of Open Access Journals (Sweden)

Mitsuyoshi Ueda

2013-09-01

Full Text Available Cell surface display of proteins/peptides, in contrast to the conventional intracellular expression, has many attractive features. This arming technology is especially effective when yeasts are used as a host, because eukaryotic modifications that are often required for functional use can be added to the surface-displayed proteins/peptides. A part of various cell wall or plasma membrane proteins can be genetically fused to the proteins/peptides of interest to be displayed. This technology, leading to the generation of so-called “arming technology”, can be employed for basic and applied research purposes. In this article, we describe various strategies for the construction of arming yeasts, and outline the diverse applications of this technology to industrial processes such as biofuel and chemical productions, pollutant removal, and health-related processes, including oral vaccines. In addition, arming technology is suitable for protein engineering and directed evolution through high-throughput screening that is made possible by the feature that proteins/peptides displayed on cell surface can be directly analyzed using intact cells without concentration and purification. Actually, novel proteins/peptides with improved or developed functions have been created, and development of diagnostic/therapeutic antibodies are likely to benefit from this powerful approach.

Lessons from Learning to Have Rational Expectations

OpenAIRE

Lindh, Thomas

1989-01-01

This paper reviews a growing literature investigating how economic agents may learn rational expectations. Fully rational learning requires implausible initial information assumptions, therefore some form of bounded rationality has come into focus. Such learning models often converge to rational expectations equilibria within certain bounds. Convergence analysis has been much simplified by methods from adaptive control theory. Learning stability as a correspondence principle show some promise...

Influence of energy concentration and source on the utilization of feed protein and NPN in lambs. 3

International Nuclear Information System (INIS)

Ulbrich, M.; Geissler, C.; Bassuny, S.M.; Borowiec, F.; Hoffmann, M.

1989-01-01

In an N balance experiment with male crossbreeding lambs at an age of 3-4 months four different rations were given differing in energy concentration (high > 700 EFU cattle /kg DM and low cattle /kg DM) and in the energy source (sugar, starch or crude fibre) with crude protein intake being almost equal. The rations contained 2% urea. Microbial protein synthesis in the rumen was assessed according to Roth and Kichgessner (1978) (1), Rys et al. (1975) (2) and Bickel-Baumann and Landis (1986) (3) on the basis of allantoin excretion in urine. The highest ruminal protein synthesis quotas were 868-921 mg protein N per kg LW 0.75 in (2). In (3) 723-766 mg protein N/kg LW 0.75 were synthesized. From the 15 N labelling of the supplemented urea and the excreted allantoin it could be calculated that 26-40% of the microbial protein resulted from the urea-N of the ration. Despite a high crude protein content of the ration of between 16 and 17% in the DM and a relation of NPN: pure protein of 0.95 the utilization of the NPN in the ration was relatively high but slightly lower than the utilization of pure protein. The variants with higher energy concentration showed as a tendency higher allantoin excretion in spite of slightly lower dry matter intake and a slightly higher NPN utilization than the variants with lower energy concentration. (author)

Strategy selection as rational metareasoning.

Science.gov (United States)

Lieder, Falk; Griffiths, Thomas L

2017-11-01

Many contemporary accounts of human reasoning assume that the mind is equipped with multiple heuristics that could be deployed to perform a given task. This raises the question of how the mind determines when to use which heuristic. To answer this question, we developed a rational model of strategy selection, based on the theory of rational metareasoning developed in the artificial intelligence literature. According to our model people learn to efficiently choose the strategy with the best cost-benefit tradeoff by learning a predictive model of each strategy's performance. We found that our model can provide a unifying explanation for classic findings from domains ranging from decision-making to arithmetic by capturing the variability of people's strategy choices, their dependence on task and context, and their development over time. Systematic model comparisons supported our theory, and 4 new experiments confirmed its distinctive predictions. Our findings suggest that people gradually learn to make increasingly more rational use of fallible heuristics. This perspective reconciles the 2 poles of the debate about human rationality by integrating heuristics and biases with learning and rationality. (PsycINFO Database Record (c) 2017 APA, all rights reserved).

Should informed consent be based on rational beliefs?

Science.gov (United States)

Savulescu, J; Momeyer, R W

1997-10-01

Our aim is to expand the regulative ideal governing consent. We argue that consent should not only be informed but also based on rational beliefs. We argue that holding true beliefs promotes autonomy. Information is important insofar as it helps a person to hold the relevant true beliefs. But in order to hold the relevant true beliefs, competent people must also think rationally. Insofar as information is important, rational deliberation is important. Just as physicians should aim to provide relevant information regarding the medical procedures prior to patients consenting to have those procedures, they should also assist patients to think more rationally. We distinguish between rational choice/action and rational belief. While autonomous choice need not necessarily be rational, it should be based on rational belief. The implication for the doctrine of informed consent and the practice of medicine is that, if physicians are to respect patient autonomy and help patients to choose and act more rationally, not only must they provide information, but they should care more about the theoretical rationality of their patients. They should not abandon their patients to irrationality. They should help their patients to deliberate more effectively and to care more about thinking rationally. We illustrate these arguments in the context of Jehovah's Witnesses refusing life-saving blood transfusions. Insofar as Jehovah's Witnesses should be informed of the consequences of their actions, they should also deliberate rationally about these consequences.

Engineering signal peptides for enhanced protein secretion from Lactococcus lactis.

Science.gov (United States)

Ng, Daphne T W; Sarkar, Casim A

2013-01-01

Lactococcus lactis is an attractive vehicle for biotechnological production of proteins and clinical delivery of therapeutics. In many such applications using this host, it is desirable to maximize secretion of recombinant proteins into the extracellular space, which is typically achieved by using the native signal peptide from a major secreted lactococcal protein, Usp45. In order to further increase protein secretion from L. lactis, inherent limitations of the Usp45 signal peptide (Usp45sp) must be elucidated. Here, we performed extensive mutagenesis on Usp45sp to probe the effects of both the mRNA sequence (silent mutations) and the peptide sequence (amino acid substitutions) on secretion. We screened signal peptides based on their resulting secretion levels of Staphylococcus aureus nuclease and further evaluated them for secretion of Bacillus subtilis α-amylase. Silent mutations alone gave an increase of up to 16% in the secretion of α-amylase through a mechanism consistent with relaxed mRNA folding around the ribosome binding site and enhanced translation. Targeted amino acid mutagenesis in Usp45sp, combined with additional silent mutations from the best clone in the initial screen, yielded an increase of up to 51% in maximum secretion of α-amylase while maintaining secretion at lower induction levels. The best sequence from our screen preserves the tripartite structure of the native signal peptide but increases the positive charge of the n-region. Our study presents the first example of an engineered L. lactis signal peptide with a higher secretion yield than Usp45sp and, more generally, provides strategies for further enhancing protein secretion in bacterial hosts.

Strategic study on energy-protein requirements for local sheep: 5. Ewes during lactation phase

Directory of Open Access Journals (Sweden)

I-W Mathius

2004-03-01

Full Text Available Thirty-six Javanese thin-tail ewes in the end of late pregnancy phase were set out to study the energy and crude protein requirements during the first eight-week of lactation phase. The ewes were penned individually in doors and randomly assigned to a 3 x 3 factorial arrangement, consisting of three levels of energy (low, medium and high and three levels of crude protein (low, medium and high diets with four ewes per treatment. The diets were pelleted and offered four times daily in approximately equal amount. Feed intake, nutrient digestibility, body weight and milk production were recorded. Results showed that, total lamb birth weights was not affected, but protein content on the ration treatments significantly altered (P0.05, while crude protein content on the ration highly significantly affected (P<0.01. Based on data recorded, the energy and protein requirements for ewes during lactation phase are highly significantly depended on ewes’ live weight, milk production and the ratio of energy metabolism and crude protein of the ration. It was concluded that in order to fulfil the crude protein and energy needs of the ewes during lactation phase, the ration given should contain crude protein and energy as much as 16% (based on dry matter and 13.4 MJ/kg dry matter respectively.

Product differentiation under bounded rationality

NARCIS (Netherlands)

Vermeulen, B.; Poutré, La J.A.; Kok, de A.G.; Pyka, A.; Handa, H.; Ishibuchi, H.; Ong, Y.-S.; Tan, K.-C.

2015-01-01

We study product differentiation equilibria and dynamics on the Salop circle under bounded rationality. Due to bounded rationality, firms tend to agglomerate in pairs. Upon adding a second tier of component suppliers, downstream assemblers may escape pairwise horizontal agglomeration. Moreover, we

Engineer Medium and Feed for Modulating N-Glycosylation of Recombinant Protein Production in CHO Cell Culture.

Science.gov (United States)

Fan, Yuzhou; Kildegaard, Helene Faustrup; Andersen, Mikael Rørdam

2017-01-01

Chinese hamster ovary (CHO) cells have become the primary expression system for the production of complex recombinant proteins due to their long-term success in industrial scale production and generating appropriate protein N-glycans similar to that of humans. Control and optimization of protein N-glycosylation is crucial, as the structure of N-glycans can largely influence both biological and physicochemical properties of recombinant proteins. Protein N-glycosylation in CHO cell culture can be controlled and tuned by engineering medium, feed, culture process, as well as genetic elements of the cell. In this chapter, we will focus on how to carry out experiments for N-glycosylation modulation through medium and feed optimization. The workflow and typical methods involved in the experiment process will be presented.

Rational Design of Thermally Stable Novel Biocatalytic Nanomaterials: Enzyme Stability in Restricted Spatial Dimensions

Science.gov (United States)

Mudhivarthi, Vamsi K.

Enzyme stability is of intense interest in bio-materials science as biocatalysts, and as sensing platforms. This is essentially because the unique properties of DNA, RNA, PAA can be coupled with the interesting and novel properties of proteins to produce systems with unprecedented control over their properties. In this article, the very first examples of enzyme/NA/inorganic hybrid nanomaterials and enzyme-Polyacrylic acid conjugates will be presented. The basic principles of design, synthesis and control of properties of these hybrid materials will be presented first, and this will be followed by a discussion of selected examples from our recent research findings. Data show that key properties of biological catalysts are improved by the inorganic framework especially when the catalyst is co-embedded with DNA. Several examples of such studies with various enzymes and proteins, including horseradish peroxidase (HRP), glucose oxidase (GO), cytochrome c (Cyt c), met-hemoglobin (Hb) and met-myoglobin (Mb) will be discussed. Additionally, key insights obtained by the standard methods of materials science including XRD, SEM and TEM as well as biochemical, calorimetric and spectroscopic methods will be discussed. Furthermore, improved structure and enhanced activities of the biocatalysts in specific cases will be demonstrated along with the potential stabilization mechanisms. Our hypothesis is that nucleic acids provide an excellent control over the enzyme-solid interactions as well as rational assembly of nanomaterials. These novel nanobiohybrid materials may aid in engineering more effective synthetic materials for gene-delivery, RNA-delivery and drug delivery applications.

An evaluation of rational-emotive imagery as a component of rational-emotive therapy in the treatment of test anxiety.

Science.gov (United States)

Hymen, S P; Warren, R

1978-06-01

This study evaluated the efficacy of rational-emotive imagery as a component of rational-emotive therapy in reduction of college students' test anxiety. 11 volunteers met for 6 1-hr. group treatment sessions over a 3-wk. period. After 2 initial treatment sessions subjects were randomly assigned to groups given either rational-emotive therapy with rational-emotive imagery or rational-emotive therapy without imagery. Contrary to predictions, improvement between groups on self-report and performance measures was nonsignificant. Failure to obtain differences was attributed to similarities in content of treatment sessions and short treatment time. Combined groups reported significant improvement on all dependent measures. Although the study did not yield the predicted benefits of the imagery, results lend further support to the efficacy of rational-emotive therapy procedures in the reduction of test anxiety.

Rationing in health systems: A critical review.

Science.gov (United States)

Keliddar, Iman; Mosadeghrad, Ali Mohammad; Jafari-Sirizi, Mehdi

2017-01-01

Background: It is difficult to provide health care services to all those in need of such services due to limited resources and unlimited demands. Thus, priority setting and rationing have to be applied. This study aimed at critically examining the concept of rationing in health sector and identifying its purposes, influencing factors, mechanisms, and outcomes. Methods: The critical interpretive synthesis methodology was used in this study. PubMed, Cochrane, and Proquest databases were searched using the related key words to find related documents published between 1970 and 2015. In total, 161 published reports were reviewed and included in the study. Thematic content analysis was applied for data analysis. Results: Health services rationing means restricting the access of some people to useful or potentially useful health services due to budgetary limitation. The inherent features of the health market and health services, limited resources, and unlimited needs necessitate health services rationing. Rationing can be applied in 4 levels: health care policy- makers, health care managers, health care providers, and patients. Health care rationing can be accomplished through fixed budget, benefit package, payment mechanisms, queuing, copayments, and deductibles. Conclusion: This paper enriched our understanding of health services rationing and its mechanisms at various levels and contributed to the literature by broadly conceptualizing health services rationing.

Investigations of the influence of the content of crude plant protein in the ration on the utilisation of urea in dairy cattle. 3

International Nuclear Information System (INIS)

Tarakanow, B.W.; Sommer, A.; Voigt, J.

1984-01-01

The rations contained 10.7 (I), 13.7 (II) and 17.1 (III)% plant crude protein and, after the supplementation of 150 g urea per animal and day, a total of 13.8 (I), 16.7 (II) and 20.2 (III)% crude protein in the dry matter. The starch content and the dry matter was 25.6, 19.6 and 13.4% from I to III. The urea was intraruminally infused during the feeding in the morning and the evening. In the morning feeding of each 1st measuring day it was labelled with 27.5 atom-% 15 N'. Samples were taken from the rumen up to 72 h after intake. 4, 8 and 12 hours after 15 N intake 100 ml rumen fluid contained 2.5, 3.3 and 3.0 (I), 3.1, 3.0 and 2.8 (II) and 4.1, 4.4 and 4.2 (III) g dry matter of the bacteria and 7.9, 9.0 and 5.9 (I), 3.8, 3.2 and 3.1 (II) and 2.1, 3.1 and 1.7 (III) g dry matter of the protozoa. In comparison to I the concentration of microbes in III decreased to 60-67%. The N level of the ration did not influence the N and carbohydrate content of the microbes as well as the amino acid composition of the microbial protein. Level and dynamics of 15 N incorporation differed between protozoa and bacteria. The latter were more intensively labelled and 15 N frequency increased more quickly. From I to III the maximum labelling degrees were 3.27, 3.09 and 1.98 for bacteria and 0.93, 0.82 and 0.80 atom-% 15 N excess. The apparent half life of 15 N on 10...12 h in the bacteria N and of 34...59 h in the protozoa N showed that 15 N metabolism in protozoa is distinctly slower. The 15 N frequency in the amino acids (AA) of the microbial protein revaled that urea N was included in the synthesis of all the 16 AA investigated, with big differences in the relative 15 N frequency in the AA both within the microbes and between the species of microbes. One can conclude that the decreasing utilization of 15 N from urea in comparison with that of the total N is caused by the reduction of the concentration of microbes in the rumen fluid and the relatively lower quota of the incorporation

Milk production is unaffected by replacing barley or sodium hydroxide wheat with maize cob silage in rations for dairy cows

DEFF Research Database (Denmark)

Hymøller, Lone; Hellwing, Anne Louise Frydendahl; Lund, Peter

2014-01-01

. The energy-corrected milk yield was unaffected by treatment. The fat content of the milk on the MCS ration was not different from the SHW ration, whereas it was higher on the barley ration. The protein content of the milk decreased when MCS was used in the ration compared with barley and SHW. From ruminal......Starch is an important energy-providing nutrient for dairy cows that is most commonly provided from cereal grains. However, ruminal fermentation of large amounts of easily degradable starch leads to excessive production and accumulation of volatile fatty acids (VFA). VFA not only play a vital role...... in the energy metabolism of dairy cows but are also the main cause of ruminal acidosis and depressed feed intake. The aim of the present study was to compare maize cob silage (MCS) as an energy supplement in rations for dairy cows with highly rumen-digestible rolled barley and with sodium hydroxide wheat (SHW...

Research on Bounded Rationality of Fuzzy Choice Functions

Directory of Open Access Journals (Sweden)

Xinlin Wu

2014-01-01

Full Text Available The rationality of a fuzzy choice function is a hot research topic in the study of fuzzy choice functions. In this paper, two common fuzzy sets are studied and analyzed in the framework of the Banerjee choice function. The complete rationality and bounded rationality of fuzzy choice functions are defined based on the two fuzzy sets. An assumption is presented to study the fuzzy choice function, and especially the fuzzy choice function with bounded rationality is studied combined with some rationality conditions. Results show that the fuzzy choice function with bounded rationality also satisfies some important rationality conditions, but not vice versa. The research gives supplements to the investigation in the framework of the Banerjee choice function.

Effects of Synchronization of Carbohydrate and Protein Supply in Total Mixed Ration with Korean Rice Wine Residue on Ruminal Fermentation, Nitrogen Metabolism and Microbial Protein Synthesis in Holstein Steers

Directory of Open Access Journals (Sweden)

Min Yu Piao

2012-11-01

Full Text Available Three Holstein steers in the growing phase, each with a ruminal cannula, were used to test the hypothesis that the synchronization of the hourly rate of carbohydrate and nitrogen (N released in the rumen would increase the amount of retained nitrogen for growth and thus improve the efficiency of microbial protein synthesis (EMPS. In Experiment 1, in situ degradability coefficients of carbohydrate and N in feeds including Korean rice wine residue (RWR were determined. In Experiment 2, three total mixed ration (TMR diets having different rates of carbohydrate and N release in the rumen were formulated using the in situ degradability of the feeds. All diets were made to contain similar contents of crude protein (CP and neutral detergent fiber (NDF but varied in their hourly pattern of nutrient release. The synchrony index of the three TMRs was 0.51 (LS, 0.77 (MS and 0.95 (HS, respectively. The diets were fed at a restricted level (2% of the animal’s body weight in a 3×3 Latin-square design. Synchronizing the hourly supply of energy and N in the rumen did not significantly alter the digestibility of dry matter, organic matter, crude protein, NDF or acid detergent fiber (ADF (p>0.05. The ruminal NH3-N content of the LS group at three hours after feeding was significantly higher (p0.05. In addition, the purine derivative (PD excretion in urine and microbial-N production (MN among the three groups were not significantly different (p>0.05. In conclusion, synchronizing dietary energy and N supply to the rumen did not have a major effect on nutrient digestion or microbial protein synthesis (MPS in Holstein steers.

Agent-based autonomous systems and abstraction engines: Theory meets practice

OpenAIRE

Dennis, L.A.; Aitken, J.M.; Collenette, J.; Cucco, E.; Kamali, M.; McAree, O.; Shaukat, A.; Atkinson, K.; Gao, Y.; Veres, S.M.; Fisher, M.

2016-01-01

We report on experiences in the development of hybrid autonomous systems where high-level decisions are made by a rational agent. This rational agent interacts with other sub-systems via an abstraction engine. We describe three systems we have developed using the EASS BDI agent programming language and framework which supports this architecture. As a result of these experiences we recommend changes to the theoretical operational semantics that underpins the EASS framework and present a fourth...

Crystals of Human Serum Albumin for Use in Genetic Engineering and Rational Drug Design

Science.gov (United States)

Carter, Daniel C. (Inventor)

1994-01-01

This invention pertains to crystals of serum albumin and processes for growing them. The purpose of the invention is to provide crystals of serum albumin which can be studied to determine binding sites for drugs. Form 2 crystals grow in the monoclinic space P2(sub 1), and possesses the following unit cell constraints: a = 58.9 +/- 7, b = 88.3 +/- 7, c = 60.7 +/- 7, Beta = 101.0 +/- 2 degrees. One advantage of the invention is that it will allow rational drug design

Using Rational-Emotive Therapy to Prevent Classroom Problems.

Science.gov (United States)

Webber, Jo; Coleman, Maggie

1988-01-01

Teachers are encouraged to utilize rational-emotive therapy to prevent and deal with classroom behavior problems. Rational-emotive therapy is defined, the ABC model of rational thinking briefly explained, types of irrational thinking identified, and suggestions for becoming a rational thinker are offered. Classroom examples are given. (DB)

Utilization of by-products and locally available feedstuffs in buffalo rations in Bangladesh

International Nuclear Information System (INIS)

Akbar, M.A.; Tareque, A.M.

1990-01-01

A series of experiments was conducted to investigate methods of urea incorporation into rice straw based rations, and to determine the optimum level of wheat bran, rice polishings, fish meal and/or broken rice supplementation of rice straw/urea based diets for growing buffaloes. The results indicate that the utilization of rice straw can be improved by ensiling with urea, soaking with urea-water, or supplementing with urea/molasses blocks, fish meal, rice polishings, or a combination of other bypass protein and energy feeds. For example, ensiling with urea increased the dry matter (DM) intake and body weight gain by 25 and 45%, respectively, compared with untreated straw. Fish meal, in combination with wheat bran, rice polishings or broken rice as supplementary sources of protein and energy, gave varying levels of body weight gain and DM intake, although, in general, all supplements improved straw utilization. Soaking of straw in urea-water is an easy and suitable method of incorporating urea into straw based rations. Supplementation of soaked straw with fish meal or rice polishings increased the live weight gain in buffalo heifers. However, the economic feasibility of using fish meal and broken rice as supplementary sources of protein and energy should be considered before these results are extended to the small farmers. (author). 12 refs, 5 tabs

STEEP STREAMS - Solid Transport Evaluation and Efficiency in Prevention: Sustainable Techniques of Rational Engineering and Advanced MethodS

Science.gov (United States)

Armanini, Aronne; Cardoso, Antonio H.; Di Baldassarre, Giuliano; Bellin, Alberto; Breinl, Korbinian; Canelas, Ricardo B.; Larcher, Michele; Majone, Bruno; Matos, Jorges; Meninno, Sabrina; Nucci, Elena; Rigon, Riccardo; Rosatti, Giorgio; Zardi, Dino

2017-04-01

The STEEP STREAMS (Solid Transport Evaluation and Efficiency in Prevention: Sustainable Techniques of Rational Engineering and Advanced MethodS) project consists of a collaboration among the Universities of Trento, Uppsala and Lisbon, who joined in a consortium within the ERANET Water JPI call WaterWorks2014. The aim of the project is to produce new rational criteria for the design of protection works against debris flows, a phenomenon consisting in hyper-concentrated flows of water and sediments, classified as catastrophic events typical of small mountainous basins (area triggered by intense rainstorms. Such events are non-stationary phenomena that arise in a very short time, and their recurrence is rather difficult to determine. Compared to flash floods, they are more difficult to anticipate, mostly since they are triggered by convective precipitation events, posing a higher risk of damage and even loss of human lives. These extreme events occur almost annually across Europe, though the formal return period in an exposed site is much larger. Recently, an increase in intensity and frequency of small-scale storm events, leading to extreme solid transport in steep channels, are recognized as one of the effects of climate change. In this context, one of the key challenges of this project is the use of comparatively coarse RCM projections to the small catchments examined in STEEP STREAMS. Given these changes, conventional protection works and their design criteria may not suffice to provide adequate levels of protection to human life and urban settlements. These structures create a storage area upstream the alluvial fans and the settlements, thereby reducing the need of channelization in areas often constrained by urban regulations. To optimize the lamination, and in particular to reduce the peak of solid mass flux, it is necessary that the deposition basin is controlled by a slit check dam, capable of inducing a controlled sedimentation of the solid mas flux. In

[Rationalization, rationing, prioritization: terminology and ethical approaches to the allocation of limited resources in hematology/oncology].

Science.gov (United States)

Winkler, Eva

2011-01-01

The field of oncology with its numerous high-priced innovations contributes considerably to the fact that medical progress is expensive. Additionally, due to the demographic changes and the increasing life expectancy, a growing number of cancer patients want to profit from this progress. Since resources are limited also in the health system, the fair distribution of the available resources urgently needs to be addressed. Dealing with scarcity is a typical problem in the domain of justice theory; therefore, this article first discusses different strategies to manage limited resources: rationalization, rationing, and prioritization. It then presents substantive as well as procedural criteria that assist in the just distribution of effective health benefits. There are various strategies to reduce the utilization of limited resources: Rationalization means that efficiency reserves are being exhausted; by means of rationing, effective health benefits are withheld due to cost considerations. Rationing can occur implicitly and thus covertly, e.g. through budgeting or the implementation of waiting periods, or explicitly, through transparent rules or policies about healthcare coverage. Ranking medical treatments according to their importance (prioritization) is often a prerequisite for rationing decisions. In terms of requirements of justice, both procedural and substantive criteria (e.g. equality, urgency, benefit) are relevant for the acceptance and quality of a decision to limit access to effective health benefits. Copyright © 2011 S. Karger AG, Basel.

Use of Sunflower Meal as a Substitute for Cottonseed Meal in Rations of Growing Lambs

International Nuclear Information System (INIS)

Saleh, S.A.; EI-Fouly, H.A.

2008-01-01

Eighteen growing male lambs with an average of 3 months old were randomly divided into three equal groups. Animals of each group were kept in separate shaded pen and fed one of the three tested rations as follows: 1st group fed ration one (RI): (CFM) concentrate feed mixture contained 30% cottonseed meal (CSM) + 0.00 sunflower meal (SFM). 2nd group fed ration two (RII): CFM contained 15% CSM + 15% SFM. 3rd group fed ration three (RIll): CFM contained 00.0% CSM + 30% SFM. Wheat straw was offered to the 3 experiment groups ad libitum. Lambs were weighed at the beginning of the experimental period then at two weeks intervals. At the end of the experimental period, four animals from each group were used to evaluate the nutrients digestibility and nutritive value of the experimental rations. Blood samples were taken each three weeks before feeding animals. The present study showed that lambs fed RII and RIll had significantly higher CP and EE digestibility values compared to those fed RI. % Nitrogen balance was currently higher for RIll than RI and RII. No significant differences among the tested blood serum parameters for the experimental lambs except for total protein, AST, triglycerides and cholesterol values. Average daily weight gain (ADG) and feed efficiency were better for RII and RIll than RI

Rational use of cognitive resources: levels of analysis between the computational and the algorithmic.

Science.gov (United States)

Griffiths, Thomas L; Lieder, Falk; Goodman, Noah D

2015-04-01

Marr's levels of analysis-computational, algorithmic, and implementation-have served cognitive science well over the last 30 years. But the recent increase in the popularity of the computational level raises a new challenge: How do we begin to relate models at different levels of analysis? We propose that it is possible to define levels of analysis that lie between the computational and the algorithmic, providing a way to build a bridge between computational- and algorithmic-level models. The key idea is to push the notion of rationality, often used in defining computational-level models, deeper toward the algorithmic level. We offer a simple recipe for reverse-engineering the mind's cognitive strategies by deriving optimal algorithms for a series of increasingly more realistic abstract computational architectures, which we call "resource-rational analysis." Copyright © 2015 Cognitive Science Society, Inc.

Differential Rationality and Personal Development.

Science.gov (United States)

Fincher, Cameron

This publication discusses differential rationality; it asserts that the development of institutions, professions, and individuals involves the differentiation of forms and styles of thinking and knowing that are, in various ways, idiosyncratic. Based on this understanding, differential rationality can be seen as a developmental construct that…

SCOWLP classification: Structural comparison and analysis of protein binding regions

Directory of Open Access Journals (Sweden)

Anders Gerd

2008-01-01

Full Text Available Abstract Background Detailed information about protein interactions is critical for our understanding of the principles governing protein recognition mechanisms. The structures of many proteins have been experimentally determined in complex with different ligands bound either in the same or different binding regions. Thus, the structural interactome requires the development of tools to classify protein binding regions. A proper classification may provide a general view of the regions that a protein uses to bind others and also facilitate a detailed comparative analysis of the interacting information for specific protein binding regions at atomic level. Such classification might be of potential use for deciphering protein interaction networks, understanding protein function, rational engineering and design. Description Protein binding regions (PBRs might be ideally described as well-defined separated regions that share no interacting residues one another. However, PBRs are often irregular, discontinuous and can share a wide range of interacting residues among them. The criteria to define an individual binding region can be often arbitrary and may differ from other binding regions within a protein family. Therefore, the rational behind protein interface classification should aim to fulfil the requirements of the analysis to be performed. We extract detailed interaction information of protein domains, peptides and interfacial solvent from the SCOWLP database and we classify the PBRs of each domain family. For this purpose, we define a similarity index based on the overlapping of interacting residues mapped in pair-wise structural alignments. We perform our classification with agglomerative hierarchical clustering using the complete-linkage method. Our classification is calculated at different similarity cut-offs to allow flexibility in the analysis of PBRs, feature especially interesting for those protein families with conflictive binding regions

Rational reconstructions of modern physics

CERN Document Server

Mittelstaedt, Peter

2013-01-01

Newton’s classical physics and its underlying ontology are loaded with several metaphysical hypotheses that cannot be justified by rational reasoning nor by experimental evidence. Furthermore, it is well known that some of these hypotheses are not contained in the great theories of Modern Physics, such as the theory of Special Relativity and Quantum Mechanics. This book shows that, on the basis of Newton’s classical physics and by rational reconstruction, the theory of Special Relativity as well as Quantum Mechanics can be obtained by partly eliminating or attenuating the metaphysical hypotheses. Moreover, it is shown that these reconstructions do not require additional hypotheses or new experimental results. In the second edition the rational reconstructions are completed with respect to General Relativity and Cosmology. In addition, the statistics of quantum objects is elaborated in more detail with respect to the rational reconstruction of quantum mechanics. The new material completes the approach of t...

Does decision documentation help junior designers rationalize their decisions? A comparative multiple-case study

OpenAIRE

Heesch, U. van; Avgeriou, P.; Tang, A.

2013-01-01

Software architecture design is challenging, especially for junior software designers. Lacking practice and experience, junior designers need process support in order to make rational architecture decisions. In this paper, we present the results of a comparative multiple-case study conducted to find out if decision viewpoints from van Heesch et al. (2012, in press) can provide such a support. The case study was conducted with four teams of software engineering students working in industrial s...

Rational Choice and the Framing of Decisions.

Science.gov (United States)

1986-05-29

survival in a competitive environment , and a minority of rational individuals can sometimes impose rationality on the whole market. Third, the...intuitive appeal of the axioms of rational choice makes it plausible that the theory derived from these axioms should provide an acceptable account of choice...rn-use U? RATIONAL CHOICE AMD THE FINNING OF KCISIOUS(U mi/ STANFORD UNIV CR A TYERSEY ET AL. 29 NAYN4-S4-K-S61SWICLASS IF lED FO 5/10S IL EEEEEEEE

Intergroup conflict and rational decision making.

Science.gov (United States)

Martínez-Tur, Vicente; Peñarroja, Vicente; Serrano, Miguel A; Hidalgo, Vanesa; Moliner, Carolina; Salvador, Alicia; Alacreu-Crespo, Adrián; Gracia, Esther; Molina, Agustín

2014-01-01

The literature has been relatively silent about post-conflict processes. However, understanding the way humans deal with post-conflict situations is a challenge in our societies. With this in mind, we focus the present study on the rationality of cooperative decision making after an intergroup conflict, i.e., the extent to which groups take advantage of post-conflict situations to obtain benefits from collaborating with the other group involved in the conflict. Based on dual-process theories of thinking and affect heuristic, we propose that intergroup conflict hinders the rationality of cooperative decision making. We also hypothesize that this rationality improves when groups are involved in an in-group deliberative discussion. Results of a laboratory experiment support the idea that intergroup conflict -associated with indicators of the activation of negative feelings (negative affect state and heart rate)- has a negative effect on the aforementioned rationality over time and on both group and individual decision making. Although intergroup conflict leads to sub-optimal decision making, rationality improves when groups and individuals subjected to intergroup conflict make decisions after an in-group deliberative discussion. Additionally, the increased rationality of the group decision making after the deliberative discussion is transferred to subsequent individual decision making.

Intergroup Conflict and Rational Decision Making

Science.gov (United States)

Martínez-Tur, Vicente; Peñarroja, Vicente; Serrano, Miguel A.; Hidalgo, Vanesa; Moliner, Carolina; Salvador, Alicia; Alacreu-Crespo, Adrián; Gracia, Esther; Molina, Agustín

2014-01-01

The literature has been relatively silent about post-conflict processes. However, understanding the way humans deal with post-conflict situations is a challenge in our societies. With this in mind, we focus the present study on the rationality of cooperative decision making after an intergroup conflict, i.e., the extent to which groups take advantage of post-conflict situations to obtain benefits from collaborating with the other group involved in the conflict. Based on dual-process theories of thinking and affect heuristic, we propose that intergroup conflict hinders the rationality of cooperative decision making. We also hypothesize that this rationality improves when groups are involved in an in-group deliberative discussion. Results of a laboratory experiment support the idea that intergroup conflict –associated with indicators of the activation of negative feelings (negative affect state and heart rate)– has a negative effect on the aforementioned rationality over time and on both group and individual decision making. Although intergroup conflict leads to sub-optimal decision making, rationality improves when groups and individuals subjected to intergroup conflict make decisions after an in-group deliberative discussion. Additionally, the increased rationality of the group decision making after the deliberative discussion is transferred to subsequent individual decision making. PMID:25461384

Intergroup conflict and rational decision making.

Directory of Open Access Journals (Sweden)

Vicente Martínez-Tur

Full Text Available The literature has been relatively silent about post-conflict processes. However, understanding the way humans deal with post-conflict situations is a challenge in our societies. With this in mind, we focus the present study on the rationality of cooperative decision making after an intergroup conflict, i.e., the extent to which groups take advantage of post-conflict situations to obtain benefits from collaborating with the other group involved in the conflict. Based on dual-process theories of thinking and affect heuristic, we propose that intergroup conflict hinders the rationality of cooperative decision making. We also hypothesize that this rationality improves when groups are involved in an in-group deliberative discussion. Results of a laboratory experiment support the idea that intergroup conflict -associated with indicators of the activation of negative feelings (negative affect state and heart rate- has a negative effect on the aforementioned rationality over time and on both group and individual decision making. Although intergroup conflict leads to sub-optimal decision making, rationality improves when groups and individuals subjected to intergroup conflict make decisions after an in-group deliberative discussion. Additionally, the increased rationality of the group decision making after the deliberative discussion is transferred to subsequent individual decision making.

Rational customs clearance technology choice

OpenAIRE

Shramenko, N.; Andriets, V.

2008-01-01

Issues concerning cargo delivery efficiencyincrease by choice of rational customs clearance technology have been considered. Three possible variants of customs clearance andmethods which allow to define the most rational version of cargo delivery in international road communication based on main efficiency criteria for definite distance have been presented.

Extracellular matrix and growth factor engineering for controlled angiogenesis in regenerative medicine

Directory of Open Access Journals (Sweden)

Mikaël M Martino

2015-04-01

Full Text Available Blood vessel growth plays a key role in regenerative medicine, both to restore blood supply to ischemic tissues and to ensure rapid vascularization of clinical-size tissue-engineered grafts. For example, vascular endothelial growth factor (VEGF is the master regulator of physiological blood vessel growth and is one of the main molecular targets of therapeutic angiogenesis approaches. However, angiogenesis is a complex process and there is a need to develop rational therapeutic strategies based on a firm understanding of basic vascular biology principles, as evidenced by the disappointing results of initial clinical trials of angiogenic factor delivery. In particular, the spatial localization of angiogenic signals in the extracellular matrix is crucial to ensure the proper assembly and maturation of new vascular structures. Here we discuss the therapeutic implications of matrix interactions of angiogenic factors, with a special emphasis on VEGF, as well as provide an overview of current approaches, based on protein and biomaterial engineering that mimic the regulatory functions of extracellular matrix to optimize the signaling microenvironment of vascular growth factors.

The Effects of Different Energy and Protein Ratio to Sheep’s Nutrient Intake and Digestibility

Directory of Open Access Journals (Sweden)

Sri Mawati

2013-06-01

different (P>0.05 among crude protein and TDN treatments. Different energy and protein ration treatments caused different DM and OM intake but were not cause different in DM and OM digestibility. Based on the research results, a study on the effects of different ration’s energy and protein ratio towards N efficiency should be conducted in order to increase cattle productivity. Normal 0 false false false IN X-NONE X-NONE MicrosoftInternetExplorer4 Doi: http://dx.doi.org/10.12777/ijse.4.2.75-79 [How to cite this article: Mawati, S., Soedarsono, S., Sunarso, S. & Purnomoadi, A. (2013. The Effects of Different Energy and Ratio to Sheep’s Nutrient Intake and Digestibility. International Journal of Science and Engineering, 4(2,76-79. Doi: http://dx.doi.org/10.12777/ijse.4.2.75-79] Cognitive Rationality and Its Logic-Mathematical Language

Science.gov (United States)

Masalova, Svetlana

2012-01-01

The article deals with the cognitive (flexible) rationality, combining rational and irrational moments of the scientific search of the cognizing subject. Linguo-cognitive model of the concept as the flexible regulative rationality reveals the activity of the cognitive processes and the mentality of the epistemological-ontic subject, its leading…

Evaluation of Biological Toxicity of CdTe Quantum Dots with Different Coating Reagents according to Protein Expression of Engineering Escherichia coli

Directory of Open Access Journals (Sweden)

Wei Xu

2015-01-01

Full Text Available The results obtained from toxicity assessment of quantum dots (QDs can be used to establish guidelines for the application of QDs in bioimaging. This paper focused on the design of a novel method to evaluate the toxicity of CdTe QDs using engineering Escherichia coli as a model. The toxicity of mercaptoacetic acid (MPA, glutathione (GSH, and L-cysteine (Cys capped CdTe QDs was analyzed according to the heterologous protein expression in BL21/DE3, engineering Escherichia coli extensively used for protein expression. The results showed that the MPA-CdTe QDs had more serious toxicity than the other two kinds of CdTe QDs. The microscopic images and SEM micrographs further proved that both the proliferation and the protein expression of engineering Escherichia coli were inhibited after treatment with MPA-CdTe QDs. The proposed method is important to evaluate biological toxicity of both QDs and other nanoparticles.

Characteristics of dairy farms in the North-Eastern part of Italy: rations, milk yield and nutrients excretion

Directory of Open Access Journals (Sweden)

Stefano Schiavon

2010-01-01

Full Text Available This survey was aimed to evaluate the characteristics of dairy farms in the North- Eastern part of Po valley in terms of ration composition, milk yield and N and P excretions. Eightynine farms, with Italian Holstein Friesian cows, were selected in order to cover different situations in term of farm size and milk yield (MY. MY and quality were obtained from the national database of functional controls. Each farm was visited in order to collect information about ingredients and chemical composition of rations used. Farms were classified in four groups differing for dietary crude protein density (LCP15.3% DM and for MY (LMY30 kg/d. N and P excretions were quantified by following a mass balance approach. Dietary crude protein content (CP was not correlated to milk yield (MY and quality. The estimated amounts of N excreted, discounted for 28% of N losses in atmosphere, were 78.5, 78.2, 87.2 and 89.1 kg/cow/year, and P excreted were 20.2, 18.6, 18.7 and 19.8 kg/cow/year for the LCPLMY, LCPHMY, HCPLMY, HCPHMY groups, respectively. On corn silage and cereals based rations, a dietary CP of 14.3% DM can support 31 kg MY/cow/day.

Technique on rationalization of using electricity and cases

International Nuclear Information System (INIS)

1988-04-01

This book deals with rationalization of using electric and cases. It is divided into four parts. The first part introduces necessity and of progression rationalization of using electric. The second part describes the technique on rationalization of using electric with management of electric energy. The third part depicts domestic cases of rationalization on using of electric such as substation and motor. The last part also introduces foreign cases of rationalization on using of electric with measure of generator circuit, design of motor, design of lighting and design of other equipment.

Book Selection, Collection Development, and Bounded Rationality.

Science.gov (United States)

Schwartz, Charles A.

1989-01-01

Reviews previously proposed schemes of classical rationality in book selection, describes new approaches to rational choice behavior, and presents a model of book selection based on bounded rationality in a garbage can decision process. The role of tacit knowledge and symbolic content in the selection process are also discussed. (102 references)…

Love and rationality: on some possible rational effects of love

Directory of Open Access Journals (Sweden)

Gustavo Ortiz-Millán

2007-01-01

Full Text Available In this paper I defend the idea that rather than disrupting rationality, as the common-sense conception has done it, love may actually help us to develop rational ways of thinking and acting. I make the case for romantic or erotic love, since this is the kind of love that is more frequently associated with irrationality in acting and thinking. I argue that this kind of love may make us develop epistemic and practical forms of rationality. Based on an analysis of its characteristic action tendencies, I argue that love may help us to develop an instrumental form of rationality in determining the best means to achieve the object of love. It may also narrow down the number of practical considerations that may help us to achieve our goals. Finally, love may generate rational ways of belief-formation by framing the parameters taken into account in perception and attention, and by bringing into light only a small portion of the epistemic information available. Love may make us perceive reality more acutely.Neste artigo defendo a idéia de que, em vez de perturbar a racionalidade, como a concepção do senso comum o faz, o amor pode, na verdade, ajudar-nos a desenvolver modos racionais de pensar e agir. Dou bons argumentos para o amor romântico ou erótico, uma vez que esse é o tipo de amor que é mais freqüentemente associado à irracionalidade no agir e no pensar. Argumento que esse tipo de amor pode fazer-nos desenvolver formas epistêmicas e práticas de racionalidade. Com base em uma análise de suas tendências características para a ação, argumento que o amor pode ajudar-nos a desenvolver uma forma instrumental de racionalidade para se determinar o melhor meio de atingir o objeto de amor. Ele também pode limitar o número de considerações práticas que podem ajudar-nos a atingir os nossos objetivos. Finalmente, o amor pode gerar modos racionais de formação de crenças ao estruturar os parâmetros considerados na percepção e na aten

A discussion of theoretical and practical rationality

Energy Technology Data Exchange (ETDEWEB)

Wahlstroem, B. [Technical Research Centre of Finland, Espoo (Finland). VTT Automation

1999-12-01

Theoretical rationality as defined in Expected Utility Theory and amended with other considerations gives a good basis for decision making. One should however always keep in mind that practical rationality often is far more complicated. People use their everyday experience when placed before new problems and this may lead to apparently irrational choices which on a closer scrutiny may be completely rational. Theories in human decision making unfortunately becomes untestable, firstly because a theory taking all considerations into account would be to complex to be practical and secondly because the data needed to test the theory cannot be collected. The benefit of EUT is that it is simple and straightforward as compared with competing theories. In the natural sciences rationality is often seen simply as a problem of optimisation. This view is practical, but it has to include also psychological and sociological considerations. The apparent controversy between natural and behavioural sciences could at least in principle be resolved by a better understanding of the complexity of human rationality. The human mind does not work in isolation, but it is adapted to a social community and a continuously changing environment. Understanding all components of human rationality is a challenge which cannot be solved on a short term basis. An important part of human rationality is connected to the intricate balance between individual and societal utility. The human mind has over thousands of years learnt to resolve that balance, but in the modern society there are decisions which may not be solvable with an intuitive approach and a strategy of trial and error. For these decisions more solid theories of rationality will be needed. EUT can in spite of its dismerits be used as the backbone for such a theory, but it has to be extended with better explanations of both individual and social rationality. If this understanding of the practical aspects of human rationality can be reached

A discussion of theoretical and practical rationality

International Nuclear Information System (INIS)

Wahlstroem, B.

1999-01-01

Theoretical rationality as defined in Expected Utility Theory and amended with other considerations gives a good basis for decision making. One should however always keep in mind that practical rationality often is far more complicated. People use their everyday experience when placed before new problems and this may lead to apparently irrational choices which on a closer scrutiny may be completely rational. Theories in human decision making unfortunately becomes untestable, firstly because a theory taking all considerations into account would be to complex to be practical and secondly because the data needed to test the theory cannot be collected. The benefit of EUT is that it is simple and straightforward as compared with competing theories. In the natural sciences rationality is often seen simply as a problem of optimisation. This view is practical, but it has to include also psychological and sociological considerations. The apparent controversy between natural and behavioural sciences could at least in principle be resolved by a better understanding of the complexity of human rationality. The human mind does not work in isolation, but it is adapted to a social community and a continuously changing environment. Understanding all components of human rationality is a challenge which cannot be solved on a short term basis. An important part of human rationality is connected to the intricate balance between individual and societal utility. The human mind has over thousands of years learnt to resolve that balance, but in the modern society there are decisions which may not be solvable with an intuitive approach and a strategy of trial and error. For these decisions more solid theories of rationality will be needed. EUT can in spite of its dismerits be used as the backbone for such a theory, but it has to be extended with better explanations of both individual and social rationality. If this understanding of the practical aspects of human rationality can be reached

A review of metabolic and enzymatic engineering strategies for designing and optimizing performance of microbial cell factories

Directory of Open Access Journals (Sweden)

Amanda K. Fisher

2014-08-01

Full Text Available Microbial cell factories (MCFs are of considerable interest to convert low value renewable substrates to biofuels and high value chemicals. This review highlights the progress of computational models for the rational design of an MCF to produce a target bio-commodity. In particular, the rational design of an MCF involves: (i product selection, (ii de novo biosynthetic pathway identification (i.e., rational, heterologous, or artificial, (iii MCF chassis selection, (iv enzyme engineering of promiscuity to enable the formation of new products, and (v metabolic engineering to ensure optimal use of the pathway by the MCF host. Computational tools such as (i de novo biosynthetic pathway builders, (ii docking, (iii molecular dynamics (MD and steered MD (SMD, and (iv genome-scale metabolic flux modeling all play critical roles in the rational design of an MCF. Genome-scale metabolic flux models are of considerable use to the design process since they can reveal metabolic capabilities of MCF hosts. These can be used for host selection as well as optimizing precursors and cofactors of artificial de novo biosynthetic pathways. In addition, recent advances in genome-scale modeling have enabled the derivation of metabolic engineering strategies, which can be implemented using the genomic tools reviewed here as well.

THE HICKSIAN RATIONAL CONSUMER

OpenAIRE

Manuel FERNÁNDEZ-GRELA

2005-01-01

The aim of this paper is to trace the evolution of the concept of ''rational consumer'' in Hicks's writings. After being one of the pioneers in the introduction of rationality assumptions about consumer behaviour in economic models, Hicks gradually developed a sceptical view about some of the uses to which those assumptions were put into. The focus of the paper is on continuity in Hicksian views, providing a picture of gradual changes in the long series of Hicks's works

Protein folding and the organization of the protein topology universe

DEFF Research Database (Denmark)

Lindorff-Larsen,, Kresten; Røgen, Peter; Paci, Emanuele

2005-01-01

residues and, in addition, that the topology of the transition state is closer to that of the native state than to that of any other fold in the protein universe. Here, we review the evidence for these conclusions and suggest a molecular mechanism that rationalizes these findings by presenting a view...... of protein folds that is based on the topological features of the polypeptide backbone, rather than the conventional view that depends on the arrangement of different types of secondary-structure elements. By linking the folding process to the organization of the protein structure universe, we propose...

Coping More Effectively Through Rational Self-Counseling.

Science.gov (United States)

Rogers, George W., Jr.

1981-01-01

Rational Self-Counseling, a variation of rational-emotive therapy, is a self-help therapeutic technique in which students are encouraged to be responsible for their own behavior and emotions. The primary function of self-counseling is to evaluate whether thoughts are rational. A list of questions which students might ask themselves is presented.…

Improved feed protein fractionation schemes for formulating rations with the cornell net carbohydrate and protein system.

Science.gov (United States)

Lanzas, C; Broderick, G A; Fox, D G

2008-12-01

Adequate predictions of rumen-degradable protein (RDP) and rumen-undegradable protein (RUP) supplies are necessary to optimize performance while minimizing losses of excess nitrogen (N). The objectives of this study were to evaluate the original Cornell Net Carbohydrate Protein System (CNCPS) protein fractionation scheme and to develop and evaluate alternatives designed to improve its adequacy in predicting RDP and RUP. The CNCPS version 5 fractionates CP into 5 fractions based on solubility in protein precipitant agents, buffers, and detergent solutions: A represents the soluble nonprotein N, B1 is the soluble true protein, B2 represents protein with intermediate rates of degradation, B3 is the CP insoluble in neutral detergent solution but soluble in acid detergent solution, and C is the unavailable N. Model predictions were evaluated with studies that measured N flow data at the omasum. The N fractionation scheme in version 5 of the CNCPS explained 78% of the variation in RDP with a root mean square prediction error (RMSPE) of 275 g/d, and 51% of the RUP variation with RMSPE of 248 g/d. Neutral detergent insoluble CP flows were overpredicted with a mean bias of 128 g/d (40% of the observed mean). The greatest improvements in the accuracy of RDP and RUP predictions were obtained with the following 2 alternative schemes. Alternative 1 used the inhibitory in vitro system to measure the fractional rate of degradation for the insoluble protein fraction in which A = nonprotein N, B1 = true soluble protein, B2 = insoluble protein, C = unavailable protein (RDP: R(2) = 0.84 and RMSPE = 167 g/d; RUP: R(2) = 0.61 and RMSPE = 209 g/d), whereas alternative 2 redefined A and B1 fractions as the non-amino-N and amino-N in the soluble fraction respectively (RDP: R(2) = 0.79 with RMSPE = 195 g/d and RUP: R(2) = 0.54 with RMSPE = 225 g/d). We concluded that implementing alternative 1 or 2 will improve the accuracy of predicting RDP and RUP within the CNCPS framework.

Freedom and Rationality : Rousseau on Citizenship

OpenAIRE

Salvat, Christophe

2008-01-01

This paper deals with Rousseau's idea of freedom in terms of rationality and deliberation. It gives support to Berlin's interpretation of the general will as a rational and objective will but dismisses the idea that Rousseau's theory necessarily leads to authoritarianism. The general will, publicly expressed by the law, may be defined as the rational and self-regarding will agents would have if put in an independent and objective state, i.e. the state of nature. The general and the particular...

TAL effectors: highly adaptable phytobacterial virulence factors and readily engineered DNA targeting proteins

Science.gov (United States)

Doyle, Erin L.; Stoddard, Barry L.; Voytas, Daniel F.; Bogdanove, Adam J.

2013-01-01

Transcription activator-like (TAL) effectors are transcription factors injected into plant cells by pathogenic bacteria in the genus Xanthomonas. They function as virulence factors by activating host genes important for disease, or as avirulence factors by turning on genes that provide resistance. DNA binding specificity is encoded by polymorphic repeats in each protein that correspond one-to-one with different nucleotides. This code has facilitated target identification and opened new avenues for engineering disease resistance. It has also enabled TAL effector customization for targeted gene control, genome editing, and other applications. This article reviews the structural basis for TAL effector-DNA specificity, the impact of the TAL effector-DNA code on plant pathology and engineered resistance, and recent accomplishments and future challenges in TAL effector-based DNA targeting. PMID:23707478

Rational choice in field archaelology

Directory of Open Access Journals (Sweden)

Cătălin Pavel

2011-11-01

Full Text Available In the present article I attempt to apply advances in the study of instrumental and epistemic rationality to field archaeology in order to gain insights into the ways archaeologists reason. The cognitive processes, particularly processes of decision making, that enable archaeologists to conduct the excavation in the trench have not been adequately studied so far. I take my cues from two different bodies of theory. I first inquire into the potential that rational choice theory (RCT may have in modeling archaeological behaviour, and I define subjective expected utility, which archaeologists attempt to maximize, in terms of knowledge acquisition and social gain. Following Elster’s criticism of RCT, I conclude that RCT’s standards for rational action do not correspond with those ostensibly used in field archaeology, but that instrumental rationality has a prominent role in the “archaeological experiment”. I further explore if models proposed as reaction to RCT may account for archaeological decision making. I focus on fast and frugal heuristics, and search for archaeological illustrations for some of the cognitive biases that are better documented in psychological literature. I document confirmation and congruence biases, the endowment effect, observer-expectancy bias, illusory correlation, clustering illusion, sunk cost bias, and anchoring, among others and I propose that some of these biases are used as cognitive tools by archaeologists at work and retain epistemic value. However, I find formal logic to be secondary in the development of archaeological reasoning, with default logic and defeasible logic being used instead. I emphasize scientific knowledge as an actively negotiated social product of human inquiry, and conclude that to describe rationality in field archaeology a bounded rationality model is the most promising avenue of investigation.

The rational maps Fλ(z)

Indian Academy of Sciences (India)

It is proved that the rational maps in the family {z → zm +λ/zd : λ ∈ C\\{0}} for integers m, d ≥ 2 ... The problem of the existence of Herman rings of a rational map has been studied by. Lyubich in [9] ..... Surveys 41(4) (1986) 35–95. [10] Milnor J ...

Heterogeneity and the (de)stabilizing role of rationality

International Nuclear Information System (INIS)

Cavalli, Fausto; Naimzada, Ahmad; Pireddu, Marina

2015-01-01

Highlights: • We analyze Cournot oligopolies with heterogeneous firms of generic size. • Rational and naive players are considered. • Stability with respect to oligopoly composition is studied. • In some settings, increasing the rational firms fraction introduces instability. - Abstract: In this paper we study oligopolies of generic size consisting of heterogeneous firms, which adopt best response adjustment mechanisms with either perfect foresight (rational firms) or static expectations (naive firms). Assuming an isoelastic demand function and possibly different marginal costs for the two groups of firms, we focus on the local stability of the Nash equilibrium. We show that, with respect to the oligopoly composition, described in terms of the fraction of rational firms, different scenarios are possible. We find that a high rationality degree may not always guarantee stability, in particular when rational firms have sufficiently larger marginal costs. In fact, in this situation, increasing the fraction of rational firms can even introduce instability. Besides the usual scenarios in which replacing some naive firms with rational ones leads to a stabilization of (or at least keeps unchanged) the dynamics, we provide a family of situations, characterized by costs ratio favorable to naive firms, in which equilibrium loses its stability when naive firms are replaced by rational ones. The results we present are both analytical and simulative.

Rational assembly of nanoparticle superlattices with designed lattice symmetries

Science.gov (United States)

Gang, Oleg; Lu, Fang; Tagawa, Miho

2017-09-05

A method for lattice design via multivalent linkers (LDML) is disclosed that introduces a rationally designed symmetry of connections between particles in order to achieve control over the morphology of their assembly. The method affords the inclusion of different programmable interactions within one linker that allow an assembly of different types of particles. The designed symmetry of connections is preferably provided utilizing DNA encoding. The linkers may include fabricated "patchy" particles, DNA scaffold constructs and Y-shaped DNA linkers, anisotropic particles, which are preferably functionalized with DNA, multimeric protein-DNA complexes, and particles with finite numbers of DNA linkers.

Rationality and the Logic of Good Reasons.

Science.gov (United States)

Fisher, Walter R.

This paper contends that the rationality of the logic of good reasons is constituted in its use. To support this claim, the paper presents an analysis of the relationship between being reasonable and being rational. It then considers how following the logic of good reasons leads to rationality in the behavior of individuals and groups; the latter…

Rationality, mental causation and social sciences

OpenAIRE

Mladenović Ivan

2009-01-01

The aim of this paper is to investigate the role of mental causation in the context of rational choice theory. The author defends psychological aspect of rational explanation against the challenge of contemporary reductive materialism.

Synthetic biology: an emerging engineering discipline.

Science.gov (United States)

Cheng, Allen A; Lu, Timothy K

2012-01-01

Over the past decade, synthetic biology has emerged as an engineering discipline for biological systems. Compared with other substrates, biology poses a unique set of engineering challenges resulting from an incomplete understanding of natural biological systems and tools for manipulating them. To address these challenges, synthetic biology is advancing from developing proof-of-concept designs to focusing on core platforms for rational and high-throughput biological engineering. These platforms span the entire biological design cycle, including DNA construction, parts libraries, computational design tools, and interfaces for manipulating and probing synthetic circuits. The development of these enabling technologies requires an engineering mindset to be applied to biology, with an emphasis on generalizable techniques in addition to application-specific designs. This review aims to discuss the progress and challenges in synthetic biology and to illustrate areas where synthetic biology may impact biomedical engineering and human health.

Physiological responses to rational-emotive self-verbalizations.

Science.gov (United States)

Master, S; Gershman, L

1983-12-01

This study tested Albert Ellis' Rational Emotive Therapy (RET) theory which predicts that cognitive beliefs, not the stimulus situation, generate human emotions. According to RET, emotions created by rational beliefs are adaptive, while irrational beliefs result in an unadaptive anxiety level. Results demonstrated that at high levels of problem relevance there was (1) a significantly greater GSR in direct response to the stimulus situation, and also to irrational statements, than to rational and control statements, and (2) no significant difference between rational and neutral control statements. The authors argue that these results are more parsimoniously explained by conditioning theory than by RET theory.

Towards a Characterization of Rational Expectations

OpenAIRE

Itai Arieli

2008-01-01

R. J. Aumann and J. H. Drèze (2008) define a rational expectation of a player i in a game G as the expected payo of some type of i in some belief system for G in which common knowledge of rationality and common priors obtain. Our goal is to characterize the set of rational expectations in terms of the game's payoff matrix. We provide such a characterization for a specific class of strategic games, called semi-elementary, which includes Myerson's "elementary" games.

Annual conference on engineering and the physical sciences in medicine

International Nuclear Information System (INIS)

Le Heron, J.

1999-01-01

The venue for the 1998 annual conference on Engineering and the Physical Sciences in Medicine was the Wrest Point Casino Convention Centre, Hobart, from 15 to 19 November. Jointly sponsored by the Australasian College of Physical Scientists and Engineers in Medicine, the College of Biomedical Engineers and the Society of Medical and Biomedical Engineering, this meeting is a major forum for professionals working in these areas in Australasia. The theme for the conference was Relevance beyond rationalism - charting a course for the future. This reviewer will consider only those presentations concerned with the use of radiation in medicine. (author)

Rational inattention or rational overreaction?

DEFF Research Database (Denmark)

Browning, Martin; Hansen, Lars Gårn; Smed, Sinne

We investigate differences in how consumers of fish react to health information in the mass media. We specify a dynamic empirical model that allows for heterogeneity in all basic parameters of consumer behavior as well as in how consumers react to information. We estimate the model using a unique...... houshold panel tracking consumption, prices, news stories and media habits over 24 quarters. We fi nd that the consumers most likely to be ’rationally ignorant’ of health effects react more dramatically to health news than the consumers who most likely are well informed....

Site-specific antibody-drug conjugates: the nexus of bioorthogonal chemistry, protein engineering, and drug development.

Science.gov (United States)

Agarwal, Paresh; Bertozzi, Carolyn R

2015-02-18

Antibody-drug conjugates (ADCs) combine the specificity of antibodies with the potency of small molecules to create targeted drugs. Despite the simplicity of this concept, generation of clinically successful ADCs has been very difficult. Over the past several decades, scientists have learned a great deal about the constraints on antibodies, linkers, and drugs as they relate to successful construction of ADCs. Once these components are in hand, most ADCs are prepared by nonspecific modification of antibody lysine or cysteine residues with drug-linker reagents, which results in heterogeneous product mixtures that cannot be further purified. With advances in the fields of bioorthogonal chemistry and protein engineering, there is growing interest in producing ADCs by site-specific conjugation to the antibody, yielding more homogeneous products that have demonstrated benefits over their heterogeneous counterparts in vivo. Here, we chronicle the development of a multitude of site-specific conjugation strategies for assembly of ADCs and provide a comprehensive account of key advances and their roots in the fields of bioorthogonal chemistry and protein engineering.

ENERGETIC VALUE OF FORAGES FROM SEMI-ARID REGION AND DIGESTIBILITY OF RATIONS FOR NAKED NECK PULLETS

Directory of Open Access Journals (Sweden)

ALEX MARTINS VARELA DE ARRUDA

2014-01-01

Full Text Available The feeding programs for naked neck chickens in semi-intensive production system from brazilian equatorial semi-arid environment, must consider regional food availability and respective nutritional values. Thus, to evaluate the digestibility and metabolizable energy of alternative forages, it was used 240 naked neck pullets (Isa Label lineage receiving water and ration ad libitum, pair-housed in cages for total collection of excreta on conventional warehouse. It was used a completely randomized design with factorial arrangement (5x2: one control ration (corn and soy meal and other four experimental rations with silk flower hay (Calotropis procera, cassava leafs hay (Manihot esculenta, kills pasture hay (Senna obtusifolia or leucaena leafs hay (Leucaena leucocephala, and all rations were balanced for two growing phases, between 8 and 10 weeks (young pullets and between 14 and 16 weeks of age (old pullets. The values of apparent digestibility of nutrients for all experimental rations were lower than control ration (P <0.05 and it was observed general means of 72.18% for dry matter, 78.12% for crude protein, 66.90% for ether extract, 28.08% for neutral detergent fiber, 18.51% for the acid detergent fiber, 71.64% for gross energy and availability of 15.61% for mineral matter. The general mean of apparent and corrected metabolizable energy of alternative forages was 1217 kcal/ kg and 1108 kcal/kg, respectively, and the higher value was determined for leucaena hay and the lower value for silk flower hay (P <0.05.

Rationality, mental causation and social sciences

Directory of Open Access Journals (Sweden)

Mladenović Ivan

2009-01-01

Full Text Available The aim of this paper is to investigate the role of mental causation in the context of rational choice theory. The author defends psychological aspect of rational explanation against the challenge of contemporary reductive materialism.

Pandemic ventilator rationing and appeals processes.

Science.gov (United States)

Patrone, Daniel; Resnik, David

2011-06-01

In a severe influenza pandemic, hospitals will likely experience serious and widespread shortages of patient pulmonary ventilators and of staff qualified to operate them. Deciding who will receive access to mechanical ventilation will often determine who lives and who dies. This prospect raises an important question whether pandemic preparedness plans should include some process by which individuals affected by ventilator rationing would have the opportunity to appeal adverse decisions. However, the issue of appeals processes to ventilator rationing decisions has been largely neglected in state pandemic planning efforts. If we are to devise just and effective plans for coping with a severe influenza pandemic, more attention to the issue of appeals processes for pandemic ventilator rationing decisions is needed. Arguments for and against appeals processes are considered, and some suggestions are offered to help efforts at devising more rational pandemic preparedness plans.

Changes in protein composition and protein phosphorylation during ...

African Journals Online (AJOL)

STORAGESEVER

2009-08-04

Aug 4, 2009 ... liquid detergent (teepol) solution (5%, v/v) for 15 min and rinsed ... stacking gel contained 2.4% bisacrylamide as a cross linker and. 0.1% SDS. The final buffer concentrations were 0.45 M Tris Hcl pH. (8.9) in resolving gel and 0.2 M Tris HCl ... rations of the protein samples by SDS-PAGE as described pre-.

RATIONAL APPROXIMATIONS TO GENERALIZED HYPERGEOMETRIC FUNCTIONS.

Science.gov (United States)

Under weak restrictions on the various free parameters, general theorems for rational representations of the generalized hypergeometric functions...and certain Meijer G-functions are developed. Upon specialization, these theorems yield a sequency of rational approximations which converge to the

Many faces of rationality: Implications of the great rationality debate for clinical decision-making

OpenAIRE

Djulbegovic, B.; Elqayam, Shira

2017-01-01

open access article Given that more than 30% of healthcare costs are wasted on inappropriate care, suboptimal care is increasingly connected to the quality of medical decisions. It has been argued that personal decisions are the leading cause of death, and 80% of healthcare expenditures result from physicians' decisions. Therefore, improving healthcare necessitates improving medical decisions, ie, making decisions (more) rational. Drawing on writings fromThe Great Rationality Debate from t...

Visualized and precise design of artificial small RNAs for regulating T7 RNA polymerase and enhancing recombinant protein folding in Escherichia coli

Directory of Open Access Journals (Sweden)

Yujia Zhao

2016-12-01

Full Text Available Small non-coding RNAs (sRNAs have received much attention in recent years due to their unique biological properties, which can efficiently and specifically tune target gene expressions in bacteria. Inspired by natural sRNAs, recent works have proposed the use of artificial sRNAs (asRNAs as genetic tools to regulate desired gene that has been applied in several fields, such as metabolic engineering and bacterial physiology studies. However, the rational design of asRNAs is still a challenge. In this study, we proposed structure and length as two criteria to implement rational visualized and precise design of asRNAs. T7 expression system was one of the most useful recombinant protein expression systems. However, it was deeply limited by the formation of inclusion body. To settle this problem, we designed a series of asRNAs to inhibit the T7 RNA polymerase (Gene1 expression to balance the rate between transcription and folding of recombinant protein. Based on the heterologous expression of Aspergillus oryzae Li-3 glucuronidase in E. coli, the asRNA-antigene1-17bp can effectively decrease the inclusion body and increase the enzyme activity by 169.9%.

Rationalizing the Promotion of Non-Rational Behaviors in Organizations.

Science.gov (United States)

Smith, Peter A. C.; Sharma, Meenakshi

2002-01-01

Organizations must balance rational/technical efficiency and emotions. Action learning has been proven to be effective for developing emotional openness in the workplace. Facilitators of action learning should draw upon the disciplines of counseling, Gestalt, psychodynamics, and Eastern philosophies. (Contains 23 references.) (SK)

Generalized NLS hierarchies from rational W algebras

International Nuclear Information System (INIS)

Toppan, F.

1993-11-01

Finite rational W algebras are very natural structures appearing in coset constructions when a Kac-Moody subalgebra is factored out. The problem of relating these algebras to integrable hierarchies of equations is studied by showing how to associate to a rational W algebra its corresponding hierarchy. Two examples are worked out, the sl(2)/U(1) coset, leading to the Non-Linear Schroedinger hierarchy, and the U(1) coset of the Polyakov-Bershadsky W algebra, leading to a 3-field representation of the KP hierarchy already encountered in the literature. In such examples a rational algebra appears as algebra of constraints when reducing a KP hierarchy to a finite field representation. This fact arises the natural question whether rational algebras are always associated to such reductions and whether a classification of rational algebras can lead to a classification of the integrable hierarchies. (author). 19 refs

Engineering Ni-Mo-S Nanoparticles for Hydrodesulfurization

DEFF Research Database (Denmark)

Bodin, Anders; Christoffersen, Ann-Louise N.; Elkjær, Christian F.

2018-01-01

Nanoparticle engineering for catalytic applications requires both a synthesis technique for the production of well-defined nanoparticles and measurements of their catalytic performance. In this paper, we present a new approach to rationally engineering highly active Ni-Mo-S nanoparticle catalysts...... for hydrodesulfurization (HDS), i.e., the removal of sulfur from fossil fuels. Nanoparticle catalysts are synthesized by the sputtering of a Mo75Ni25 metal target in a reactive atmosphere of Ar and H2S followed by the gas aggregation of the sputtered material into nanoparticles. The nanoparticles are filtered...

Lying for the Greater Good: Bounded Rationality in a Team

Directory of Open Access Journals (Sweden)

Oktay Sürücü

2014-10-01

Full Text Available This paper is concerned with the interaction between fully and boundedly rational agents in situations where their interests are perfectly aligned. The cognitive limitations of the boundedly rational agent do not allow him to fully understand the market conditions and lead him to take non-optimal decisions in some situations. Using categorization to model bounded rationality, we show that the fully rational agent can nudge, i.e., he can manipulate the information he sends and decrease the expected loss caused by the boundedly rational agent. Assuming different types for the boundedly rational agent, who differ only in the categories used, we show that the fully rational agent may learn the type of the boundedly rational agent along their interaction. Using this additional information, the outcome can be improved and the amount of manipulated information can be decreased. Furthermore, as the length of the interaction increases the probability that the fully rational agent learns the type of the boundedly rational agent grows

Normative and descriptive rationality: from nature to artifice and back

Science.gov (United States)

Besold, T. R.; Uckelman, S. L.

2018-03-01

Rationality plays a key role in both the study of human reasoning and Artificial Intelligence (AI). Certain notions of rationality have been adopted in AI as guides for the development of intelligent machines and these notions have been given a normative function. The notions of rationality in AI are often taken to be closely related to conceptions of rationality in human contexts. In this paper, we argue that the normative role of rationality differs in the human and artificial contexts. While rationality in human-focused fields of study is normative, prescribing how humans ought to reason, the normative conception in AI is built on a notion of human rationality which is descriptive, not normative, in the human context, as AI aims at building agents which reason as humans do. In order to make this point, we review prominent notions of rationality used in psychology, cognitive science, and (the history of) philosophy, as well as in AI, and discuss some factors that contributed to rationality being assigned the differing normative statuses in the differing fields of study. We argue that while 'rationality' is a normative notion in both AI and in human reasoning, the normativity of the AI conception of 'rationality' is grounded in a descriptive account of human rationality.

Engineering Aromatic-Aromatic Interactions To Nucleate Folding in Intrinsically Disordered Regions of Proteins.

Science.gov (United States)

Balakrishnan, Swati; Sarma, Siddhartha P

2017-08-22

Aromatic interactions are an important force in protein folding as they combine the stability of a hydrophobic interaction with the selectivity of a hydrogen bond. Much of our understanding of aromatic interactions comes from "bioinformatics" based analyses of protein structures and from the contribution of these interactions to stabilizing secondary structure motifs in model peptides. In this study, the structural consequences of aromatic interactions on protein folding have been explored in engineered mutants of the molten globule protein apo-cytochrome b 5 . Structural changes from disorder to order due to aromatic interactions in two variants of the protein, viz., WF-cytb5 and FF-cytb5, result in significant long-range secondary and tertiary structure. The results show that 54 and 52% of the residues in WF-cytb5 and FF-cytb5, respectively, occupy ordered regions versus 26% in apo-cytochrome b 5 . The interactions between the aromatic groups are offset-stacked and edge-to-face for the Trp-Phe and Phe-Phe mutants, respectively. Urea denaturation studies indicate that both mutants have a C m higher than that of apo-cytochrome b 5 and are more stable to chaotropic agents than apo-cytochrome b 5 . The introduction of these aromatic residues also results in "trimer" interactions with existing aromatic groups, reaffirming the selectivity of the aromatic interactions. These studies provide insights into the aromatic interactions that drive disorder-to-order transitions in intrinsically disordered regions of proteins and will aid in de novo protein design beyond small peptide scaffolds.

Momentum effect in stocks’ returns between the rational and the behavioural financial theories: Proposition of the progressive rationality

Directory of Open Access Journals (Sweden)

Faten Zoghlami

2013-01-01

Full Text Available     The puzzling momentum strategies’ payoffs defied the rational financial theory asserting the stocks returns’ unpredictability. Moreover, the momentum effect persist the main stocks returns’ anomaly escaping any risk-based explanation. The resilience of this phenomenon had favoured the development of behavioural financial field, which breaks with the investor’ full rationality hypothesis. This paper attempts to reconcile between the rational and behavioural financial theories, through the introduction of the progressive rationality concept. Especially, we argue that recognizing the temporary inappropriate investors’ reactions; can resolve the puzzling momentum anomaly. To fulfil our objective, we identify the appropriate autoregressive level that captures the significant autocorrelations involved by the investors’ over and under reactions. Then, we explore the profitability of the 6/6 momentum strategy implemented on the adjusted stocks’ returns. The adjusted momentum strategy is still profitable but no longer puzzling, since the related excess return is henceforth fully captured by a β and a size effect.Key words: Tunisian momentum effect, the rational finance theory, the behavioural finance theory, the three-factorial model and the autoregressive process.

A divide and conquer approach to determine the Pareto frontier for optimization of protein engineering experiments

Science.gov (United States)

He, Lu; Friedman, Alan M.; Bailey-Kellogg, Chris

2016-01-01

In developing improved protein variants by site-directed mutagenesis or recombination, there are often competing objectives that must be considered in designing an experiment (selecting mutations or breakpoints): stability vs. novelty, affinity vs. specificity, activity vs. immunogenicity, and so forth. Pareto optimal experimental designs make the best trade-offs between competing objectives. Such designs are not “dominated”; i.e., no other design is better than a Pareto optimal design for one objective without being worse for another objective. Our goal is to produce all the Pareto optimal designs (the Pareto frontier), in order to characterize the trade-offs and suggest designs most worth considering, but to avoid explicitly considering the large number of dominated designs. To do so, we develop a divide-and-conquer algorithm, PEPFR (Protein Engineering Pareto FRontier), that hierarchically subdivides the objective space, employing appropriate dynamic programming or integer programming methods to optimize designs in different regions. This divide-and-conquer approach is efficient in that the number of divisions (and thus calls to the optimizer) is directly proportional to the number of Pareto optimal designs. We demonstrate PEPFR with three protein engineering case studies: site-directed recombination for stability and diversity via dynamic programming, site-directed mutagenesis of interacting proteins for affinity and specificity via integer programming, and site-directed mutagenesis of a therapeutic protein for activity and immunogenicity via integer programming. We show that PEPFR is able to effectively produce all the Pareto optimal designs, discovering many more designs than previous methods. The characterization of the Pareto frontier provides additional insights into the local stability of design choices as well as global trends leading to trade-offs between competing criteria. PMID:22180081

A divide-and-conquer approach to determine the Pareto frontier for optimization of protein engineering experiments.

Science.gov (United States)

He, Lu; Friedman, Alan M; Bailey-Kellogg, Chris

2012-03-01

In developing improved protein variants by site-directed mutagenesis or recombination, there are often competing objectives that must be considered in designing an experiment (selecting mutations or breakpoints): stability versus novelty, affinity versus specificity, activity versus immunogenicity, and so forth. Pareto optimal experimental designs make the best trade-offs between competing objectives. Such designs are not "dominated"; that is, no other design is better than a Pareto optimal design for one objective without being worse for another objective. Our goal is to produce all the Pareto optimal designs (the Pareto frontier), to characterize the trade-offs and suggest designs most worth considering, but to avoid explicitly considering the large number of dominated designs. To do so, we develop a divide-and-conquer algorithm, Protein Engineering Pareto FRontier (PEPFR), that hierarchically subdivides the objective space, using appropriate dynamic programming or integer programming methods to optimize designs in different regions. This divide-and-conquer approach is efficient in that the number of divisions (and thus calls to the optimizer) is directly proportional to the number of Pareto optimal designs. We demonstrate PEPFR with three protein engineering case studies: site-directed recombination for stability and diversity via dynamic programming, site-directed mutagenesis of interacting proteins for affinity and specificity via integer programming, and site-directed mutagenesis of a therapeutic protein for activity and immunogenicity via integer programming. We show that PEPFR is able to effectively produce all the Pareto optimal designs, discovering many more designs than previous methods. The characterization of the Pareto frontier provides additional insights into the local stability of design choices as well as global trends leading to trade-offs between competing criteria. Copyright © 2011 Wiley Periodicals, Inc.

Emerging Computational Methods for the Rational Discovery of Allosteric Drugs.

Science.gov (United States)

Wagner, Jeffrey R; Lee, Christopher T; Durrant, Jacob D; Malmstrom, Robert D; Feher, Victoria A; Amaro, Rommie E

2016-06-08

Allosteric drug development holds promise for delivering medicines that are more selective and less toxic than those that target orthosteric sites. To date, the discovery of allosteric binding sites and lead compounds has been mostly serendipitous, achieved through high-throughput screening. Over the past decade, structural data has become more readily available for larger protein systems and more membrane protein classes (e.g., GPCRs and ion channels), which are common allosteric drug targets. In parallel, improved simulation methods now provide better atomistic understanding of the protein dynamics and cooperative motions that are critical to allosteric mechanisms. As a result of these advances, the field of predictive allosteric drug development is now on the cusp of a new era of rational structure-based computational methods. Here, we review algorithms that predict allosteric sites based on sequence data and molecular dynamics simulations, describe tools that assess the druggability of these pockets, and discuss how Markov state models and topology analyses provide insight into the relationship between protein dynamics and allosteric drug binding. In each section, we first provide an overview of the various method classes before describing relevant algorithms and software packages.

Synthetic biology approaches to engineer T cells.

Science.gov (United States)

Wu, Chia-Yung; Rupp, Levi J; Roybal, Kole T; Lim, Wendell A

2015-08-01

There is rapidly growing interest in learning how to engineer immune cells, such as T lymphocytes, because of the potential of these engineered cells to be used for therapeutic applications such as the recognition and killing of cancer cells. At the same time, our knowhow and capability to logically engineer cellular behavior is growing rapidly with the development of synthetic biology. Here we describe how synthetic biology approaches are being used to rationally alter the behavior of T cells to optimize them for therapeutic functions. We also describe future developments that will be important in order to construct safe and precise T cell therapeutics. Copyright © 2015 Elsevier Ltd. All rights reserved.

Neurophysiology and Rationality in Political Thinking.

Science.gov (United States)

Peterson, Steven A.

Research both in cognitive psychology and psychobiology suggests that political behavior is often less rational than individuals believe it to be. Information processing, memory, and decision making are interlinked processes. Studies in cognitive psychology reveal that even though decision making requires rationality, individuals often adopt…

Positroids Induced by Rational Dyck Paths

OpenAIRE

Gotti, Felix

2017-01-01

A rational Dyck path of type $(m,d)$ is an increasing unit-step lattice path from $(0,0)$ to $(m,d) \\in \\mathbb{Z}^2$ that never goes above the diagonal line $y = (d/m)x$. On the other hand, a positroid of rank $d$ on the ground set $[d+m]$ is a special type of matroid coming from the totally nonnegative Grassmannian. In this paper we describe how to naturally assign a rank $d$ positroid on the ground set $[d+m]$, which we name rational Dyck positroid, to each rational Dyck path of type $(m,d...

Rationalization and the Role of the School Counselor.

Science.gov (United States)

Clark, Arthur J.

1995-01-01

Examines rationalization in counselors' interactions with students, parents, and teachers--provides examples of each kind of interaction. Describes the dynamics of rationalization in the schools and outlines interventions that may be used with students, parents, and teachers. Also explores counselors' use of rationalization and gives examples of…

The effect of dietary protein on reproduction in the mare. I. The composition and evaluation of the digestibility of dietary protein from different sources

Directory of Open Access Journals (Sweden)

F.E. Van Niekerk

1997-07-01

Full Text Available Four rations that differed in their crude protein and essential amino-acid content were compiled. Digestibility of the crude protein and essential amino-acid contents were determined biologically in a feeding trial using 4 Anglo-Arab stallions. Their respective daily diets were: Diet 1: 2 kg cubes, 5 kg tef hay (Eragrostis tef; Diet 2: 2 kg cubes, 5 kg lucerne hay (Medicago sativa; Diet 3: 2 kg cubes, 5 kg tef hay, 200 g fishmeal; Diet 4: 2 kg cubes, 5 kg lucerne hay, 200 g fishmeal. The concentrations of the amino-acids threonine, iso-leucine, leucine and arginine were increased in the total ration when lucerne hay replaced the tef hay while fishmeal supplementation increased the methionine and lysine contents, which provided a wide range of concentrations of digestible amino-acids in each of the 4 rations.

An antibody based approach for multi-coloring osteogenic and chondrogenic proteins in tissue engineered constructs.

Science.gov (United States)

Leferink, Anne M; Reis, Diogo Santos; van Blitterswijk, Clemens A; Moroni, Lorenzo

2018-04-11

When tissue engineering strategies rely on the combination of three-dimensional (3D) polymeric or ceramic scaffolds with cells to culture implantable tissue constructs in vitro, it is desirable to monitor tissue growth and cell fate to be able to more rationally predict the quality and success of the construct upon implantation. Such a 3D construct is often referred to as a 'black-box' since the properties of the scaffolds material limit the applicability of most imaging modalities to assess important construct parameters. These parameters include the number of cells, the amount and type of tissue formed and the distribution of cells and tissue throughout the construct. Immunolabeling enables the spatial and temporal identification of multiple tissue types within one scaffold without the need to sacrifice the construct. In this report, we concisely review the applicability of antibodies (Abs) and their conjugation chemistries in tissue engineered constructs. With some preliminary experiments, we show an efficient conjugation strategy to couple extracellular matrix Abs to fluorophores. The conjugated probes proved to be effective in determining the presence of collagen type I and type II on electrospun and additive manufactured 3D scaffolds seeded with adult human bone marrow derived mesenchymal stromal cells. The conjugation chemistry applied in our proof of concept study is expected to be applicable in the coupling of any other fluorophore or particle to the Abs. This could ultimately lead to a library of probes to permit high-contrast imaging by several imaging modalities.

Rationality in the Cryptographic Model

DEFF Research Database (Denmark)

Hubacek, Pavel

This thesis presents results in the field of rational cryptography. In the first part we study the use of cryptographic protocols to avoid mediation and binding commitment when implementing game theoretic equilibrium concepts. First, we concentrate on the limits of cryptographic cheap talk...... to implement correlated equilibria of two-player strategic games in a sequentially rational way. We show that there exist two-player games for which no cryptographic protocol can implement the mediator in a sequentially rational way; that is, without introducing empty threats. In the context of computational...... with appealing economic applications. Our implementation puts forward a notion of cryptographically blinded games that exploits the power of encryption to selectively restrict the information available to players about sampled action profiles, such that these desirable equilibria can be stably achieved...

Heteronuclear 2D NMR studies on an engineered insulin monomer: Assignments and characterization of the receptor-binding surface by selective 2H and 13C labeling with application to protein design

International Nuclear Information System (INIS)

Weiss, M.A.; Hua, Qingxin; Lynch, C.S.; Shoelson, S.E.; Frank, B.H.

1991-01-01

Insulin provides an important model for the application of genetic engineering to rational protein design and has been well characterized in the crystal state. However, self-association of insulin in solution has precluded complementary 2D NMR study under physiological conditions. The authors demonstrate here that such limitations may be circumvented by the use of a monomeric analogue that contains three amino acid substitutions on the protein surface (HisB10 → Asp, ProB28 → Lys, and LysB29 → Pro); this analogue (designated DKP-insulin) retains native receptor-binding potency. Comparative 1 H NMR studies of native human insulin and a series of three related analogues-(i) the singly substituted analogue [HisB10→Asp], (ii) the doubly substituted analogue [ProB28→Lys; LysB29→Pro], and (iii) DKP-insulin-demonstrate progressive reduction in concentration-dependent line-broadening in accord with the results of analytical ultracentrifugation. Extensive nonlocal interactions are observed in the NOESY spectrum of DKP-insulin, indicating that this analogue adopts a compact and stably folded structure as a monomer in overall accord with crystal models. Site-specific 2 H and 13 C isotopic labels are introduced by semisynthesis as probes for the structure and dynamics of the receptor-binding surface. These studies confirm and extend under physiological conditions the results of a previous 2D NMR analysis of native insulin in 20% acetic acid. Implications for the role of protein flexibility in receptor recognition are discussed with application to the design of novel insulin analogues

Specific Internalisation of Gold Nanoparticles into Engineered Porous Protein Cages via Affinity Binding.

Science.gov (United States)

Paramelle, David; Peng, Tao; Free, Paul; Fernig, David G; Lim, Sierin; Tomczak, Nikodem

2016-01-01

Porous protein cages are supramolecular protein self-assemblies presenting pores that allow the access of surrounding molecules and ions into their core in order to store and transport them in biological environments. Protein cages' pores are attractive channels for the internalisation of inorganic nanoparticles and an alternative for the preparation of hybrid bioinspired nanoparticles. However, strategies based on nanoparticle transport through the pores are largely unexplored, due to the difficulty of tailoring nanoparticles that have diameters commensurate with the pores size and simultaneously displaying specific affinity to the cages' core and low non-specific binding to the cages' outer surface. We evaluated the specific internalisation of single small gold nanoparticles, 3.9 nm in diameter, into porous protein cages via affinity binding. The E2 protein cage derived from the Geobacillus stearothermophilus presents 12 pores, 6 nm in diameter, and an empty core of 13 nm in diameter. We engineered the E2 protein by site-directed mutagenesis with oligohistidine sequences exposing them into the cage's core. Dynamic light scattering and electron microscopy analysis show that the structures of E2 protein cages mutated with bis- or penta-histidine sequences are well conserved. The surface of the gold nanoparticles was passivated with a self-assembled monolayer made of a mixture of short peptidols and thiolated alkane ethylene glycol ligands. Such monolayers are found to provide thin coatings preventing non-specific binding to proteins. Further functionalisation of the peptide coated gold nanoparticles with Ni2+ nitrilotriacetic moieties enabled the specific binding to oligohistidine tagged cages. The internalisation via affinity binding was evaluated by electron microscopy analysis. From the various mutations tested, only the penta-histidine mutated E2 protein cage showed repeatable and stable internalisation. The present work overcomes the limitations of currently

The effect of selected cereals contained in feed ration on the amino acid composition of cows’ milk

Directory of Open Access Journals (Sweden)

Markéta Šípalová

2010-01-01

Full Text Available The aim of this study was to evaluate the possible effects of maize replacement in feeding rations on the amino acid content in cows´ milk. Cows were fed total mix ration based on the maize, clover silage and hay. There was a difference in the concentrate of the feeding ration. The first group (fed maize was the control group, another two groups were experimental, one fed wheat and second fed triticale. During six weeks, totally 26 milk samples were taken from dairy cows of Czech Pied breed. Feed groups were preferably balanced in terms of milk yield, stage and number of lactations. The samples of feedstuffs as well as milk were modified for the analysis using acidic and oxidative hydrolyses. The analysis of amino acids content and composition of the sample hydrolysates was performed chromatographically by an AAA 400 analyzer, using Na-citrate buffers and ninhydrin detection. Total nitrogen content was determined according to Kjehldahl and the crude protein of the samples was determined by conversion from the nitrogen content multiplied by appropriate factor. The high content of crude protein in wheat did not influenced composition of milk from dairy cows fed this type of feedstuff. With respect to resulting amino acid content and composition of milk samples, none of the tested grains can be re­com­men­ded as a full-value maize replacement. Each feedstuff is an abundant source of several and ty­pi­cal amino acids in milk. However, triticale (cultivar Kitaro seems to be acceptable replacement of maize owing to better crude protein efficiency, composition and health indicators of milk quality.

Proposed standby gasoline rationing plan: public comments

Energy Technology Data Exchange (ETDEWEB)

1978-12-01

Under the proposed plan, DOE would allocate ration rights (rights to purchase gasoline) to owners of registered vehicles. All vehicles in a given class would receive the same entitlement. Essential services would receive supplemental allotments of ration rights as pririty firms. Once every 3 months, ration checks would be mailed out to all vehicle registrants, allotting them a certain amount of ration rights. These checks would then be cashed at Coupon Issuance Points, where the bearer would receive ration coupons to be used at gasoline stations. Large users of gasoline could deposit their allotment checks in accounts at ration banks. Coupons or checks would be freely exchangeable in a white market. A certain percentage of the gasoline supply would be set aside in reserve for use in national emergencies. When the plan was published in the Federal Register, public comments were requested. DOE also solicited comments from private citizens, public interest groups, business and industry, state and local governments. A total of 1126 responses were reveived and these are analyzed in this paper. The second part of the report describes how the comments were classified, and gives a statistical breakdown of the major responses. The last section is a discussion and analysis of theissue raised by commenting agencies, firms, associations, and individuals. (MCW)

Privacy-Enhancing Auctions Using Rational Cryptography

DEFF Research Database (Denmark)

Miltersen, Peter Bro; Nielsen, Jesper Buus; Triandopoulos, Nikolaos

2009-01-01

show how to use rational cryptography to approximately implement any given ex interim individually strictly rational equilibrium of such an auction without a trusted mediator through a cryptographic protocol that uses only point-to-point authenticated channels between the players. By “ex interim...

Principles of Economic Rationality in Mice.

Science.gov (United States)

Rivalan, Marion; Winter, York; Nachev, Vladislav

2017-12-12

Humans and non-human animals frequently violate principles of economic rationality, such as transitivity, independence of irrelevant alternatives, and regularity. The conditions that lead to these violations are not completely understood. Here we report a study on mice tested in automated home-cage setups using rewards of drinking water. Rewards differed in one of two dimensions, volume or probability. Our results suggest that mouse choice conforms to the principles of economic rationality for options that differ along a single reward dimension. A psychometric analysis of mouse choices further revealed that mice responded more strongly to differences in probability than to differences in volume, despite equivalence in return rates. This study also demonstrates the synergistic effect between the principles of economic rationality and psychophysics in making quantitative predictions about choices of healthy laboratory mice. This opens up new possibilities for the analyses of multi-dimensional choice and the use of mice with cognitive impairments that may violate economic rationality.

Elastin-like polypeptide switches: A design strategy to detect multimeric proteins.

Science.gov (United States)

Dhandhukia, Jugal P; Brill, Dab A; Kouhi, Aida; Pastuszka, Martha K; MacKay, J Andrew

2017-09-01

Elastin-Like Polypeptides (ELPs) reversibly phase separate in response to changes in temperature, pressure, concentration, pH, and ionic species. While powerful triggers, biological microenvironments present a multitude of more specific biological cues, such as antibodies, cytokines, and cell-surface receptors. To develop better biosensors and bioresponsive drug carriers, rational strategies are required to sense and respond to these target proteins. We recently reported that noncovalent association of two ELP fusion proteins to a "chemical inducer of dimerization" small molecule (1.5 kDa) induces phase separation at physiological temperatures. Having detected a small molecule, here we present the first evidence that ELP multimerization can also detect a much larger (60 kDa) protein target. To demonstrate this strategy, ELPs were biotinylated at their amino terminus and mixed with tetrameric streptavidin. At a stoichiometric ratio of [4:1], two to three biotin-ELPs associate with streptavidin into multimeric complexes with an apparent K d of 5 nM. The increased ELP density around a streptavidin core strongly promotes isothermal phase separation, which was tuned to occur at physiological temperature. This phase separation reverses upon saturation with excess streptavidin, which only favors [1:1] complexes. Together, these findings suggest that ELP association with multimeric biomolecules is a viable strategy to deliberately engineer ELPs that respond to multimeric protein substrates. © 2017 The Protein Society.

Engineered mutations in fibrillin-1 leading to Marfan syndrome act at the protein, cellular and organismal levels.

Science.gov (United States)

Zeyer, Karina A; Reinhardt, Dieter P

2015-01-01

Fibrillins are the major components of microfibrils in the extracellular matrix of elastic and non-elastic tissues. They are multi-domain proteins, containing primarily calcium binding epidermal growth factor-like (cbEGF) domains and 8-cysteine/transforming growth factor-beta binding protein-like (TB) domains. Mutations in the fibrillin-1 gene give rise to Marfan syndrome, a connective tissue disorder with clinical complications in the cardiovascular, skeletal, ocular and other organ systems. Here, we review the consequences of engineered Marfan syndrome mutations in fibrillin-1 at the protein, cellular and organismal levels. Representative point mutations associated with Marfan syndrome in affected individuals have been introduced and analyzed in recombinant fibrillin-1 fragments. Those mutations affect fibrillin-1 on a structural and functional level. Mutations which impair folding of cbEGF domains can affect protein trafficking. Protein folding disrupted by some mutations can lead to defective secretion in mutant fibrillin-1 fragments, whereas fragments with other Marfan mutations are secreted normally. Many Marfan mutations render fibrillin-1 more susceptible to proteolysis. There is also evidence that some mutations affect heparin binding. Few mutations have been further analyzed in mouse models. An extensively studied mouse model of Marfan syndrome expresses mouse fibrillin-1 with a missense mutation (p.C1039G). The mice display similar characteristics to human patients with Marfan syndrome. Overall, the analyses of engineered mutations leading to Marfan syndrome provide important insights into the pathogenic molecular mechanisms exerted by mutated fibrillin-1. Copyright © 2015 Elsevier B.V. All rights reserved.

Decision Making: Rational, Nonrational, and Irrational.

Science.gov (United States)

Simon, Herbert A.

1993-01-01

Describes the current state of knowledge about human decision-making and problem-solving processes, explaining recent developments and their implications for management and management training. Rational goal-setting is the key to effective decision making and accomplishment. Bounded rationality is a realistic orientation, because the world is too…

Dietary protein level and performance of growing Baladi kids.

Science.gov (United States)

Abdelrahman, M M; Aljumaah, R S

2014-01-01

A study was conducted to evaluate the effect of feeding different levels of protein to black Baladi breed kids. Weanling Baladi kids (n=18; 75 to 90 days old) were selected and individually housed at our experimental farm. Kids were divided randomly to one of the three treatments for 12 weeks. The three dietary treatments were: T1: control ration, formulated according to NRC to cover the protein (level 1) and other nutrients requirements. T2: ration formulated to cover only 75% of protein (level 2) recommended by NRC. T3: control diet + 2.4 g undegradable methionine (Smartamine®)/day/kid (level 3). Feed intake, initial and monthly body weights were recorded. Blood samples were collected monthly and analyzed for metabolites and Co, Zn and Cu levels. Decreasing the dietary level of protein (T2) negatively affected (Pkids below the NRC requirements of protein negatively affect the growth performance and feed efficiency. The recommended protein level by NRC for growing kids cover the requirements of growing black Baladi kids for maximum growth and productivity.

Simplifications of rational matrices by using UML

OpenAIRE

Tasić, Milan B.; Stanimirović, Ivan P.

2013-01-01

The simplification process on rational matrices consists of simplifying each entry represented by a rational function. We follow the classic approach of dividing the numerator and denominator polynomials by their common GCD polynomial, and provide the activity diagram in UML for this process. A rational matrix representation as the quotient of a polynomial matrix and a polynomial is also discussed here and illustrated via activity diagrams. Also, a class diagram giving the links between the c...

Free-Standing Metal Oxide Nanoparticle Superlattices Constructed with Engineered Protein Containers Show in Crystallo Catalytic Activity.

Science.gov (United States)

Lach, Marcel; Künzle, Matthias; Beck, Tobias

2017-12-11

The construction of defined nanostructured catalysts is challenging. In previous work, we established a strategy to assemble binary nanoparticle superlattices with oppositely charged protein containers as building blocks. Here, we show that these free-standing nanoparticle superlattices are catalytically active. The metal oxide nanoparticles inside the protein scaffold are accessible for a range of substrates and show oxidase-like and peroxidase-like activity. The stable superlattices can be reused for several reaction cycles. In contrast to bulk nanoparticle-based catalysts, which are prone to aggregation and difficult to characterize, nanoparticle superlattices based on engineered protein containers provide an innovative synthetic route to structurally defined heterogeneous catalysts with control over nanoparticle size and composition. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

THE ROLE OF P-GLYCOPROTEIN IN RATIONAL PHARMACOTHERAPY IN CARDIOLOGY

Directory of Open Access Journals (Sweden)

A. V. Shulkin

2015-09-01

Full Text Available On the basis of the analysis of published data the role of P-glycoprotein, carrier protein, in rational pharmacotherapy in cardiology was shown on the example of its substrates – digoxin, antiplatelet agents and anticoagulants. Determination of C3435T polymorphism of multidrug resistance gene (MDR1, encoding P-glycoprotein, in pharmacotherapy with digoxin, antiplatelet drugs (clopidogrel tikagrelol, prasugrel and anticoagulants (dabigatran etexilate, rivaroxaban, edoxaban is not feasible in routine practice. Drug in- teractions have clinical implications for the efficacy and safety of pharmacotherapy in coadministration of these drugs with P-glycoprotein substrates, inducers and inhibitors.

Engineering multifunctional protein nanoparticles by in vitro disassembling and reassembling of heterologous building blocks

Science.gov (United States)

Unzueta, Ugutz; Serna, Naroa; Sánchez-García, Laura; Roldán, Mónica; Sánchez-Chardi, Alejandro; Mangues, Ramón; Villaverde, Antonio; Vázquez, Esther

2017-12-01

The engineering of protein self-assembling at the nanoscale allows the generation of functional and biocompatible materials, which can be produced by easy biological fabrication. The combination of cationic and histidine-rich stretches in fusion proteins promotes oligomerization as stable protein-only regular nanoparticles that are composed by a moderate number of building blocks. Among other applications, these materials are highly appealing as tools in targeted drug delivery once empowered with peptidic ligands of cell surface receptors. In this context, we have dissected here this simple technological platform regarding the controlled disassembling and reassembling of the composing building blocks. By applying high salt and imidazole in combination, nanoparticles are disassembled in a process that is fully reversible upon removal of the disrupting agents. By taking this approach, we accomplish here the in vitro generation of hybrid nanoparticles formed by heterologous building blocks. This fact demonstrates the capability to generate multifunctional and/or multiparatopic or multispecific materials usable in nanomedical applications.

Integrating Protein Engineering and Bioorthogonal Click Conjugation for Extracellular Vesicle Modulation and Intracellular Delivery.

Directory of Open Access Journals (Sweden)

Ming Wang

Full Text Available Exosomes are small, cell-secreted vesicles that transfer proteins and genetic information between cells. This intercellular transmission regulates many physiological and pathological processes. Therefore, exosomes have emerged as novel biomarkers for disease diagnosis and as nanocarriers for drug delivery. Here, we report an easy-to-adapt and highly versatile methodology to modulate exosome composition and conjugate exosomes for intracellular delivery. Our strategy combines the metabolic labeling of newly synthesized proteins or glycan/glycoproteins of exosome-secreting cells with active azides and bioorthogonal click conjugation to modify and functionalize the exosomes. The azide-integrated can be conjugated to a variety of small molecules and proteins and can efficiently deliver conjugates into cells. The metabolic engineering of exosomes diversifies the chemistry of exosomes and expands the functions that can be introduced into exosomes, providing novel, powerful tools to study the roles of exosomes in biology and expand the biomedical potential of exosomes.

Local Choices: Rationality and the Contextuality of Decision-Making

Science.gov (United States)

Vlaev, Ivo

2018-01-01

Rational explanation is ubiquitous in psychology and social sciences, ranging from rational analysis, expectancy-value theories, ideal observer models, mental logic to probabilistic frameworks, rational choice theory, and informal “folk psychological” explanation. However, rational explanation appears to be challenged by apparently systematic irrationality observed in psychological experiments, especially in the field of judgement and decision-making (JDM). Here, it is proposed that the experimental results require not that rational explanation should be rejected, but that rational explanation is local, i.e., within a context. Thus, rational models need to be supplemented with a theory of contextual shifts. We review evidence in JDM that patterns of choices are often consistent within contexts, but unstable between contexts. We also demonstrate that for a limited, though reasonably broad, class of decision-making domains, recent theoretical models can be viewed as providing theories of contextual shifts. It is argued that one particular significant source of global inconsistency arises from a cognitive inability to represent absolute magnitudes, whether for perceptual variables, utilities, payoffs, or probabilities. This overall argument provides a fresh perspective on the scope and limits of human rationality. PMID:29301289

Local Choices: Rationality and the Contextuality of Decision-Making.

Science.gov (United States)

Vlaev, Ivo

2018-01-02

Rational explanation is ubiquitous in psychology and social sciences, ranging from rational analysis, expectancy-value theories, ideal observer models, mental logic to probabilistic frameworks, rational choice theory, and informal "folk psychological" explanation. However, rational explanation appears to be challenged by apparently systematic irrationality observed in psychological experiments, especially in the field of judgement and decision-making (JDM). Here, it is proposed that the experimental results require not that rational explanation should be rejected, but that rational explanation is local , i.e., within a context. Thus, rational models need to be supplemented with a theory of contextual shifts. We review evidence in JDM that patterns of choices are often consistent within contexts, but unstable between contexts. We also demonstrate that for a limited, though reasonably broad, class of decision-making domains, recent theoretical models can be viewed as providing theories of contextual shifts. It is argued that one particular significant source of global inconsistency arises from a cognitive inability to represent absolute magnitudes, whether for perceptual variables, utilities, payoffs, or probabilities. This overall argument provides a fresh perspective on the scope and limits of human rationality.

Techno-Optimism and Rational Superstition