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1

Heterogeneous properties of individual molecules of beta-galactosidase from the thermophilic bacteria Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Single enzyme molecule assays were performed on beta-galactosidase from the thermophilic bacteria Geobacillus stearothermophilus using a capillary electrophoresis-based continuous flow assay and the substrate DDAO-beta-D: -galactoside. The enzyme was found to be heterogeneous with respect to catalytic rate, electrophoretic mobility and activation energy of catalysis. Catalytic rate was also found to vary over time for individual molecules at elevated temperature. Comparison with beta-galactosidase from the mesophilic bacteria Escherichia coli showed that the variation in activity over time was less pronounced and the average activation energy of catalysis was lower for the Geobacillus stearothermophilus enzyme. Attempts to measure the properties of individual beta-galactosidase molecules from the thermophilic bacteria Thermus thermophilus and the cold-adapted bacteria Pseudoalteromas haloplanktis using this assay were unsuccessful.

Craig DB

2010-01-01

2

Cadmium ion biosorption by the thermophilic bacteria Geobacillus stearothermophilus and G. thermocatenulatus.  

UK PubMed Central (United Kingdom)

This study reports surface complexation models (SCMs) for quantifying metal ion adsorption by thermophilic microorganisms. In initial cadmium ion toxicity tests, members of the genus Geobacillus displayed the highest tolerance to CdCl2 (as high as 400 to 3,200 microM). The thermophilic, gram-positive bacteria Geobacillus stearothermophilus and G. thermocatenulatus were selected for further electrophoretic mobility, potentiometric titration, and Cd2+ adsorption experiments to characterize Cd2+ complexation by functional groups within and on the cell wall. Distinct one-site SCMs described the extent of cadmium ion adsorption by both studied Geobacillus sp. strains over a range of pH values and metal/bacteria concentration ratios. The results indicate that a functional group with a deprotonation constant pK value of approximately 3.8 accounts for 66% and 80% of all titratable sites for G. thermocatenulatus and G. stearothermophilus, respectively, and is dominant in Cd2+ adsorption reactions. The results suggest a different type of functional group may be involved in cadmium biosorption for both thermophilic strains investigated here, compared to previous reports for mesophilic bacteria.

Hetzer A; Daughney CJ; Morgan HW

2006-06-01

3

Cadmium ion biosorption by the thermophilic bacteria Geobacillus stearothermophilus and G. thermocatenulatus.  

Science.gov (United States)

This study reports surface complexation models (SCMs) for quantifying metal ion adsorption by thermophilic microorganisms. In initial cadmium ion toxicity tests, members of the genus Geobacillus displayed the highest tolerance to CdCl2 (as high as 400 to 3,200 microM). The thermophilic, gram-positive bacteria Geobacillus stearothermophilus and G. thermocatenulatus were selected for further electrophoretic mobility, potentiometric titration, and Cd2+ adsorption experiments to characterize Cd2+ complexation by functional groups within and on the cell wall. Distinct one-site SCMs described the extent of cadmium ion adsorption by both studied Geobacillus sp. strains over a range of pH values and metal/bacteria concentration ratios. The results indicate that a functional group with a deprotonation constant pK value of approximately 3.8 accounts for 66% and 80% of all titratable sites for G. thermocatenulatus and G. stearothermophilus, respectively, and is dominant in Cd2+ adsorption reactions. The results suggest a different type of functional group may be involved in cadmium biosorption for both thermophilic strains investigated here, compared to previous reports for mesophilic bacteria. PMID:16751511

Hetzer, Adrian; Daughney, Christopher J; Morgan, Hugh W

2006-06-01

4

Biosynthesis of ?-alicyclic fatty acids induced by cyclic precursors and change of membrane fluidity in thermophilic bacteria Geobacillus stearothermophilus and Meiothermus ruber.  

UK PubMed Central (United Kingdom)

Two thermophilic strains belonging to Geobacillus stearothermophilus and Meiothermus ruber, which naturally do not synthesize ?-alicyclic fatty acids (?-FAs) were cultivated with cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl carboxylic acids. Gas chromatography-mass spectrometry analysis of fatty acid methyl and picolinyl esters showed that both strains are able to synthesize ?-FAs when cultivated with the appropriate precursor. The incorporation of cyclic acids influenced the whole FA composition as well as membrane fluidity. Membrane fluidity of intact cells was studied by measuring the fluorescence polarisation of the probe l,6-diphenyl-1,3,5-hexatriene incorporated into membrane lipid bilayers. Cytoplasmic membrane became more fluid with increasing content of ?-FAs. This is caused by considerable changes in lipid packing within the membrane induced by the presence of ?-FAs not found in the natural environment of Geobacillus and Meiothermus strains.

Siristova L; Luhovy R; Sigler K; Rezanka T

2011-05-01

5

Transformation of chenodeoxycholic acid by thermophilic Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

We performed a series of experiments with Geobacillus stearothermophilus, a thermophile isolated from oil-contaminated soil in the Kuwaiti desert. The organism has a good potential for the transformation of a broad spectrum of organic molecules such as steroids, amino acids, and aromatic hydrocarbons. In the present study, we tested its potential for the transformation of a bile component, chenodeoxycholic acid (CDCA). Five transformed products, namely, cholic acid, methylcholate, methylchenodeoxycholate, 3?-hydroxy-7-oxo-5?-cholanic acid, and 7?-hydroxy-3-oxo-5?-cholanic acid, were the major transformation products catalyzed by G. stearothermophilus. Under aerobic conditions, no evidence of side chain degradation, ring cleavage, or dehydrogenation was found among the metabolites of CDCA. CDCA transformation by a thermophile is reported for the first time.

Afzal M; Oommen S; Al-Awadi S

2011-07-01

6

Biotransformation of 2,6-diaminopurine nucleosides by immobilized Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

An efficient and green bioprocess to obtain 2,6-diaminopurine nucleosides using thermophilic bacteria is herein reported. Geobacillus stearothermophilus CECT 43 showed a conversion rate of 90 and 83% at 2 h to obtain 2,6-diaminopurine-2'-deoxyriboside and 2,6-diaminopurine riboside, respectively. The selected biocatalyst was successfully stabilized in an agarose matrix and used to produce up to 23.4 g of 2,6-diaminopurine-2'-deoxyriboside in 240 h of process. These nucleoside analogues can be used as prodrug precursors or in antisense oligonucleotide synthesis.

De Benedetti EC; Rivero CW; Britos CN; Lozano ME; Trelles JA

2012-09-01

7

Biotransformation of 2,6-diaminopurine nucleosides by immobilized Geobacillus stearothermophilus.  

Science.gov (United States)

An efficient and green bioprocess to obtain 2,6-diaminopurine nucleosides using thermophilic bacteria is herein reported. Geobacillus stearothermophilus CECT 43 showed a conversion rate of 90 and 83% at 2 h to obtain 2,6-diaminopurine-2'-deoxyriboside and 2,6-diaminopurine riboside, respectively. The selected biocatalyst was successfully stabilized in an agarose matrix and used to produce up to 23.4 g of 2,6-diaminopurine-2'-deoxyriboside in 240 h of process. These nucleoside analogues can be used as prodrug precursors or in antisense oligonucleotide synthesis. PMID:22837142

De Benedetti, Eliana C; Rivero, Cintia W; Britos, Claudia N; Lozano, Mario E; Trelles, Jorge A

2012-08-28

8

FORMALDEHYDE GAS INACTIVATION OF BACILLUS ANTHRACIS, BACILLUS SUBTILIS AND GEOBACILLUS STEAROTHERMOPHILUS SPORES ON INDOOR SURFACE MATERIALS.  

Science.gov (United States)

Research evaluated the decontamination of Bacillus anthracis, Bacillus subtilis, and Geobacillus stearothermophilus spores on indoor surface material using formaldehyde gas. Spores were dried on seven types of indoor surfaces and exposed to 1100 ppm formaldehyde gas for 10 hr. Fo...

9

Accelerated inactivation of Geobacillus stearothermophilus spores by ohmic heating  

UK PubMed Central (United Kingdom)

Until recently, ohmic heating was commonly thought to kill microorganisms through a thermal effect. However a growing body of evidence suggests that non-thermal effects may occur. Our aim was to determine the kinetics of inactivation of Geobacillus stearothermophilus spores (ATCC 7953) under ohmic and conventional heating using a specially constructed test chamber with capillary sized cells to eliminate potential sources of error and ensure that identical thermal histories were experienced both by conventionally and ohmically heated samples. Ohmic treatments at frequencies of 60Hz and 10kHz were compared with conventional heating at 121, 125 and 130°C for four different holding times. Both ohmic treatments showed a general trend of accelerated spore inactivation. It is hypothesized that vibration of polar dipicolinic acid molecules (DPA) and spore proteins to electric fields at high temperature conditions may result in the accelerated inactivation.

Somavat R; Mohamed HMH; Chung YK; Yousef AE; Sastry SK

2012-01-01

10

DECONTAMINATION ASSESSMENT OF BACILLUS ANTHRACIS, BACILLUS SUBTILIS, AND GEOBACILLUS STEAROTHERMOPHILUS SPORES ON INDOOR SURFACTS USING A HYDROGEN PERIOXIDE GAS GENERATOR  

Science.gov (United States)

Aims: To evaluate the decontamination of Bacillus anthracis, Bacillus subtilis, and Geobacillus stearothermophilus spores on indoor surface materials using hydrogen peroxide gas. Methods and Results: B. anthracis, B. subtilis, and G. Stearothermophilus spores were dried on seven...

11

Functional characterization of the galactan utilization system of Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

UNLABELLED: Type I galactan is a pectic polysaccharide composed of ?-1,4 linked units of d-galactose and is part of the main plant cell wall polysaccharides, which are the most abundant sources of renewable carbon in the biosphere. The thermophilic bacterium Geobacillus stearothermophilus T-6 possesses an extensive system for the utilization of plant cell wall polysaccharides, including a 9.4-kb gene cluster, ganREFGBA, which encodes galactan-utilization elements. Based on enzyme activity assays, the ganEFGBA genes, which probably constitute an operon, are induced by short galactosaccharides but not by galactose. GanA is a glycoside hydrolase family 53 ?-1,4-galactanase, active on high molecular weight galactan, producing galactotetraose as the main product. Homology modelling of the active site residues suggests that the enzyme can accommodate at least eight galactose molecules (at subsites -4 to +4) in the active site. GanB is a glycoside hydrolase family 42 ?-galactosidase capable of hydrolyzing short ?-1,4 galactosaccharides into galactose. Applying both GanA and GanB on galactan resulted in the full degradation of the polymer into galactose. The ganEFG genes encode an ATP-binding cassette sugar transport system whose sugar-binding lipoprotein, GanE, was shown to bind galacto-oligosaccharides. The utilization of galactan by G. stearothermophilus involves the extracellular galactanase GanA cleaving galactan into galacto-oligosaccharides that enter the cell via a specific transport system GanEFG. The galacto-oligosaccharides are further degraded by the intracellular ?-galactosidase GanB into galactose, which is then metabolized into UDP-glucose via the Leloir pathway by the galKET gene products. DATABASE: Nucleotide sequence data have been deposited in the GenBank database under the accession number JF327803.

Tabachnikov O; Shoham Y

2013-02-01

12

Safety evaluation of a thermolysin enzyme produced from Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Thermolysin is a zinc metalloprotease that has potential uses in the food industry. The safety of thermolysin has not been demonstrated before, and therefore a series of standard toxicological tests to assess its potential toxicity was undertaken. The thermolysin used in this study was derived from the thermophilic bacterium Geobacillus stearothermophilus, which had undergone chemical mutagenesis to generate strains with increased thermolysin production. Acute toxicity studies in rats and mice showed that thermolysin powder is not acutely toxic with an oral LD50 of more than 18,000mg/kg (2520mg/kg thermolysin protein) in rats and more than 24,000mg/kg (3360mg/kg protein) in mice. Subchronic feeding studies in rats for 91days at doses up to 1000mg/kg (390mg/kg protein) revealed no significant differences between treated and non-treated groups and a No Observed Effect Level (NOEL) of 1000mg/kg (390mg/kg protein) per day was established. Results from genotoxicity tests such as in vitro chromosomal aberration assay and in vivo mouse micronucleus were negative. Allergenicity sequence analysis revealed no evidence suggesting that thermolysin is an allergen. The data presented in this study support the conclusion that thermolysin is safe for use in food production.

Ke Q; Chen A; Minoda M; Yoshida H

2013-09-01

13

Safety evaluation of a thermolysin enzyme produced from Geobacillus stearothermophilus.  

Science.gov (United States)

Thermolysin is a zinc metalloprotease that has potential uses in the food industry. The safety of thermolysin has not been demonstrated before, and therefore a series of standard toxicological tests to assess its potential toxicity was undertaken. The thermolysin used in this study was derived from the thermophilic bacterium Geobacillus stearothermophilus, which had undergone chemical mutagenesis to generate strains with increased thermolysin production. Acute toxicity studies in rats and mice showed that thermolysin powder is not acutely toxic with an oral LD50 of more than 18,000 mg/kg (2520 mg/kg thermolysin protein) in rats and more than 24,000 mg/kg (3360 mg/kg protein) in mice. Subchronic feeding studies in rats for 91 days at doses up to 1000 mg/kg (390 mg/kg protein) revealed no significant differences between treated and non-treated groups and a No Observed Effect Level (NOEL) of 1000 mg/kg (390 mg/kg protein) per day was established. Results from genotoxicity tests such as in vitro chromosomal aberration assay and in vivo mouse micronucleus were negative. Allergenicity sequence analysis revealed no evidence suggesting that thermolysin is an allergen. The data presented in this study support the conclusion that thermolysin is safe for use in food production. PMID:23831195

Ke, Qingdong; Chen, Alice; Minoda, Masashi; Yoshida, Hiromichi

2013-07-03

14

The L-Arabinan utilization system of Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Geobacillus stearothermophilus T-6 is a thermophilic soil bacterium that has a 38-kb gene cluster for the utilization of arabinan, a branched polysaccharide that is part of the plant cell wall. The bacterium encodes a unique three-component regulatory system (araPST) that includes a sugar-binding lipoprotein (AraP), a histidine sensor kinase (AraS), and a response regulator (AraT) and lies adjacent to an ATP-binding cassette (ABC) arabinose transport system (araEGH). The lipoprotein (AraP) specifically bound arabinose, and gel mobility shift experiments showed that the response regulator, AraT, binds to a 139-bp fragment corresponding to the araE promoter region. Taken together, the results showed that the araPST system appeared to sense extracellular arabinose and to activate a specific ABC transporter for arabinose (AraEGH). The promoter regions of the arabinan utilization genes contain a 14-bp inverted repeat motif resembling an operator site for the arabinose repressor, AraR. AraR was found to bind specifically to these sequences, and binding was efficiently prevented in the presence of arabinose, suggesting that arabinose is the molecular inducer of the arabinan utilization system. The expression of the arabinan utilization genes was reduced in the presence of glucose, indicating that regulation is also mediated via a catabolic repression mechanism. The cluster also encodes a second putative ABC sugar transporter (AbnEFJ) whose sugar-binding lipoprotein (AbnE) was shown to interact specifically with linear and branched arabino-oligosaccharides. The final degradation of the arabino-oligosaccharides is likely carried out by intracellular enzymes, including two ?-l-arabinofuranosidases (AbfA and AbfB), a ?-l-arabinopyranosidase (Abp), and an arabinanase (AbnB), all of which are encoded in the 38-kb cluster.

Shulami S; Raz-Pasteur A; Tabachnikov O; Gilead-Gropper S; Shner I; Shoham Y

2011-06-01

15

A method of increasing test range and accuracy of bioindicators: Geobacillus stearothermophilus spores.  

UK PubMed Central (United Kingdom)

Spores of Geobacillus stearothermophilus are very sensitive to changes in temperature. When validating sterilizing processes, the most common bioindicator (BI) is spores of Geobacillus stearothermophilus ATCC12980 and ATCC7953 with about 10(6) spores /BI and a D121-value of about 2 minutes in water. Because these spores of Geobacillus stearothermophilus do not survive at a F0-value above 12 minutes, it has not been possible to evaluate the agreement between the biological F-value (F(BIO)) and physical measurements (time and temperature) when the physical F0-value exceeds that limit. However, it has been proven that glycerin substantially increases the heat resistance of the spores, and it is possible to utilize that property when manufacturing BIs suitable to use in processes with longer sterilization time or high temperature (above 121 degrees C). By the method described, it is possible to make use of the sensitivity and durability of Geobacillus stearothermophilus' spores when glycerin has increased both test range and accuracy. Experience from years of development and validation work with the use of the highly sensitive glycerin-water-spore-suspension sensor (GWS-sensor) is reported. Validation of the steam sterilization process at high temperature has been possible with the use of GWS-sensors. It has also been shown that the spores in suspension keep their characteristics for a period of 19 months when stored cold (8 degrees C).

Lundahl G

2003-07-01

16

A method of increasing test range and accuracy of bioindicators: Geobacillus stearothermophilus spores.  

Science.gov (United States)

Spores of Geobacillus stearothermophilus are very sensitive to changes in temperature. When validating sterilizing processes, the most common bioindicator (BI) is spores of Geobacillus stearothermophilus ATCC12980 and ATCC7953 with about 10(6) spores /BI and a D121-value of about 2 minutes in water. Because these spores of Geobacillus stearothermophilus do not survive at a F0-value above 12 minutes, it has not been possible to evaluate the agreement between the biological F-value (F(BIO)) and physical measurements (time and temperature) when the physical F0-value exceeds that limit. However, it has been proven that glycerin substantially increases the heat resistance of the spores, and it is possible to utilize that property when manufacturing BIs suitable to use in processes with longer sterilization time or high temperature (above 121 degrees C). By the method described, it is possible to make use of the sensitivity and durability of Geobacillus stearothermophilus' spores when glycerin has increased both test range and accuracy. Experience from years of development and validation work with the use of the highly sensitive glycerin-water-spore-suspension sensor (GWS-sensor) is reported. Validation of the steam sterilization process at high temperature has been possible with the use of GWS-sensors. It has also been shown that the spores in suspension keep their characteristics for a period of 19 months when stored cold (8 degrees C). PMID:14558699

Lundahl, Gunnel

17

Isolation of Lipase Gene of the Thermophilic Geobacillus stearothermophilus Strain-5  

Directory of Open Access Journals (Sweden)

Full Text Available In earlier study a new thermophilic strain Geobacillus stearothermophilus strain-5 producing thermostable lipase was isolated and identified based on 16S rRNA sequencing. Phylogenetic analysis revealed its closeness to geobacilli especially the thermophilic Geobacillus stearothermophilus with optimal growth and lipolytic enzyme activity at 60°C and pH 7.0. In this study thermostable lipase gene from this bacterium was isolated by PCR using degenerate primers. The DNA fragment coding for lipase gene was cloned in the pCR 4-TOPO plasmid and the ligation products were transformed into Escherichia coli XL1-blue cells. Partial sequencing of the gene was carried out (accession number DQ923401). Analysis by BLAST program showed some sequence similarity to that, of several lipase genes from thermophilic Geobacillus and Bacillus submitted to Genbank.

M. Sifour; H.M. Saeed; T.I. Zaghloul; M.M. Berekaa; Y.R. Abdel-Fattah

2010-01-01

18

Taxonomic revision of the genus Geobacillus: emendation of Geobacillus, G. stearothermophilus, G. jurassicus, G. toebii, G. thermodenitrificans and G. thermoglucosidans (nom. corrig., formerly 'thermoglucosidasius'); transfer of Bacillus thermantarcticus to the genus as G. thermantarcticus comb. nov.; proposal of Caldibacillus debilis gen. nov., comb. nov.; transfer of G. tepidamans to Anoxybacillus as A. tepidamans comb. nov.; and proposal of Anoxybacillus caldiproteolyticus sp. nov.  

UK PubMed Central (United Kingdom)

Sixty-two strains of thermophilic aerobic endospore-forming bacteria were subjected to polyphasic taxonomic study including 16S rRNA gene sequence analysis, polar lipid and fatty acid analysis, phenotypic characterization, and DNA-DNA hybridization experiments. Distinct clusters of the species Geobacillus stearothermophilus, Geobacillus thermodenitrificans, Geobacillus toebii and Geobacillus thermoglucosidasius were formed, allowing their descriptions to be emended, and the distinctiveness of the poorly represented species Geobacillus jurassicus, Geobacillus subterraneus and Geobacillus caldoxylosilyticus was confirmed. It is proposed that the name Geobacillus thermoglucosidasius be corrected to Geobacillus thermoglucosidans nom. corrig. Bacillus thermantarcticus clustered between Geobacillus species on the basis of 16S rRNA gene sequence analysis, and its transfer to the genus Geobacillus as Geobacillus thermantarcticus comb. nov. (type strain LMG 23032(T)=DSM 9572(T)=strain M1(T)=R-35644(T)) is proposed. The above-mentioned species, together with Geobacillus thermoleovorans and Geobacillus thermocatenulatus, form a monophyletic cluster representing the genus Geobacillus. The distinctiveness of 'Geobacillus caldoproteolyticus' was confirmed and it is proposed that it be accommodated, along with Geobacillus tepidamans, in the genus Anoxybacillus as Anoxybacillus caldiproteolyticus sp. nov. (type strain DSM 15730(T)=ATCC BAA-818(T)=LMG 26209(T)=R-35652(T)) and Anoxybacillus tepidamans comb. nov. (type strain LMG 26208(T)=ATCC BAA-942(T)=DSM 16325(T)=R-35643(T)), respectively. The type strain of Geobacillus debilis was not closely related to any members of the genera Anoxybacillus and Geobacillus, and it is proposed that this species be placed in the new genus Caldibacillus as Caldibacillus debilis gen. nov. comb. nov. The type strain of the type species, Caldibacillus debilis, is LMG 23386(T) (=DSM 16016(T)=NCIMB 13995(T)=Tf(T)=R-35653(T)).

Coorevits A; Dinsdale AE; Halket G; Lebbe L; De Vos P; Van Landschoot A; Logan NA

2012-07-01

19

Taxonomic revision of the genus Geobacillus: emendation of Geobacillus, G. stearothermophilus, G. jurassicus, G. toebii, G. thermodenitrificans and G. thermoglucosidans (nom. corrig., formerly 'thermoglucosidasius'); transfer of Bacillus thermantarcticus to the genus as G. thermantarcticus comb. nov.; proposal of Caldibacillus debilis gen. nov., comb. nov.; transfer of G. tepidamans to Anoxybacillus as A. tepidamans comb. nov.; and proposal of Anoxybacillus caldiproteolyticus sp. nov.  

Science.gov (United States)

Sixty-two strains of thermophilic aerobic endospore-forming bacteria were subjected to polyphasic taxonomic study including 16S rRNA gene sequence analysis, polar lipid and fatty acid analysis, phenotypic characterization, and DNA-DNA hybridization experiments. Distinct clusters of the species Geobacillus stearothermophilus, Geobacillus thermodenitrificans, Geobacillus toebii and Geobacillus thermoglucosidasius were formed, allowing their descriptions to be emended, and the distinctiveness of the poorly represented species Geobacillus jurassicus, Geobacillus subterraneus and Geobacillus caldoxylosilyticus was confirmed. It is proposed that the name Geobacillus thermoglucosidasius be corrected to Geobacillus thermoglucosidans nom. corrig. Bacillus thermantarcticus clustered between Geobacillus species on the basis of 16S rRNA gene sequence analysis, and its transfer to the genus Geobacillus as Geobacillus thermantarcticus comb. nov. (type strain LMG 23032(T)=DSM 9572(T)=strain M1(T)=R-35644(T)) is proposed. The above-mentioned species, together with Geobacillus thermoleovorans and Geobacillus thermocatenulatus, form a monophyletic cluster representing the genus Geobacillus. The distinctiveness of 'Geobacillus caldoproteolyticus' was confirmed and it is proposed that it be accommodated, along with Geobacillus tepidamans, in the genus Anoxybacillus as Anoxybacillus caldiproteolyticus sp. nov. (type strain DSM 15730(T)=ATCC BAA-818(T)=LMG 26209(T)=R-35652(T)) and Anoxybacillus tepidamans comb. nov. (type strain LMG 26208(T)=ATCC BAA-942(T)=DSM 16325(T)=R-35643(T)), respectively. The type strain of Geobacillus debilis was not closely related to any members of the genera Anoxybacillus and Geobacillus, and it is proposed that this species be placed in the new genus Caldibacillus as Caldibacillus debilis gen. nov. comb. nov. The type strain of the type species, Caldibacillus debilis, is LMG 23386(T) (=DSM 16016(T)=NCIMB 13995(T)=Tf(T)=R-35653(T)). PMID:21856988

Coorevits, An; Dinsdale, Anna E; Halket, Gillian; Lebbe, Liesbeth; De Vos, Paul; Van Landschoot, Anita; Logan, Niall A

2011-08-19

20

Inactivation kinetics of Geobacillus stearothermophilus spores in water using high-pressure processing at elevated temperatures.  

UK PubMed Central (United Kingdom)

Establishment of a high-pressure sterilization process requires data on pressure and temperature-dependent inactivation kinetics of target pathogenic, spoilage, or surrogate spore-forming bacteria in the food being tested. The objective of this study was to examine the response of Geobacillus stearothermophilus ATCC 10149 spores to various temperature, time, and pressure combination treatments (500 to 700 MPa; 92 degrees C to 111 degrees C, 0.01 to 360 s). The pressure inactivation of spores was characterized at elevated temperatures under isobaric and isothermal conditions during the holding time using a laboratory-scale high-pressure unit. The inactivation kinetics was well described by the log-linear regression model. As expected, the rate of spore inactivation increased with increasing pressure and temperature. Decimal reduction times at constant pressure (D(T,P) values) varied from 29.4 to 108.8 s at 92 degrees C, 17.4 to 76 s at 100 degrees C, and 6.1 to 51.3 s at 111 degrees C within the pressure range of 500 to 700 MPa. The resistance of spores to temperature and pressure was characterized with z(T) and z(P) values and compared with their resistance to conventional steam heating. The conventional thermal resistance of G. stearothermophilus species did not correlate to the thermal resistance at high pressure. The study provides kinetic data on the effects of pressure and temperature on the inactivation of a heat-resistant bacterial spore species under conditions applicable to the commercial processing of low-acid foods.

Patazca E; Koutchma T; Ramaswamy HS

2006-04-01

 
 
 
 
21

Properties of geobacillus stearothermophilus levansucrase as potential biocatalyst for the synthesis of levan and fructooligosaccharides.  

UK PubMed Central (United Kingdom)

The production of levansucrase (LS) by thermophilic Geobacillus stearothermophilus was investigated. LS production was more effective in the presence of sucrose (1%, w/v) than fructose, glucose, glycerol or raffinose. The results (Top 57°C; stable for 6h at 47°C) indicate the high stability of the transfructosylation activity of G. stearothermophilus LS as compared to LSs from other microbial sources. Contrary to temperature, the pH had a significant effect on the selectivity of G. stearothermophilus LS-catalyzed reaction, favoring the transfructosylation reaction in the pH range of 6.0 to 6.5. The kinetic parameter study revealed that the catalytic efficiency of transfructosylation activity was higher as compared to the hydrolytic one. In addition to levan, G. stearothermophilus LS synthesized fructooligosaccharides in the presence of sucrose as the sole substrate. The results also demonstrated the wide acceptor specificity of G. stearothermophilus LS with maltose being the best fructosyl acceptor. This study is the first on the catalytic properties and the acceptor specificity of LS from G. stearothermophilus. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 2013.

Inthanavong L; Tian F; Khodadadi M; Karboune S

2013-08-01

22

Inactivation of Geobacillus stearothermophilus spores by high-pressure carbon dioxide treatment.  

UK PubMed Central (United Kingdom)

High-pressure CO2 treatment has been studied as a promising method for inactivating bacterial spores. In the present study, we compared this method with other sterilization techniques, including heat and pressure treatment. Spores of Bacillus coagulans, Bacillus subtilis, Bacillus cereus, Bacillus licheniformis, and Geobacillus stearothermophilus were subjected to CO2 treatment at 30 MPa and 35 degrees C, to high-hydrostatic-pressure treatment at 200 MPa and 65 degrees C, or to heat treatment at 0.1 MPa and 85 degrees C. All of the bacterial spores except the G. stearothermophilus spores were easily inactivated by the heat treatment. The highly heat- and pressure-resistant spores of G. stearothermophilus were not the most resistant to CO2 treatment. We also investigated the influence of temperature on CO2 inactivation of G. stearothermophilus. Treatment with CO2 and 30 MPa of pressure at 95 degrees C for 120 min resulted in 5-log-order spore inactivation, whereas heat treatment at 95 degrees C for 120 min and high-hydrostatic-pressure treatment at 30 MPa and 95 degrees C for 120 min had little effect. The activation energy required for CO2 treatment of G. stearothermophilus spores was lower than the activation energy for heat or pressure treatment. Although heat was not necessary for inactivationby CO2 treatment of G. stearothermophilus spores, CO2 treatment at 95 degrees C was more effective than treatment at 95 degrees C alone.

Watanabe T; Furukawa S; Hirata J; Koyama T; Ogihara H; Yamasaki M

2003-12-01

23

Inactivation of Geobacillus stearothermophilus spores by high-pressure carbon dioxide treatment.  

Science.gov (United States)

High-pressure CO2 treatment has been studied as a promising method for inactivating bacterial spores. In the present study, we compared this method with other sterilization techniques, including heat and pressure treatment. Spores of Bacillus coagulans, Bacillus subtilis, Bacillus cereus, Bacillus licheniformis, and Geobacillus stearothermophilus were subjected to CO2 treatment at 30 MPa and 35 degrees C, to high-hydrostatic-pressure treatment at 200 MPa and 65 degrees C, or to heat treatment at 0.1 MPa and 85 degrees C. All of the bacterial spores except the G. stearothermophilus spores were easily inactivated by the heat treatment. The highly heat- and pressure-resistant spores of G. stearothermophilus were not the most resistant to CO2 treatment. We also investigated the influence of temperature on CO2 inactivation of G. stearothermophilus. Treatment with CO2 and 30 MPa of pressure at 95 degrees C for 120 min resulted in 5-log-order spore inactivation, whereas heat treatment at 95 degrees C for 120 min and high-hydrostatic-pressure treatment at 30 MPa and 95 degrees C for 120 min had little effect. The activation energy required for CO2 treatment of G. stearothermophilus spores was lower than the activation energy for heat or pressure treatment. Although heat was not necessary for inactivationby CO2 treatment of G. stearothermophilus spores, CO2 treatment at 95 degrees C was more effective than treatment at 95 degrees C alone. PMID:14660357

Watanabe, Taisuke; Furukawa, Soichi; Hirata, Junichi; Koyama, Tetsuya; Ogihara, Hirokazu; Yamasaki, Makari

2003-12-01

24

Structural basis of substrate binding in WsaF, a rhamnosyltransferase from Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Carbohydrate polymers are medically and industrially important. The S-layer of many Gram-positive organisms comprises protein and carbohydrate polymers and forms an almost paracrystalline array on the cell surface. Not only is this array important for the bacteria but it has potential application in the manufacture of commercially important polysaccharides and glycoconjugates as well. The S-layer glycoprotein glycan from Geobacillus stearothermophilus NRS 2004/3a is mainly composed of repeating units of three rhamnose sugars linked by alpha-1,3-, alpha-1,2-, and beta-1,2-linkages. The formation of the beta-1,2-linkage is catalysed by the enzyme WsaF. The rational use of this system is hampered by the fact that WsaF and other enzymes in the pathway share very little homology to other enzymes. We report the structural and biochemical characterisation of WsaF, the first such rhamnosyltransferase to be characterised. Structural work was aided by the surface entropy reduction method. The enzyme has two domains, the N-terminal domain, which binds the acceptor (the growing rhamnan chain), and the C-terminal domain, which binds the substrate (dTDP-beta-l-rhamnose). The structure of WsaF bound to dTDP and dTDP-beta-l-rhamnose coupled to biochemical analysis identifies the residues that underlie catalysis and substrate recognition. We have constructed and tested by site-directed mutagenesis a model for acceptor recognition.

Steiner K; Hagelueken G; Messner P; Schäffer C; Naismith JH

2010-03-01

25

Novel thermostable and alkalitolerant amylase production by Geobacillus stearothermophilus HP 3.  

UK PubMed Central (United Kingdom)

This study reports the production and characterisation of a novel thermostable alkaline amylase from the newly isolated Geobacillus stearothermophilus HP 3. The optimum temperature and pH for enzyme production was 55°C and 9.0, respectively. The amylase powder obtained from the culture filtrate by pre-chilled acetone treatment was stable over a wide pH range and liquefied thick starch slurries at 75°C. Ca²? and Co²? were required for the thermostability of the enzyme preparation. The present purified amylase therefore could be defined as thermostable and alkalitolerant with new properties that make the enzyme applicable for many starch processing applications for use in the food industry.

Selim SA

2012-01-01

26

Kinetics of CO Recombination to the Heme in Geobacillus Stearothermophilus Nitric Oxide Synthase.  

UK PubMed Central (United Kingdom)

We report the kinetics of CO rebinding to the heme in His134Ser, Ile223Val and His134Ser/Ile223Ser mutants of Geobacillus stearothermophilus nitric oxide synthase (gsNOS). The amplitudes of the two observed kinetics phases, which are insensitive to CO concentration, depend on enzyme concentration. We suggest that two forms of gsNOS are in equilibrium under the conditions employed (6.1-27 µM gsNOS with 20 or 100% CO atmosphere). The kinetics of CO rebinding to the heme do not depend on the identity of the NO-gate residues at positions 134 and 223.

Whited CA; Warren JJ; Lavoie KD; Winkler JR; Gray HB

2013-07-01

27

Formaldehyde gas inactivation of Bacillus anthracis, Bacillus subtilis, and Geobacillus stearothermophilus spores on indoor surface materials.  

UK PubMed Central (United Kingdom)

AIMS: To evaluate the decontamination of Bacillus anthracis, Bacillus subtilis, and Geobacillus stearothermophilus spores on indoor surface materials using formaldehyde gas. METHODS AND RESULTS: B. anthracis, B. subtilis, and G. stearothermophilus spores were dried on seven types of indoor surfaces and exposed to approx. 1100 ppm formaldehyde gas for 10 h. Formaldehyde exposure significantly decreased viable B. anthracis, B. subtilis, and G. stearothermophilus spores on all test materials. Significant differences were observed when comparing the reduction in viable spores of B. anthracis with B. subtilis (galvanized metal and painted wallboard paper) and G. stearothermophilus (industrial carpet and painted wallboard paper). Formaldehyde gas inactivated>or=50% of the biological indicators and spore strips (approx. 1x10(6) CFU) when analyzed after 1 and 7 days. CONCLUSIONS: Formaldehyde gas significantly reduced the number of viable spores on both porous and nonporous materials in which the two surrogates exhibited similar log reductions to that of B. anthracis on most test materials. SIGNIFICANCE AND IMPACT OF THE STUDY: These results provide new comparative information for the decontamination of B. anthracis spores with surrogates on indoor surfaces using formaldehyde gas.

Rogers JV; Choi YW; Richter WR; Rudnicki DC; Joseph DW; Sabourin CL; Taylor ML; Chang JC

2007-10-01

28

A group IIC-type intron interrupts the rRNA methylase gene of Geobacillus stearothermophilus strain 10.  

UK PubMed Central (United Kingdom)

Group IIC introns insert next to the stem-loop structure of rho-independent transcription terminators, thus avoiding intact genes. The insertion sites of 17 copies of the G.st.I1 intron from Geobacillus stearothermophilus were compared. One copy of the intron was found to interrupt an open reading frame (ORF) encoding an rRNA methylase.

Moretz SE; Lampson BC

2010-10-01

29

Structural Analysis of Xylanase from Marine Thermophilic Geobacillus stearothermophilus in Tanjung Api, Poso, Indonesia  

Directory of Open Access Journals (Sweden)

Full Text Available A xylanase gene, xynA, has been cloned from thermophilic strain Geobacillus stearothermophilus, which was isolated from marine Tanjung Api, Indonesia. The polymerase chain reaction product of 1266 bp of xynA gene consisted of 1221 bp open reading frame and encoded 407 amino acids including 30 residues of signal peptide. The sequence exhibited highest identity of 98.7% in the level of amino acid, with an extracellular endo-1,4-?-xylanase from G. stearothermophilus T-6 (E-GSX T-6) of the glycoside hydrolase family 10 (GH10). A comparative study between the local strain G. stearothermophilus (GSX L) and E-GSX T-6 on homology of amino acid sequence indicated five differents amino acids in the gene. They were Threonine/Alanine (T/A), Asparagine/Aspartate (N/D), Lysine/Asparagine (K/N), Isoleucine/Methionine (I/M), Serine/Threonine (S/T) at the position 220, 227, 228, 233, and 245, respectively. Protein structural analysis of those differences suggested that those amino acids may play role in biochemical properties such as enzyme stability, in particular its thermostability.

BUDI SAKSONO; LINDA SUKMARINI

2010-01-01

30

EFFECT OF HIGH HYDROSTATIC PRESSURE ON SPORES OF GEOBACILLUS STEAROTHERMOPHILUS SUSPENDED IN SOYMILK  

UK PubMed Central (United Kingdom)

The inactivation of Geobacillus stearothermophilus spores (ATCC 7953) inoculated in soymilk was investigated using high hydrostatic pressure (550, 585 and 620 MPa) in combination with temperature (70, 80 and 90C) for selected times (2 s to 15 min). Inactivation of spores occurred at all selected treatments. Less than 10 CFU/mL of G. stearothermophilus were observed after 7 min of treatment at 620 MPa and 90C. An increase in the inactivation rate constant, at the highest pressure, was observed, resulting in a decrease in D values at all temperatures. D values were calculated as 10.6, 6.2 and 3.5 min for 70, 80 and 90C, respectively after pressurization at 620 MPa. zp values decreased as temperature increased with values ranging from 142 to 238 MPa. The activation energy required for inactivation of G. stearothermophilus spores in soymilk, at the selected treatments, was in the range of 37.9-57.4 kJ/mol.

ESTRADA-GIRÓN Y; GUERRERO-BELTRÁN JA; SWANSON BG; BARBOSA-CÁNOVAS GV

2007-10-01

31

The effect of organic acid type on thermal inactivation of Geobacillus stearothermophilus spores  

UK PubMed Central (United Kingdom)

The effect of organic acid (either citric or acetic) on thermal inactivation of Geobacillus stearothermophilus (former Bacillus stearothermophilus) spores was analysed focusing on individuals of population with different thermal sensitivity. The survival of spores was investigated in media (tryptone solution, red beet juice) of natural pH or acidified with organic acid at pH ranging from 6.0 to 4.0. Thermal sterilisation was carried out in the temperature range from 115°C to 125°C. It was confirmed that survival curves of G. stearothermophilus spores could have three stages, which caused that the decimal reduction time was not always a proper parameter to set the conditions required for the media sterilisation. Based on performed experiments it was found out that spore inactivation in acidified red beet juice medium was faster than in acidified tryptone solution. It was shown that the presence of both acetic and citric acids in tryptone solution had a similar influence on the rate of spore inactivation. However, in the case of red beet juice, the acetic acid had a stronger effect on spore destruction than the citric acid only at pH 4.0. It was proved that medium composition could accelerate the activation of dormant spores, what resulted in elimination of “tailing” on the spore survival curve and in shorter time necessary for inactivation of particular spore population.

Iciek Jan; B?aszczyk Ilona; Papiewska Agnieszka

2008-07-01

32

The abp gene in Geobacillus stearothermophilus T-6 encodes a GH27 ?-L-arabinopyranosidase.  

Science.gov (United States)

In this study we demonstrate that the abp gene in Geobacillus stearothermophilus T-6 encodes a family 27 glycoside hydrolase ?-L-arabinopyranosidase. The catalytic constants towards the chromogenic substrate pNP-?-L-arabinopyranoside were 0.8±0.1 mM, 6.6±0.3 s(-1), and 8.2±0.3 s(-1) mM(-1) for K(m), k(cat) and k(cat)/K(m), respectively. (13)C NMR spectroscopy unequivocally showed that Abp is capable of removing ?-L-arabinopyranose residues from the natural arabino-polysaccharide, larch arabinogalactan. Most family 27 enzymes are active on galactose and contain a conserved Asp residue, whereas in Abp this residue is Ile67, which shifts the specificity of the enzyme towards arabinopyranoside. PMID:22687242

Salama, Rachel; Alalouf, Onit; Tabachnikov, Orly; Zolotnitsky, Gennady; Shoham, Gil; Shoham, Yuval

2012-06-08

33

The abp gene in Geobacillus stearothermophilus T-6 encodes a GH27 ?-L-arabinopyranosidase.  

UK PubMed Central (United Kingdom)

In this study we demonstrate that the abp gene in Geobacillus stearothermophilus T-6 encodes a family 27 glycoside hydrolase ?-L-arabinopyranosidase. The catalytic constants towards the chromogenic substrate pNP-?-L-arabinopyranoside were 0.8±0.1 mM, 6.6±0.3 s(-1), and 8.2±0.3 s(-1) mM(-1) for K(m), k(cat) and k(cat)/K(m), respectively. (13)C NMR spectroscopy unequivocally showed that Abp is capable of removing ?-L-arabinopyranose residues from the natural arabino-polysaccharide, larch arabinogalactan. Most family 27 enzymes are active on galactose and contain a conserved Asp residue, whereas in Abp this residue is Ile67, which shifts the specificity of the enzyme towards arabinopyranoside.

Salama R; Alalouf O; Tabachnikov O; Zolotnitsky G; Shoham G; Shoham Y

2012-07-01

34

Augmentation of PCR efficiency using highly thermostable gold nanoparticles synthesized from a thermophilic bacterium, Geobacillus stearothermophilus.  

Science.gov (United States)

Biogenic gold nanoparticles were synthesized from the Geobacillus stearothermophilus cell-free extract. These nanoparticles were characterized by UV-vis spectroscopy, FTIR, TEM, and XRD. The gold nanoparticles showed an absorption maximum at 522 nm. The TEM micrograph revealed the formation of monodispersed particles. The high degree of stability of the nanoparticle solution could be attributed to the secretion of certain capping proteins by the bacterium in the reaction mixture which was confirmed by the FTIR and UV-vis spectrometric analyses. The heat transfer property of the gold nanoparticles in aqueous solution has been explored in the current study for augmenting the PCR efficiency. The highly thermostable biogenic gold nanoparticles effectively increased the yield, product specificity besides reducing the reaction time of the PCR. PMID:23434707

Girilal, M; Mohammed Fayaz, A; Mohan Balaji, P; Kalaichelvan, P T

2013-01-20

35

Physicochemical characteristics of a thermostable gellan lyase from Geobacillus stearothermophilus 98.  

UK PubMed Central (United Kingdom)

A purified thermostable gellan lyase, produced by a thermophilic bacterium, Geobacillus stearothermophilus 98, was characterized in relation to its physicochemical properties. The gellan lyase was established to have a molecular weight of 216 kDa, defined by capillary gel electrophoresis. Amino acid analysis revealed high quantities of Lys, His, Ala, Val, Ile, Glx, and Pro residues. The circular dichroism revealed 45% beta-structure and practically lack of a-spiral domains. Kinetic studies showed high affinity of the enzyme to gellan as a substrate (Km = 0.21 microM). The thermal denaturation investigated by cicular dichroism showed a highly cooperative transition with a midpoint (Tm) at about 75 degrees C. A single product was identified after enzyme action on gellan. Large exothermic aggregation near Tm was observed by differential scanning calorimetry. Two types of gellan lyase crystals were reproducibly isolated.

Derekova A; Atanassova M; Christova P; Tchorbanov B; Shosheva A; Mandeva R; Rodríguez-Alonso P; Garabal JI; Kambourova M

2010-03-01

36

Augmentation of PCR efficiency using highly thermostable gold nanoparticles synthesized from a thermophilic bacterium, Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Biogenic gold nanoparticles were synthesized from the Geobacillus stearothermophilus cell-free extract. These nanoparticles were characterized by UV-vis spectroscopy, FTIR, TEM, and XRD. The gold nanoparticles showed an absorption maximum at 522 nm. The TEM micrograph revealed the formation of monodispersed particles. The high degree of stability of the nanoparticle solution could be attributed to the secretion of certain capping proteins by the bacterium in the reaction mixture which was confirmed by the FTIR and UV-vis spectrometric analyses. The heat transfer property of the gold nanoparticles in aqueous solution has been explored in the current study for augmenting the PCR efficiency. The highly thermostable biogenic gold nanoparticles effectively increased the yield, product specificity besides reducing the reaction time of the PCR.

Girilal M; Mohammed Fayaz A; Mohan Balaji P; Kalaichelvan PT

2013-06-01

37

Enhanced production of lipase by the thermophilic Geobacillus stearothermophilus strain-5 using statistical experimental designs.  

UK PubMed Central (United Kingdom)

Statistically based experimental designs were applied to optimize the cultural conditions for the production of a glycerol-inducible lipase from the thermophilic Geobacillus stearothermophilus strain-5. The effect of nineteen culture conditions on enzyme production was evaluated using Plackett-Burman factorial design. Tween 80, K(2)HPO(4), glycerol and glucose were the most significant factors in improving enzyme production. The selected parameters were then further investigated using central composite design to define the optimal process conditions. Maximal enzyme activity (578 U/ml) was reached under the following conditions: glycerol, 2.24% (v/v); Tween 80, 0.76% (v/v); glucose, 0.76% (w/v) and K(2)HPO(4), 0.38% (w/v) which is about five folds the activity in basal medium. A verification experiment was carried out to examine model validation and revealed more than 98% validity.

Sifour M; Zaghloul TI; Saeed HM; Berekaa MM; Abdel-Fattah YR

2010-09-01

38

Sporicidal Activity of the KMT reagent in its vapor phase against Geobacillus stearothermophilus Spores.  

UK PubMed Central (United Kingdom)

In an investigation of the sporicidal activity of the KMT reagent, a vapor phase study was performed using five kinds of carriers contaminated with Geobacillus stearothermophilus spores. When 25 ml of the KMT reagent was vaporized in a chamber (capacity; approximately 95 liters), the 2-step heating method (vaporization by a combination of low temperature and high temperature) showed the most effective sporicidal activity in comparison with the 1-step heating method (rapid vaporization). The 2-step heating method appeared to be related to the sporicidal activity of vaporized KMT reagent, i.e., ethanol and iodine, which vaporized mainly when heated at a low temperature such as 55 C, and acidic water, which vaporized mainly when heated at a high temperature such as 300 C. We proposed that the KMT reagent can be used as a new disinfectant not only in the liquid phase but also in the vapor phase in the same way as peracetic acid and hydrogen peroxide.

Kida N; Mochizuki Y; Taguchi F

2007-01-01

39

Sporicidal Activity of the KMT reagent in its vapor phase against Geobacillus stearothermophilus Spores.  

Science.gov (United States)

In an investigation of the sporicidal activity of the KMT reagent, a vapor phase study was performed using five kinds of carriers contaminated with Geobacillus stearothermophilus spores. When 25 ml of the KMT reagent was vaporized in a chamber (capacity; approximately 95 liters), the 2-step heating method (vaporization by a combination of low temperature and high temperature) showed the most effective sporicidal activity in comparison with the 1-step heating method (rapid vaporization). The 2-step heating method appeared to be related to the sporicidal activity of vaporized KMT reagent, i.e., ethanol and iodine, which vaporized mainly when heated at a low temperature such as 55 C, and acidic water, which vaporized mainly when heated at a high temperature such as 300 C. We proposed that the KMT reagent can be used as a new disinfectant not only in the liquid phase but also in the vapor phase in the same way as peracetic acid and hydrogen peroxide. PMID:17237604

Kida, Nori; Mochizuki, Yasushi; Taguchi, Fumiaki

2007-01-01

40

Kinetics of Germination of Individual Spores of Geobacillus stearothermophilus as Measured by Raman Spectroscopy and Differential Interference Contrast Microscopy  

Science.gov (United States)

Geobacillus stearothermophilus is a gram-positive, thermophilic bacterium, spores of which are very heat resistant. Raman spectroscopy and differential interference contrast microscopy were used to monitor the kinetics of germination of individual spores of G. stearothermophilus at different temperatures, and major conclusions from this work were as follows. 1) The CaDPA level of individual G. stearothermophilus spores was similar to that of Bacillus spores. However, the Raman spectra of protein amide bands suggested there are differences in protein structure in spores of G. stearothermophilus and Bacillus species. 2) During nutrient germination of G. stearothermophilus spores, CaDPA was released beginning after a lag time (Tlag) between addition of nutrient germinants and initiation of CaDPA release. CaDPA release was complete at Trelease, and ?Trelease (Trelease – Tlag) was 1–2 min. 3) Activation by heat or sodium nitrite was essential for efficient nutrient germination of G. stearothermophilus spores, primarily by decreasing Tlag values. 4) Values of Tlag and Trelease were heterogeneous among individual spores, but ?Trelease values were relatively constant. 5) Temperature had major effects on nutrient germination of G. stearothermophilus spores, as at temperatures below 65°C, average Tlag values increased significantly. 6) G. stearothermophilus spore germination with exogenous CaDPA or dodecylamine was fastest at 65°C, with longer Tlag values at lower temperatures. 7) Decoating of G. stearothermophilus spores slowed nutrient germination slightly and CaDPA germination significantly, but increased dodecylamine germination markedly. These results indicate that the dynamics and heterogeneity of the germination of individual G. stearothermophilus spores are generally similar to that of Bacillus species.

Zhou, Tingting; Dong, Zhiyang; Setlow, Peter; Li, Yong-qing

2013-01-01

 
 
 
 
41

Kinetics of Germination of Individual Spores of Geobacillus stearothermophilus as Measured by Raman Spectroscopy and Differential Interference Contrast Microscopy.  

UK PubMed Central (United Kingdom)

Geobacillus stearothermophilus is a gram-positive, thermophilic bacterium, spores of which are very heat resistant. Raman spectroscopy and differential interference contrast microscopy were used to monitor the kinetics of germination of individual spores of G. stearothermophilus at different temperatures, and major conclusions from this work were as follows. 1) The CaDPA level of individual G. stearothermophilus spores was similar to that of Bacillus spores. However, the Raman spectra of protein amide bands suggested there are differences in protein structure in spores of G. stearothermophilus and Bacillus species. 2) During nutrient germination of G. stearothermophilus spores, CaDPA was released beginning after a lag time (T lag) between addition of nutrient germinants and initiation of CaDPA release. CaDPA release was complete at T release, and ?T release (T release - T lag) was 1-2 min. 3) Activation by heat or sodium nitrite was essential for efficient nutrient germination of G. stearothermophilus spores, primarily by decreasing T lag values. 4) Values of T lag and T release were heterogeneous among individual spores, but ?T release values were relatively constant. 5) Temperature had major effects on nutrient germination of G. stearothermophilus spores, as at temperatures below 65°C, average T lag values increased significantly. 6) G. stearothermophilus spore germination with exogenous CaDPA or dodecylamine was fastest at 65°C, with longer Tlag values at lower temperatures. 7) Decoating of G. stearothermophilus spores slowed nutrient germination slightly and CaDPA germination significantly, but increased dodecylamine germination markedly. These results indicate that the dynamics and heterogeneity of the germination of individual G. stearothermophilus spores are generally similar to that of Bacillus species.

Zhou T; Dong Z; Setlow P; Li YQ

2013-01-01

42

Characterization of thermostable FMN-dependent NADH azoreductase from the moderate thermophile Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

The gene encoding an FMN-dependent NADH azoreductase, AzrG, from thermophilic Geobacillus stearothermophilus was cloned and functionally expressed in recombinant Escherichia coli. Purified recombinant AzrG is a homodimer of 23 kDa and bore FMN as a flavin cofactor. The optimal temperature of AzrG was 85 degrees C for the degradation of Methyl Red (MR). AzrG remained active for 1 h at 65 degrees C and for 1 month at 30 degrees C, demonstrating both superior thermostability and long-term stability of the enzyme. AzrG efficiently decolorized MR, Ethyl Red at 30 degrees C. Furthermore, the enzyme exhibited a wide-range of degrading activity towards several tenacious azo dyes, such as Acid Red 88, Orange I, and Congo Red. These results suggested the sustainable utilization of G. stearothermophilus as an azo-degrading strain for AzrG carrying whole-cell wastewater treatments for azo pollutants under high temperature conditions.

Matsumoto K; Mukai Y; Ogata D; Shozui F; Nduko JM; Taguchi S; Ooi T

2010-05-01

43

Crystallization and preliminary crystallographic analysis of Abp, a GH27 ?-L-arabinopyranosidase from Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Geobacillus stearothermophilus T-6 is a thermophilic soil bacterium that possesses an extensive system for the utilization of hemicellulose. The bacterium produces a small number of endo-acting extracellular enzymes that cleave high-molecular-weight hemicellulolytic polymers into short decorated oligosaccharides, which are further hydrolysed into the respective sugar monomers by a battery of intracellular glycoside hydrolases. One of these intracellular processing enzymes is ?-L-arabinopyranosidase (Abp), which is capable of removing ?-L-arabinopyranose residues from naturally occurring arabino-polysaccharides. As arabino-polymers constitute a significant part of the hemicellulolytic content of plant biomass, their efficient enzymatic degradation presents an important challenge for many potential biotechnological applications. This aspect has led to an increasing interest in the biochemical characterization and structural analysis of this and related hemicellulases. Abp from G. stearothermophilus T-6 has recently been cloned, overexpressed, purified, biochemically characterized and crystallized in our laboratory, as part of its complete structure-function study. The best crystals obtained for this enzyme belonged to the primitive orthorhombic space group P2(1)2(1)2(1), with average unit-cell parameters a = 107.7, b = 202.2, c = 287.3 Å. Full diffraction data sets to 2.3 Å resolution have been collected for both the wild-type enzyme and its D197A catalytic mutant from flash-cooled crystals at 100 K, using synchrotron radiation. These data are currently being used for a high-resolution three-dimensional structure determination of Abp.

Lansky S; Salama R; Solomon VH; Belrhali H; Shoham Y; Shoham G

2013-06-01

44

Crystallization and preliminary crystallographic analysis of Abp, a GH27 ?-L-arabinopyranosidase from Geobacillus stearothermophilus.  

Science.gov (United States)

Geobacillus stearothermophilus T-6 is a thermophilic soil bacterium that possesses an extensive system for the utilization of hemicellulose. The bacterium produces a small number of endo-acting extracellular enzymes that cleave high-molecular-weight hemicellulolytic polymers into short decorated oligosaccharides, which are further hydrolysed into the respective sugar monomers by a battery of intracellular glycoside hydrolases. One of these intracellular processing enzymes is ?-L-arabinopyranosidase (Abp), which is capable of removing ?-L-arabinopyranose residues from naturally occurring arabino-polysaccharides. As arabino-polymers constitute a significant part of the hemicellulolytic content of plant biomass, their efficient enzymatic degradation presents an important challenge for many potential biotechnological applications. This aspect has led to an increasing interest in the biochemical characterization and structural analysis of this and related hemicellulases. Abp from G. stearothermophilus T-6 has recently been cloned, overexpressed, purified, biochemically characterized and crystallized in our laboratory, as part of its complete structure-function study. The best crystals obtained for this enzyme belonged to the primitive orthorhombic space group P2(1)2(1)2(1), with average unit-cell parameters a = 107.7, b = 202.2, c = 287.3 Å. Full diffraction data sets to 2.3 Å resolution have been collected for both the wild-type enzyme and its D197A catalytic mutant from flash-cooled crystals at 100 K, using synchrotron radiation. These data are currently being used for a high-resolution three-dimensional structure determination of Abp. PMID:23722857

Lansky, Shifra; Salama, Rachel; Solomon, Vered H; Belrhali, Hassan; Shoham, Yuval; Shoham, Gil

2013-05-29

45

Physicochemical characterization of tensio-active produced by Geobacillus stearothermophilus isolated from petroleum-contaminated soil.  

Science.gov (United States)

Biosurfactants are surface-active agents of microbial origin, and have a property of lowering the interfacial tension between two liquids. They act on the interface and are amphiphathic molecules; in with both hydrophilic and hydrophobic portions are present in the same molecule. However, the economics of producing biosurfactant has limited its commercial applications, and the costs can be reduced using cheap substrates or industrial waste. The present study showed the biosurfactant production using corn steep liquor and palm oil as carbon and nitrogen sources for reduction the costs of production. The biosurfactant production by Geobacillus stearothermophilus UCP 986 was carried out using optimized culture medium constituted by palm oil (7.5%) and corn steep liquor (4.5%) using Bioflo fermentor, at temperature of 45°C, during 32 h and agitation of 300 rpm. The biosurfactant showed a reduction of the water surface tension of 72-31 mN/m and interfacial tension of 0.3 mN/m. The biosurfactant was obtained from the net metabolic liquid by acetone precipitation corresponding to the yield of 2.3g/L. The isolate biosurfactant showed a CMC of 2.5% and non-ionic profile. The best emulsification index (E(24)) obtained was 87% using motor oil burned. The biosurfactant solution (2.5%) used in oil spreading test increases the viscosity of engine burning oil of 149.2% and 138.2% to vegetable fat post-frying, respectively. The gas chromatography-mass spectrometer indicated at 29.52 min a molecular weight of 207 Da and eight peaks by FT-IR identified the chemical structure of the biosurfactant produced by G. stearothermophilus. PMID:23010035

Jara, Alícia M A T; Andrade, Rosileide F S; Campos-Takaki, Galba M

2012-06-27

46

Physicochemical characterization of tensio-active produced by Geobacillus stearothermophilus isolated from petroleum-contaminated soil.  

UK PubMed Central (United Kingdom)

Biosurfactants are surface-active agents of microbial origin, and have a property of lowering the interfacial tension between two liquids. They act on the interface and are amphiphathic molecules; in with both hydrophilic and hydrophobic portions are present in the same molecule. However, the economics of producing biosurfactant has limited its commercial applications, and the costs can be reduced using cheap substrates or industrial waste. The present study showed the biosurfactant production using corn steep liquor and palm oil as carbon and nitrogen sources for reduction the costs of production. The biosurfactant production by Geobacillus stearothermophilus UCP 986 was carried out using optimized culture medium constituted by palm oil (7.5%) and corn steep liquor (4.5%) using Bioflo fermentor, at temperature of 45°C, during 32 h and agitation of 300 rpm. The biosurfactant showed a reduction of the water surface tension of 72-31 mN/m and interfacial tension of 0.3 mN/m. The biosurfactant was obtained from the net metabolic liquid by acetone precipitation corresponding to the yield of 2.3g/L. The isolate biosurfactant showed a CMC of 2.5% and non-ionic profile. The best emulsification index (E(24)) obtained was 87% using motor oil burned. The biosurfactant solution (2.5%) used in oil spreading test increases the viscosity of engine burning oil of 149.2% and 138.2% to vegetable fat post-frying, respectively. The gas chromatography-mass spectrometer indicated at 29.52 min a molecular weight of 207 Da and eight peaks by FT-IR identified the chemical structure of the biosurfactant produced by G. stearothermophilus.

Jara AM; Andrade RF; Campos-Takaki GM

2013-01-01

47

Decontamination assessment of Bacillus anthracis, Bacillus subtilis, and Geobacillus stearothermophilus spores on indoor surfaces using a hydrogen peroxide gas generator.  

UK PubMed Central (United Kingdom)

AIMS: To evaluate the decontamination of Bacillus anthracis, Bacillus subtilis, and Geobacillus stearothermophilus spores on indoor surface materials using hydrogen peroxide gas. METHODS AND RESULTS: Bacillus anthracis, B. subtilis, and G. stearothermophilus spores were dried on seven types of indoor surfaces and exposed to > or =1000 ppm hydrogen peroxide gas for 20 min. Hydrogen peroxide exposure significantly decreased viable B. anthracis, B. subtilis, and G. stearothermophilus spores on all test materials except G. stearothermophilus on industrial carpet. Significant differences were observed when comparing the reduction in viable spores of B. anthracis with both surrogates. The effectiveness of gaseous hydrogen peroxide on the growth of biological indicators and spore strips was evaluated in parallel as a qualitative assessment of decontamination. At 1 and 7 days postexposure, decontaminated biological indicators and spore strips exhibited no growth, while the nondecontaminated samples displayed growth. CONCLUSIONS: Significant differences in decontamination efficacy of hydrogen peroxide gas on porous and nonporous surfaces were observed when comparing the mean log reduction in B. anthracis spores with B. subtilis and G. stearothermophilus spores. SIGNIFICANCE AND IMPACT OF THE STUDY: These results provide comparative information for the decontamination of B. anthracis spores with surrogates on indoor surfaces using hydrogen peroxide gas.

Rogers JV; Sabourin CL; Choi YW; Richter WR; Rudnicki DC; Riggs KB; Taylor ML; Chang J

2005-01-01

48

Enhancement of the hydrolysis activity of ?-galactosidase from Geobacillus stearothermophilus by saturation mutagenesis.  

UK PubMed Central (United Kingdom)

Thermostable ?-galactosidase (BgaB) from Geobacillus stearothermophilus is characterized by its thermoactivity in the hydrolysis of lactose to produce lactose-free milk products. However, BgaB has limited activity toward lactose. We established a method for screening evolved mutants with high hydrolysis activity based on prediction of substrate binding sites. Seven amino acid residues were identified as candidates for substrate binding to galactose. To study the hydrolysis activity of these residues, we constructed mutants by site-saturation mutagenesis of these residue sites, and each variant was screened for its hydrolysis activity. The first round of mutagenesis showed that changes in amino acid residues of Arg109, Tyr272, and Glu351 resulted in altered hydrolysis activity, including greater activity toward ortho-nitrophenyl-?-d-galactopyranoside (oNPG). The mutants R109V and R109L displayed changes in the optimum pH from 7.0 to 6.5, and the mutant R109V/L displayed different substrate affinity and catalytic efficiency (k(cat)/K(m)). Mutant R109G showed complete loss of BgaB enzymatic activity, suggesting that Arg109 plays a significant role in maintaining hydrolysis activity. The optimum pH of mutant E351R increased from 7.0 to 7.5 and this mutant showed a prominent increase in catalytic efficiency with oNPG and lactose as substrates.

Dong YN; Liu XM; Chen HQ; Xia Y; Zhang HP; Zhang H; Chen W

2011-03-01

49

Structure-Specificity Relationships of an Intracellular Xylanase from Geobacillus stearothermophilus  

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Geobacillus stearothermophilus T-6 is a thermophilic Gram-positive bacterium that produces two selective family 10 xylanases which both take part in the complete degradation and utilization of the xylan polymer. The two xylanases exhibit significantly different substrate specificities. While the extracellular xylanase (XT6; MW 43.8 kDa) hydrolyzes the long and branched native xylan polymer, the intracellular xylanase (IXT6; MW 38.6 kDa) preferentially hydrolyzes only short xylo-oligosaccharides. In this study, the detailed three-dimensional structure of IXT6 is reported, as determined by X-ray crystallography. It was initially solved by molecular replacement and then refined at 1.45 {angstrom} resolution to a final R factor of 15.0% and an R{sub free} of 19.0%. As expected, the structure forms the classical ({alpha}/{beta}){sub 8} fold, in which the two catalytic residues (Glu134 and Glu241) are located on the inner surface of the central cavity. The structure of IXT6 was compared with the highly homologous extracellular xylanase XT6, revealing a number of structural differences between the active sites of the two enzymes. In particular, structural differences derived from the unique subdomain in the carboxy-terminal region of XT6, which is completely absent in IXT6. These structural modifications may account for the significant differences in the substrate specificities of these otherwise very similar enzymes.

Solomon,V.; Teplitsky, A.; Shulami, S.; Zolotnitsky, G.; Shoham, Y.; Shoham, G.

2007-01-01

50

Efficacy of 3M™ Petrifilm™ aerobic count plates for enumerating Bacillus sporothermodurans and Geobacillus stearothermophilus in UHT milk  

UK PubMed Central (United Kingdom)

The effectiveness of the 3M™ Petrifilm™ aerobic count plate for determining the amount of Bacillus sporothermodurans and Geobacillus stearothermophilus in ultra high temperature milk was determined and its efficacy was compared with the pour-plate agar and one-streak nutrient agar methods. Tubes containing milk were inoculated with spore suspensions, heat shocked and incubated. Aliquots were collected after 0, 24, 48, and 72h of incubation, inoculated using the three different plate methods, and incubated at 55°C. B. sporothermodurans was detected at 9h on the Petrifilm plate, compared with 18h for both the pour plate and one-streak methods. Depending on the strain, G. stearothermophilus was detected between 6 and 12h by the Petrifilm plate method and between 9 and 12h by the other methods. The Petrifilm plates gave reproducible results compared with the traditional methods and are compatible with industrial requirements for milk quality control.

Casillas-Buenrostro RM; Heredia NL; Benesh DL; García S

2012-08-01

51

Crystallization and preliminary crystallographic analysis of Axe2, an acetylxylan esterase from Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Acetylxylan esterases are part of the hemi-cellulolytic system of many microorganisms which utilize plant biomass for growth. Xylans, which are polymeric sugars that constitute a significant part of the plant biomass, are usually substituted with acetyl side groups attached at position 2 or 3 of the xylose backbone units. Acetylxylan esterases hydrolyse the ester linkages of the xylan acetyl groups and thus improve the ability of main-chain hydrolysing enzymes to break down the sugar backbone units. As such, these enzymes play an important part in the hemi-cellulolytic utilization system of many microorganisms that use plant biomass for growth. Interest in the biochemical characterization and structural analysis of these enzymes stems from their numerous potential biotechnological applications. An acetylxylan esterase (Axe2) of this type from Geobacillus stearothermophilus T-6 has recently been cloned, overexpressed, purified, biochemically characterized and crystallized. One of the crystal forms obtained (RB1) belonged to the tetragonal space group I422, with unit-cell parameters a = b = 110.2, c = 213.1 Å. A full diffraction data set was collected to 1.85 Å resolution from flash-cooled crystals of the wild-type enzyme at 100 K using synchrotron radiation. A selenomethionine derivative of Axe2 has also been prepared and crystallized for single-wavelength anomalous diffraction experiments. The crystals of the selenomethionine-derivatized Axe2 appeared to be isomorphous to those of the wild-type enzyme and enabled the measurement of a full 1.85 Å resolution diffraction data set at the selenium absorption edge and a full 1.70 Å resolution data set at a remote wavelength. These data are currently being used for three-dimensional structure determination of the Axe2 protein.

Lansky S; Alalouf O; Solomon V; Alhassid A; Govada L; Chayan NE; Belrhali H; Shoham Y; Shoham G

2013-04-01

52

Crystallization and preliminary crystallographic analysis of Axe2, an acetylxylan esterase from Geobacillus stearothermophilus.  

Science.gov (United States)

Acetylxylan esterases are part of the hemi-cellulolytic system of many microorganisms which utilize plant biomass for growth. Xylans, which are polymeric sugars that constitute a significant part of the plant biomass, are usually substituted with acetyl side groups attached at position 2 or 3 of the xylose backbone units. Acetylxylan esterases hydrolyse the ester linkages of the xylan acetyl groups and thus improve the ability of main-chain hydrolysing enzymes to break down the sugar backbone units. As such, these enzymes play an important part in the hemi-cellulolytic utilization system of many microorganisms that use plant biomass for growth. Interest in the biochemical characterization and structural analysis of these enzymes stems from their numerous potential biotechnological applications. An acetylxylan esterase (Axe2) of this type from Geobacillus stearothermophilus T-6 has recently been cloned, overexpressed, purified, biochemically characterized and crystallized. One of the crystal forms obtained (RB1) belonged to the tetragonal space group I422, with unit-cell parameters a = b = 110.2, c = 213.1 Å. A full diffraction data set was collected to 1.85 Å resolution from flash-cooled crystals of the wild-type enzyme at 100 K using synchrotron radiation. A selenomethionine derivative of Axe2 has also been prepared and crystallized for single-wavelength anomalous diffraction experiments. The crystals of the selenomethionine-derivatized Axe2 appeared to be isomorphous to those of the wild-type enzyme and enabled the measurement of a full 1.85 Å resolution diffraction data set at the selenium absorption edge and a full 1.70 Å resolution data set at a remote wavelength. These data are currently being used for three-dimensional structure determination of the Axe2 protein. PMID:23545652

Lansky, Shifra; Alalouf, Onit; Solomon, Vered; Alhassid, Anat; Govada, Lata; Chayan, Naomi E; Belrhali, Hassan; Shoham, Yuval; Shoham, Gil

2013-03-28

53

Plasma sterilization of Geobacillus Stearothermophilus by O{mathsf2}:N{mathsf2} RF inductively coupled plasma  

Science.gov (United States)

The aim of this work is to identify the main process responsible for sterilization of Geobacillus Stearothermophilus spores in O{2}:N{2} RF inductively coupled plasma. In order to meet this objective the sterilization efficiencies of discharges in mixtures differing in the initial O{2}/N{2} ratios are compared with plasma properties and with scanning electron microscopy images of treated spores. According to the obtained results it can be concluded that under our experimental conditions the time needed to reach complete sterilization is more related to O atom density than UV radiation intensity, i.e. complete sterilization is not related only to DNA damage as in UV sterilization but more likely to the etching of the spore.

Kylián, O.; Sasaki, T.; Rossi, F.

2006-05-01

54

Improving the thermostability of Geobacillus stearothermophilus xylanase XT6 by directed evolution and site-directed mutagenesis.  

UK PubMed Central (United Kingdom)

Protein engineering of the thermostable xylanase XT6 from Geobacillus stearothermophilus was performed to obtain enzymes with improved thermal tolerance. Mutants producing such enzymes were obtained after several rounds of directed evolution using error-prone PCR and sequence family shuffling, in combination with a consensus-based semi-rational approach. The most thermostable mutant enzyme contained 13 amino acid substitutions and its half-life of inactivation was 52-fold of that of the wild-type. Its reaction temperature for maximum activity increased from 77 degrees C to 87 degrees C, and catalytic efficiency (k(cat)/K(m)) increased by 90%. The mutant is of potential interest for industrial applications.

Zhang ZG; Yi ZL; Pei XQ; Wu ZL

2010-12-01

55

Tryptophan oxidative metabolism catalyzed by geobacillus stearothermophilus: a thermophile isolated from kuwait soil contaminated with petroleum hydrocarbons.  

UK PubMed Central (United Kingdom)

Tryptophan metabolism has been extensively studied in humans as well as in soil. Its metabolism takes place mainly through kynurenine pathway yielding hydroxylated, deaminated and many other products of physiological significance. However, tryptophan metabolism has not been studied in an isolated thermophilic bacterium. Geobacillus stearothermophilus is a local thermophile isolated from Kuwait desert soil contaminated with petroleum hydrocarbons. The bacterium grows well at 65 °C in 0.05 M phosphate buffer (pH 7), when supplied with organic compounds as a carbon source and has a good potential for transformation of steroids and related molecules. In the present study, we used tryptophan ethyl ester as a carbon source for the bacterium to study the catabolism of the amino acid at pH 5 and pH 7. In this endeavor, we have resolved twenty one transformation products of tryptophan by GC/LC and have identified them through their mass spectral fragmentation.

Al-Hassan JM; Al-Awadi S; Oommen S; Alkhamis A; Afzal M

2011-01-01

56

Analysis of reduction of Geobacillus stearothermophilus spores treated with high hydrostatic pressure and mild heat in milk buffer.  

UK PubMed Central (United Kingdom)

Our unpublished experimental results of fractional factorial experiments showed that the significant external factors affecting high pressure processing (HPP) inactivation were pressure, temperature and pressure holding time. Based on these results, response surface methodology (RSM) was employed in the present work and a quadratic equation for HPP inactivation was built. By analyzing the response surface plots and their corresponding contour plots as well as solving the quadratic equation, the experimental values were shown to be significantly in good agreement with predicted values since the adjusted determination coefficient (R(Adj)(2)) was 0.9747. The optimum process parameters for six log-cycles reduction of Geobacillus stearothermophilus spores were obtained as: temperature, 86 degrees C; pressure, 625.0 MPa and pressure holding time, 14.0 min. The adequacy of the model equation for predicting the optimum response values was verified effectively by the validation data.

Gao YL; Ju XR; Jiang HH

2006-09-01

57

Analysis of reduction of Geobacillus stearothermophilus spores treated with high hydrostatic pressure and mild heat in milk buffer.  

Science.gov (United States)

Our unpublished experimental results of fractional factorial experiments showed that the significant external factors affecting high pressure processing (HPP) inactivation were pressure, temperature and pressure holding time. Based on these results, response surface methodology (RSM) was employed in the present work and a quadratic equation for HPP inactivation was built. By analyzing the response surface plots and their corresponding contour plots as well as solving the quadratic equation, the experimental values were shown to be significantly in good agreement with predicted values since the adjusted determination coefficient (R(Adj)(2)) was 0.9747. The optimum process parameters for six log-cycles reduction of Geobacillus stearothermophilus spores were obtained as: temperature, 86 degrees C; pressure, 625.0 MPa and pressure holding time, 14.0 min. The adequacy of the model equation for predicting the optimum response values was verified effectively by the validation data. PMID:16621090

Gao, Yu-Long; Ju, Xing-Rong; Jiang, Han-Hu

2006-04-18

58

Cloning of araA Gene Encoding L-Arabinose Isomerase from Marine Geobacillus stearothermophilus Isolated from Tanjung Api, Poso, Indonesia  

Directory of Open Access Journals (Sweden)

Full Text Available L-arabinose isomerase is an enzyme converting D-galactose to D-tagatose. D-tagatose is a potential sweetener-sucrose substitute which has low calorie. This research was to clone and sequence araA gene from marine bacterial strain Geobacillus stearothermophilus isolated from Tanjung Api Poso Indonesia. The amplified araA gene consisted of 1494 bp nucleotides encoding 497 amino acids. DNA alignment analysis showed that the gene had high homology with that of G. stearothermophilus T6. The enzyme had optimum activity at high temperature and alkalin condition.

DEWI FITRIANI; BUDI SAKSONO

2010-01-01

59

A double mutant of highly purified Geobacillus stearothermophilus lactate dehydrogenase recognises l-mandelic acid as a substrate.  

Science.gov (United States)

Lactate dehydrogenase from the thermophilic organism Geobacillus stearothermophilus (formerly Bacillus stearothermophilus) (bsLDH) has a crucial role in producing chirally pure hydroxyl compounds. ?-Hydroxy acids are used in many industrial situations, ranging from pharmaceutical to cosmetic dermatology products. One drawback of this enzyme is its limited substrate specificity. For instance, l-lactate dehydrogenase exhibits no detectable activity towards the large side chain of 2-hydroxy acid l-mandelic acid, an ?-hydroxy acid with anti-bacterial activity. Despite many attempts to engineer bsLDH to accept ?-hydroxy acid substrates, there have been no attempts to introduce the industrially important l-mandelic acid to bsLDH. Herein, we describe attempts to change the reactivity of bsLDH towards l-mandelic acid. Using the Insight II molecular modelling programme (except 'program' in computers) and protein engineering techniques, we have successfully introduced substantial mandelate dehydrogenase activity to the enzyme. Energy minimisation modelling studies suggested that two mutations, T246G and I240A, would allow the enzyme to utilise l-mandelic acid as a substrate. Genes encoding for the wild-type and mutant enzymes were constructed, and the resulting bsLDH proteins were overexpressed in Escherichia coli and purified using the TAGZyme system. Enzyme assays showed that insertion of this double mutation into highly purified bsLDH switched the substrate specificity from lactate to l-mandelic acid. PMID:23608509

Binay, Bar??; Sessions, Richard B; Karagüler, Nevin Gül

2013-02-27

60

A double mutant of highly purified Geobacillus stearothermophilus lactate dehydrogenase recognises l-mandelic acid as a substrate.  

UK PubMed Central (United Kingdom)

Lactate dehydrogenase from the thermophilic organism Geobacillus stearothermophilus (formerly Bacillus stearothermophilus) (bsLDH) has a crucial role in producing chirally pure hydroxyl compounds. ?-Hydroxy acids are used in many industrial situations, ranging from pharmaceutical to cosmetic dermatology products. One drawback of this enzyme is its limited substrate specificity. For instance, l-lactate dehydrogenase exhibits no detectable activity towards the large side chain of 2-hydroxy acid l-mandelic acid, an ?-hydroxy acid with anti-bacterial activity. Despite many attempts to engineer bsLDH to accept ?-hydroxy acid substrates, there have been no attempts to introduce the industrially important l-mandelic acid to bsLDH. Herein, we describe attempts to change the reactivity of bsLDH towards l-mandelic acid. Using the Insight II molecular modelling programme (except 'program' in computers) and protein engineering techniques, we have successfully introduced substantial mandelate dehydrogenase activity to the enzyme. Energy minimisation modelling studies suggested that two mutations, T246G and I240A, would allow the enzyme to utilise l-mandelic acid as a substrate. Genes encoding for the wild-type and mutant enzymes were constructed, and the resulting bsLDH proteins were overexpressed in Escherichia coli and purified using the TAGZyme system. Enzyme assays showed that insertion of this double mutation into highly purified bsLDH switched the substrate specificity from lactate to l-mandelic acid.

Binay B; Sessions RB; Karagüler NG

2013-05-01

 
 
 
 
61

Meticillin-resistant Staphylococcus aureus is more resistant to vaporized hydrogen peroxide than commercial Geobacillus stearothermophilus biological indicators.  

UK PubMed Central (United Kingdom)

BACKGROUND: Vaporized hydrogen peroxide (VHP) is increasingly used in the decontamination of hospital isolation rooms. Commercially available bioindicators, most frequently Geobacillus stearothermophilus spores, are used to assess the efficacy of the decontamination phase. Staphylococcus aureus, including meticillin-resistant S. aureus (MRSA), produce catalase, which breaks down VHP, therefore potentially making it resistant to the decontamination phase. AIM: This investigation was designed to assess the resistance of meticillin-resistant S. aureus to VHP in comparison with commercially available biological indicators loaded with spores. METHODS: Stainless steel indicators were prepared with the same loading of MRSA (NCTC 13142) as commercially available indicators of G. stearothermophilus (ATCC 7953) (?3.1×10(6) spores) and both indicators were exposed to a vapour hydrogen peroxide cycle (750 ppm). At set time-points during the exposure period, indicators containing both organisms were removed for processing and enumeration to compare survivability. FINDINGS: During the exposure period the recovery of MRSA from the coupons was between 1.5 and 3.5 log(10) higher than the recovery of G. stearothermophilus spores (P<0.05). This greater resistance may be due to the production of catalase which could break down the hydrogen peroxide, resulting in a reduction of the effectiveness of VHP. CONCLUSION: These findings highlight that the reduction achieved with a commercially available biological indicator cannot always be extrapolated to other micro-organisms. It must be recognized that although gaseous decontamination is the final step of the decontamination process, pre-cleaning of surfaces must be carried out to reduce the microbial loading being exposed.

Pottage T; Macken S; Walker JT; Bennett AM

2012-01-01

62

A meta-analysis accounting for sources of variability to estimate heat resistance reference parameters of bacteria using hierarchical Bayesian modeling: Estimation of D at 121.1 °C and pH 7, zT and zpH of Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Predicting microbial survival requires reference parameters for each micro-organism of concern. When data are abundant and publicly available, a meta-analysis is a useful approach for assessment of these parameters, which can be performed with hierarchical Bayesian modeling. Geobacillus stearothermophilus is a major agent of microbial spoilage of canned foods and is therefore a persistent problem in the food industry. The thermal inactivation parameters of G. stearothermophilus (D(ref), i.e.the decimal reduction time D at the reference temperature 121.1°C and pH 7.0, z(T) and z(pH)) were estimated from a large set of 430 D values mainly collected from scientific literature. Between-study variability hypotheses on the inactivation parameters D(ref), z(T) and z(pH) were explored, using three different hierarchical Bayesian models. Parameter estimations were made using Bayesian inference and the models were compared with a graphical and a Bayesian criterion. Results show the necessity to account for random effects associated with between-study variability. Assuming variability on D(ref), z(T) and z(pH), the resulting distributions for D(ref), z(T) and z(pH) led to a mean of 3.3 min for D(ref) (95% Credible Interval CI=[0.8; 9.6]), to a mean of 9.1°C for z(T) (CI=[5.4; 13.1]) and to a mean of 4.3 pH units for z(pH) (CI=[2.9; 6.3]), in the range pH 3 to pH 7.5. Results are also given separating variability and uncertainty in these distributions, as well as adjusted parametric distributions to facilitate further use of these results in aqueous canned foods such as canned vegetables.

Rigaux C; Denis JB; Albert I; Carlin F

2013-02-01

63

Identifying assembly-inhibiting and assembly-tolerant sites in the SbsB S-layer protein from Geobacillus stearothermophilus.  

Science.gov (United States)

Surface layer (S-layer) proteins self-assemble into two-dimensional crystalline lattices that cover the cell wall of all archaea and many bacteria. We have generated assembly-negative protein variants of high solubility that will facilitate high-resolution structure determination. Assembly-negative versions of the S-layer protein SbsB from Geobacillus stearothermophilus PV72/p2 were obtained using an insertion mutagenesis screen. The haemagglutinin epitope tag was inserted at 23 amino acid positions known to be located on the monomer protein surface from a previous cysteine accessibility screen. Limited proteolysis, circular dichroism, and fluorescence were used to probe whether the epitope insertion affected the secondary and tertiary structures of the monomer, while electron microscopy and size-exclusion chromatography were employed to examine proteins' ability to self-assemble. The screen not only identified assembly-compromised mutants with native fold but also yielded correctly folded, self-assembling mutants suitable for displaying epitopes for biomedical and biophysical applications, as well as cryo-electron microscopy imaging. Our study marks an important step in the analysis of the S-layer structure. In addition, the approach of concerted insertion and cysteine mutagenesis can likely be applied for other supramolecular assemblies. PMID:19836402

Kinns, Helen; Badelt-Lichtblau, Helga; Egelseer, Eva Maria; Sleytr, Uwe B; Howorka, Stefan

2009-10-15

64

Identifying assembly-inhibiting and assembly-tolerant sites in the SbsB S-layer protein from Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Surface layer (S-layer) proteins self-assemble into two-dimensional crystalline lattices that cover the cell wall of all archaea and many bacteria. We have generated assembly-negative protein variants of high solubility that will facilitate high-resolution structure determination. Assembly-negative versions of the S-layer protein SbsB from Geobacillus stearothermophilus PV72/p2 were obtained using an insertion mutagenesis screen. The haemagglutinin epitope tag was inserted at 23 amino acid positions known to be located on the monomer protein surface from a previous cysteine accessibility screen. Limited proteolysis, circular dichroism, and fluorescence were used to probe whether the epitope insertion affected the secondary and tertiary structures of the monomer, while electron microscopy and size-exclusion chromatography were employed to examine proteins' ability to self-assemble. The screen not only identified assembly-compromised mutants with native fold but also yielded correctly folded, self-assembling mutants suitable for displaying epitopes for biomedical and biophysical applications, as well as cryo-electron microscopy imaging. Our study marks an important step in the analysis of the S-layer structure. In addition, the approach of concerted insertion and cysteine mutagenesis can likely be applied for other supramolecular assemblies.

Kinns H; Badelt-Lichtblau H; Egelseer EM; Sleytr UB; Howorka S

2010-01-01

65

Taxonomic study of aerobic thermophilic bacilli: descriptions of Geobacillus subterraneus gen. nov., sp. nov. and Geobacillus uzenensis sp. nov. from petroleum reservoirs and transfer of Bacillus stearothermophilus, Bacillus thermocatenulatus, Bacillus thermoleovorans, Bacillus kaustophilus, Bacillus thermodenitrificans to Geobacillus as the new combinations G. stearothermophilus, G. th.  

UK PubMed Central (United Kingdom)

Five hydrocarbon-oxidizing strains were isolated from formation waters of oilfields in Russia, Kazakhstan and China. These strains were moderately thermophilic, neutrophilic, motile, spore-forming rods, aerobic or facultatively anaerobic. The G+C content of their DNA ranged from 49.7 to 52.3 mol%. The major isoprenoid quinone was menaquinone-7; cellular fatty acid profiles consisted of significant amounts of iso-15:0, iso-16:0 and iso-17:0 fatty acids (61.7-86.8% of the total). Based on data from 16S rDNA analysis and DNA-DNA hybridization, the subsurface isolates could be divided into two groups, one of which consisted of strains UT and X and the other of which consisted of strains K, Sam and 34T. The new strains exhibited a close phylogenetic relationship to thermophilic bacilli of 'Group 5' of Ash et al. [Ash, C., Farrow, J. A. E., Wallbanks, S. & Collins, M. D. (1991). Lett Appl Microbiol 13, 202-206] and a set of corresponding signature positions of 16S rRNA. Comparative analysis of the 16S rDNA sequences and fatty acid compositions of the novel isolates and established species of thermophilic bacilli indicated that the subsurface strains represent two new species within a new genus, for which the names Geobacillus subterraneus gen. nov., sp. nov., and Geobacillus uzenensis sp. nov. are proposed. It is also proposed that Bacillus stearothermophilus, Bacillus thermoleovorans, Bacillus thermocatenulatus, Bacillus kaustophilus, Bacillus thermoglucosidasius and Bacillus thermodenitrificans be transferred to this new genus, with Geobacillus stearothermophilus (formerly Bacillus stearothermophilus) as the type species.

Nazina TN; Tourova TP; Poltaraus AB; Novikova EV; Grigoryan AA; Ivanova AE; Lysenko AM; Petrunyaka VV; Osipov GA; Belyaev SS; Ivanov MV

2001-03-01

66

Taxonomic study of aerobic thermophilic bacilli: descriptions of Geobacillus subterraneus gen. nov., sp. nov. and Geobacillus uzenensis sp. nov. from petroleum reservoirs and transfer of Bacillus stearothermophilus, Bacillus thermocatenulatus, Bacillus thermoleovorans, Bacillus kaustophilus, Bacillus thermodenitrificans to Geobacillus as the new combinations G. stearothermophilus, G. th.  

Science.gov (United States)

Five hydrocarbon-oxidizing strains were isolated from formation waters of oilfields in Russia, Kazakhstan and China. These strains were moderately thermophilic, neutrophilic, motile, spore-forming rods, aerobic or facultatively anaerobic. The G+C content of their DNA ranged from 49.7 to 52.3 mol%. The major isoprenoid quinone was menaquinone-7; cellular fatty acid profiles consisted of significant amounts of iso-15:0, iso-16:0 and iso-17:0 fatty acids (61.7-86.8% of the total). Based on data from 16S rDNA analysis and DNA-DNA hybridization, the subsurface isolates could be divided into two groups, one of which consisted of strains UT and X and the other of which consisted of strains K, Sam and 34T. The new strains exhibited a close phylogenetic relationship to thermophilic bacilli of 'Group 5' of Ash et al. [Ash, C., Farrow, J. A. E., Wallbanks, S. & Collins, M. D. (1991). Lett Appl Microbiol 13, 202-206] and a set of corresponding signature positions of 16S rRNA. Comparative analysis of the 16S rDNA sequences and fatty acid compositions of the novel isolates and established species of thermophilic bacilli indicated that the subsurface strains represent two new species within a new genus, for which the names Geobacillus subterraneus gen. nov., sp. nov., and Geobacillus uzenensis sp. nov. are proposed. It is also proposed that Bacillus stearothermophilus, Bacillus thermoleovorans, Bacillus thermocatenulatus, Bacillus kaustophilus, Bacillus thermoglucosidasius and Bacillus thermodenitrificans be transferred to this new genus, with Geobacillus stearothermophilus (formerly Bacillus stearothermophilus) as the type species. PMID:11321089

Nazina, T N; Tourova, T P; Poltaraus, A B; Novikova, E V; Grigoryan, A A; Ivanova, A E; Lysenko, A M; Petrunyaka, V V; Osipov, G A; Belyaev, S S; Ivanov, M V

2001-03-01

67

Proton transfer in the quinol-dependent nitric oxide reductase from Geobacillus stearothermophilus during reduction of oxygen.  

UK PubMed Central (United Kingdom)

Bacterial nitric oxide reductases (NOR) are integral membrane proteins that catalyse the reduction of nitric oxide to nitrous oxide, often as a step in the process of denitrification. Most functional data has been obtained with NORs that receive their electrons from a soluble cytochrome c in the periplasm and are hence termed cNOR. Very recently, the structure of a different type of NOR, the quinol-dependent (q)-NOR from the thermophilic bacterium Geobacillus stearothermophilus was solved to atomic resolution [Y. Matsumoto, T. Tosha, A.V. Pisliakov, T. Hino, H. Sugimoto, S. Nagano, Y. Sugita and Y. Shiro, Nat. Struct. Mol. Biol. 19 (2012) 238-246]. In this study, we have investigated the reaction between this qNOR and oxygen. Our results show that, like some cNORs, the G. stearothermophilus qNOR is capable of O(2) reduction with a turnover of ~3electronss(-1) at 40°C. Furthermore, using the so-called flow-flash technique, we show that the fully reduced (with three available electrons) qNOR reacts with oxygen in a reaction with a time constant of 1.8ms that oxidises the low-spin heme b. This reaction is coupled to proton uptake from solution and presumably forms a ferryl intermediate at the active site. The pH dependence of the reaction is markedly different from a corresponding reaction in cNOR from Paracoccus denitrificans, indicating that possibly the proton uptake mechanism and/or pathway differs between qNOR and cNOR. This study furthermore forms the basis for investigation of the proton transfer pathway in qNOR using both variants with putative proton transfer elements modified and measurements of the vectorial nature of the proton transfer. This article is part of a Special Issue entitled: 17th European Bioenergetics Conference (EBEC 2012).

Salomonsson L; Reimann J; Tosha T; Krause N; Gonska N; Shiro Y; Adelroth P

2012-10-01

68

Proton transfer in the quinol-dependent nitric oxide reductase from Geobacillus stearothermophilus during reduction of oxygen.  

Science.gov (United States)

Bacterial nitric oxide reductases (NOR) are integral membrane proteins that catalyse the reduction of nitric oxide to nitrous oxide, often as a step in the process of denitrification. Most functional data has been obtained with NORs that receive their electrons from a soluble cytochrome c in the periplasm and are hence termed cNOR. Very recently, the structure of a different type of NOR, the quinol-dependent (q)-NOR from the thermophilic bacterium Geobacillus stearothermophilus was solved to atomic resolution [Y. Matsumoto, T. Tosha, A.V. Pisliakov, T. Hino, H. Sugimoto, S. Nagano, Y. Sugita and Y. Shiro, Nat. Struct. Mol. Biol. 19 (2012) 238-246]. In this study, we have investigated the reaction between this qNOR and oxygen. Our results show that, like some cNORs, the G. stearothermophilus qNOR is capable of O(2) reduction with a turnover of ~3electronss(-1) at 40°C. Furthermore, using the so-called flow-flash technique, we show that the fully reduced (with three available electrons) qNOR reacts with oxygen in a reaction with a time constant of 1.8ms that oxidises the low-spin heme b. This reaction is coupled to proton uptake from solution and presumably forms a ferryl intermediate at the active site. The pH dependence of the reaction is markedly different from a corresponding reaction in cNOR from Paracoccus denitrificans, indicating that possibly the proton uptake mechanism and/or pathway differs between qNOR and cNOR. This study furthermore forms the basis for investigation of the proton transfer pathway in qNOR using both variants with putative proton transfer elements modified and measurements of the vectorial nature of the proton transfer. This article is part of a Special Issue entitled: 17th European Bioenergetics Conference (EBEC 2012). PMID:22538294

Salomonsson, Lina; Reimann, Joachim; Tosha, Takehiko; Krause, Nils; Gonska, Nathalie; Shiro, Yoshitsugu; Adelroth, Pia

2012-04-17

69

The structure of DinB from Geobacillus stearothermophilus: a representative of a unique four-helix-bundle superfamily.  

UK PubMed Central (United Kingdom)

The crystal structure of the dinB gene product from Geobacillus stearothermophilus (GsDinB) is reported at 2.5 A resolution. The dinB gene is one of the DNA-damage-induced genes and the corresponding protein, DinB, is the founding member of a Pfam family with no known function. The protein contains a four-helix up-down-down-up bundle that has previously been described in the literature in three disparate proteins: the enzyme MDMPI (mycothiol-dependent maleylpyruvate isomerase), YfiT and TTHA0303, a member of a small DUF (domain of unknown function). However, a search of the DALI structural database revealed similarities to a further 11 new unpublished structures contributed by structural genomics centers. The sequences of these proteins are quite divergent and represent several Pfam families, yet their structures are quite similar and most (but not all) seem to have the ability to coordinate a metal ion using a conserved histidine-triad motif. The structural similarities of these diverse proteins suggest that a new Pfam clan encompassing the families that share this fold should be created. The proteins that share this fold exhibit four different quaternary structures: monomeric and three different dimeric forms.

Cooper DR; Grelewska K; Kim CY; Joachimiak A; Derewenda ZS

2010-03-01

70

How to Switch Off a Histidine Kinase: Crystal Structure of Geobacillus Stearothermophilus KinB with the Inhibitor Sda  

Energy Technology Data Exchange (ETDEWEB)

Entry to sporulation in bacilli is governed by a histidine kinase phosphorelay, a variation of the predominant signal transduction mechanism in prokaryotes. Sda directly inhibits sporulation histidine kinases in response to DNA damage and replication defects. We determined a 2.0-Angstroms-resolution X-ray crystal structure of the intact cytoplasmic catalytic core [comprising the dimerization and histidine phosphotransfer domain (DHp domain), connected to the ATP binding catalytic domain] of the Geobacillus stearothermophilus sporulation kinase KinB complexed with Sda. Structural and biochemical analyses reveal that Sda binds to the base of the DHp domain and prevents molecular transactions with the DHp domain to which it is bound by acting as a simple molecular barricade. Sda acts to sterically block communication between the catalytic domain and the DHp domain, which is required for autophosphorylation, as well as to sterically block communication between the response regulator Spo0F and the DHp domain, which is required for phosphotransfer and phosphatase activities.

Bick, M.; Lamour, V; Rajashankar, K; Gordiyenko, Y; Robinson, C; Darst, S

2009-01-01

71

Geobacillus stearothermophilus LV cadA gene mediates resistance to cadmium, lead and zinc in zntA mutants of Salmonella entérica serovar Typhimurium  

Scientific Electronic Library Online (English)

Full Text Available Abstract in english Salmonella entérica serovar Typhimurium cells expressing the cadA gene of Geobacillus stearothermophilus LV exhibit a hypersensitive phenotype to cadmium chloride. Deletion of the ORF STM3576 from the Salmonella genome resulted in cadmium, lead and zinc sensitivity, confirming that this ORF is a homologue of the zntA gene. The observed sensitivity was reverted upon expression of the G. stearothermophilus LV cadA gene. These results indicate that the cadA gene product is (more) involved in Cd, Pb and Zn resistance as a classical P-type ATPase and strongly suggest that the observed hypersensitive phenotype to these metals can be related to the function of the host ·zntA gene product

PÉREZ, JOSÉ M; PRADEÑAS, GONZALO A; NAVARRO, CLAUDIO A; HENRÍQUEZ, DANIEL R; PICHUANTES, SERGIO E; VÁSQUEZ, CLAUDIO C

2006-01-01

72

Geobacillus stearothermophilus LV cadA gene mediates resistance to cadmium, lead and zinc in zntA mutants of Salmonella entérica serovar Typhimurium  

Directory of Open Access Journals (Sweden)

Full Text Available Salmonella entérica serovar Typhimurium cells expressing the cadA gene of Geobacillus stearothermophilus LV exhibit a hypersensitive phenotype to cadmium chloride. Deletion of the ORF STM3576 from the Salmonella genome resulted in cadmium, lead and zinc sensitivity, confirming that this ORF is a homologue of the zntA gene. The observed sensitivity was reverted upon expression of the G. stearothermophilus LV cadA gene. These results indicate that the cadA gene product is involved in Cd, Pb and Zn resistance as a classical P-type ATPase and strongly suggest that the observed hypersensitive phenotype to these metals can be related to the function of the host ·zntA gene product

JOSÉ M PÉREZ; GONZALO A PRADEÑAS; CLAUDIO A NAVARRO; DANIEL R HENRÍQUEZ; SERGIO E PICHUANTES; CLAUDIO C VÁSQUEZ

2006-01-01

73

Crystal structure of quinol-dependent nitric oxide reductase from Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

The structure of quinol-dependent nitric oxide reductase (qNOR) from G. stearothermophilus, which catalyzes the reduction of NO to produce the major ozone-depleting gas N(2)O, has been characterized at 2.5 Å resolution. The overall fold of qNOR is similar to that of cytochrome c-dependent NOR (cNOR), and some structural features that are characteristic of cNOR, such as the calcium binding site and hydrophilic cytochrome c domain, are observed in qNOR, even though it harbors no heme c. In contrast to cNOR, structure-based mutagenesis and molecular dynamics simulation studies of qNOR suggest that a water channel from the cytoplasm can serve as a proton transfer pathway for the catalytic reaction. Further structural comparison of qNOR with cNOR and aerobic and microaerobic respiratory oxidases elucidates their evolutionary relationship and possible functional conversions.

Matsumoto Y; Tosha T; Pisliakov AV; Hino T; Sugimoto H; Nagano S; Sugita Y; Shiro Y

2012-02-01

74

GH52 xylosidase from Geobacillus stearothermophilus: characterization and introduction of xylanase activity by site-directed mutagenesis of Tyr509.  

UK PubMed Central (United Kingdom)

A xylosidase gene, gsxyn, was cloned from the deep-sea thermophilic Geobacillus stearothermophilus, which consisted of 2,118 bp and encoded a protein of 705 amino acids with a calculated molecular mass of 79.8 kDa. The GSxyn of glycoside hydrolase family 52 (GH52) displayed its maximum activity at 70 °C and pH 5.5. The K m and k cat values of GSxyn for ?NPX were 0.48 mM and 36.64 s(-1), respectively. Interestingly, a new exo-xylanase activity was introduced into GSxyn by mutating the tyrosine509 into glutamic acid, whereas the resultant enzyme variant, Y509E, retained the xylosidase activity. The optimum xylanase activity of theY509E mutant displayed at pH 6.5 and 50 °C, and retained approximately 45 % of its maximal activity at 55 °C, pH 6.5 for 60 min. The K m and k cat values of the xylanase activity of Y509E mutant for beechwood xylan were 5.10 mg/ml and 22.53 s(-1), respectively. The optimum xylosidase activity of theY509E mutant displayed at pH 5.5 and 60 °C. The K m and k cat values of the xylosidase activity of Y509E mutant for ?NPX were 0.51 mM and 22.53 s(-1), respectively. This report demonstrated that GH52 xylosidase has provided a platform for generating bifunctional enzymes for industrially significant and complex substrates, such as plant cell wall.

Huang Z; Liu X; Zhang S; Liu Z

2013-10-01

75

A new family of carbohydrate esterases is represented by a GDSL hydrolase/acetylxylan esterase from Geobacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Acetylxylan esterases hydrolyze the ester linkages of acetyl groups at positions 2 and/or 3 of the xylose moieties in xylan and play an important role in enhancing the accessibility of xylanases to the xylan backbone. The hemicellulolytic system of the thermophilic bacterium Geobacillus stearothermophilus T-6 comprises a putative acetylxylan esterase gene, axe2. The gene product belongs to the GDSL hydrolase family and does not share sequence homology with any of the carbohydrate esterases in the CAZy Database. The axe2 gene is induced by xylose, and the purified gene product completely deacetylates xylobiose peracetate (fully acetylated) and hydrolyzes the synthetic substrates 2-naphthyl acetate, 4-nitrophenyl acetate, 4-methylumbelliferyl acetate, and phenyl acetate. The pH profiles for k(cat) and k(cat)/K(m) suggest the existence of two ionizable groups affecting the binding of the substrate to the enzyme. Using NMR spectroscopy, the regioselectivity of Axe2 was directly determined with the aid of one-dimensional selective total correlation spectroscopy. Methyl 2,3,4-tri-O-acetyl-?-d-xylopyranoside was rapidly deacetylated at position 2 or at positions 3 and 4 to give either diacetyl or monoacetyl intermediates, respectively; methyl 2,3,4,6-tetra-O-acetyl-?-d-glucopyranoside was initially deacetylated at position 6. In both cases, the complete hydrolysis of the intermediates occurred at a much slower rate, suggesting that the preferred substrate is the peracetate sugar form. Site-directed mutagenesis of Ser-15, His-194, and Asp-191 resulted in complete inactivation of the enzyme, consistent with their role as the catalytic triad. Overall, our results show that Axe2 is a serine acetylxylan esterase representing a new carbohydrate esterase family.

Alalouf O; Balazs Y; Volkinshtein M; Grimpel Y; Shoham G; Shoham Y

2011-12-01

76

High-Affinity Interaction between the S-Layer Protein SbsC and the Secondary Cell Wall Polymer of Geobacillus stearothermophilus ATCC 12980 Determined by Surface Plasmon Resonance Technology? †  

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Surface plasmon resonance studies using C-terminal truncation forms of the S-layer protein SbsC (recombinant SbsC consisting of amino acids 31 to 270 [rSbsC31-270] and rSbsC31-443) and the secondary cell wall polymer (SCWP) isolated from Geobacillus stearothermophilus ATCC 12980 confirmed the exclus...

Ferner-Ortner, Judith; Mader, Christoph; Ilk, Nicola; Sleytr, Uwe B.; Egelseer, Eva M.

77

Strain Improvement of Bacillus coagulans and Geobacillus stearothermophilus for Enhanced Thermostable Cellulase Production and the Effect of Different Metal Ions on Cellulase Activity  

Directory of Open Access Journals (Sweden)

Full Text Available The current study was focused on the strain improvement of Bacillus coagulans and Geobacillus stearothermophilus for thermostable cellulase production with higher enzyme activity. For strain improvement UV radiations, NTG and Sodium azide were used as mutagenic agents.NTG was found to be best mutagenic agent among all in term of highest cellulase activity. Mutant strain C11 exhibited the highest cellulase specific activity at 45 U/mg followed by C15 (39 U/mg) in case of B.coagulans while Mutant strain S18 exhibited thehighest cellulase specific activity at 69 U/mg followed by S12 (62 U/mg) in case of G. stearothermophilus. Specific activity of cellulase was 92 U/mg in case of B.coagulans C11 and 118 U/mg in case of G. stearothermophilus S18. Ag+, Mg+, Se2+,Ca2+,Co2+,Mn2+,K+, Zn2+ ,Fe3+, Hg2+ and Cu2+ showed positive change in specific activity while Na+, Ni2+ negative change in specific activity of cellulase with respect to specific activity of cellulase in absence of any additive in case of B.coagulans C11 and Ag+, Mg+, Se2+,Co2+,Mn2+ andHg2+ showed positive change in specific activity, Na+, K+ showed no change in specific activity while Ca2+, Zn2+, Ni2+, Fe3+ and Cu2+ showed negative change in specific activity of cellulase with respect to specific activity of cellulase in absence of any additive in case of G. stearothermophilus S18.

Vikas Sharma; Prakash Kumar Singh

2012-01-01

78

Structural basis for thermostability revealed through the identification and characterization of a highly thermostable phosphotriesterase-like lactonase from Geobacillus stearothermophilus  

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A new enzyme homologous to phosphotriesterase was identified from the bacterium Geobacillus stearothermophilus (GsP). This enzyme belongs to the amidohydrolase family and possesses the ability to hydrolyze both lactone and organophosphate (OP) compounds, making it a phosphotriesterase-like lactonase (PLL). GsP possesses higher OP-degrading activity than recently characterized PLLs, and it is extremely thermostable. GsP is active up to 100 C with an energy of activation of 8.0 kcal/mol towards ethyl paraoxon, and it can withstand an incubation temperature of 60 C for two days. In an attempt to understand the thermostability of PLLs, the X-ray structure of GsP was determined and compared to those of existing PLLs. Based upon a comparative analysis, a new thermal advantage score and plot was developed and reveals that a number of different factors contribute to the thermostability of PLLs.

Hawwa, Renda; Aikens, John; Turner, Robert J.; Santarsiero, Bernard D.; Mescar, Andrew D.; (Lybradyn Inc.); (UIC)

2009-08-31

79

Study of the influence of sporulation conditions on heat resistance of Geobacillus stearothermophilus used in the development of biological indicators for steam sterilization.  

UK PubMed Central (United Kingdom)

Biological indicators are important tools in infection control via sterilization process monitoring. The use of a standardized spore crop with a well-defined heat resistance will guarantee the quality of a biological indicator. Ambient factors during sporulation can affect spore characteristics and properties, including heat resistance. The aim of this study is to evaluate the main sporulation factors responsible for heat resistance in Geobacillus stearothermophilus, a useful biological indicator for steam sterilization. A sequence of a three-step optimization of variables (initial pH, nutrient concentration, tryptone, peptone, beef extract, yeast extract, manganese sulfate, magnesium sulfate, calcium chloride and potassium phosphate) was carried out to screen those that have a significant influence on heat resistance of produced spores. The variable exerting greatest influence on G. stearothermophilus heat resistance during sporulation was found to be the initial pH. Lower nutrient concentration and alkaline pH around 8.5 tended to enhance decimal reduction time at 121 °C (D(121°C)). A central composite design enabled a fourfold enhancement in heat resistance, and the model obtained accurately describes positive pH and negative manganese sulfate concentration influence on spore heat resistance.

Guizelini BP; Vandenberghe LP; Sella SR; Soccol CR

2012-12-01

80

Study of the influence of sporulation conditions on heat resistance of Geobacillus stearothermophilus used in the development of biological indicators for steam sterilization.  

Science.gov (United States)

Biological indicators are important tools in infection control via sterilization process monitoring. The use of a standardized spore crop with a well-defined heat resistance will guarantee the quality of a biological indicator. Ambient factors during sporulation can affect spore characteristics and properties, including heat resistance. The aim of this study is to evaluate the main sporulation factors responsible for heat resistance in Geobacillus stearothermophilus, a useful biological indicator for steam sterilization. A sequence of a three-step optimization of variables (initial pH, nutrient concentration, tryptone, peptone, beef extract, yeast extract, manganese sulfate, magnesium sulfate, calcium chloride and potassium phosphate) was carried out to screen those that have a significant influence on heat resistance of produced spores. The variable exerting greatest influence on G. stearothermophilus heat resistance during sporulation was found to be the initial pH. Lower nutrient concentration and alkaline pH around 8.5 tended to enhance decimal reduction time at 121 °C (D(121°C)). A central composite design enabled a fourfold enhancement in heat resistance, and the model obtained accurately describes positive pH and negative manganese sulfate concentration influence on spore heat resistance. PMID:22872104

Guizelini, Belquis P; Vandenberghe, Luciana P S; Sella, Sandra Regina B R; Soccol, Carlos Ricardo

2012-08-08

 
 
 
 
81

Phylogenetic, inter, and intraspecific sequence analysis of spo0A gene of the genus Geobacillus.  

Science.gov (United States)

In this work, the variability of spo0A gene in the genus Geobacillus and applicability of this gene for the taxonomy within this genus were evaluated. The protein Spo0A is the master regulator of the endospore-forming process in the all endospore-forming bacteria. Geobacillus genus-specific primers GEOSPO were designed based on the sequences of Geobacillus spo0A gene available through the public databases. Inter and intraspecific variability of Geobacillus spo0A gene was determined after sequencing of the GEOSPO-PCR products. Geobacillus spo0A sequence analysis showed that three species--Geobacillus thermodenitrificans, G. stearothermophilus, and G. jurassicus--could be easily identified. Similarity between the sequences of these species and the other species were in the range of 83.3%-92.0%. In contrast, intraspecific similarity of G. thermodenitrificans and G. stearothermophilus was high--above 99.0%. Similarity of spo0A sequences of G. subterraneus-G. uzenensis species cluster also matched this interval. Intercluster similarity between G. lituanicus-G. thermoleovorans-G. kaustophilus-G. vulcani and G. thermocatenulatus-G. gargensis-G. caldoxylosilyticus-G. toebii-G. thermoglucosidasius species clusters, as well as interspecific similarity within these two clusters was in the range of the intraspecific similarity determined for G. thermodenitrificans and G. stearothermophilus. It was also determined that spo0A cannot be used as the phylogenetic marker for the genus Geobacillus. PMID:19205799

Kuisiene, Nomeda; Raugalas, Juozas; Chitavichius, Donaldas

2009-02-10

82

Phylogenetic, inter, and intraspecific sequence analysis of spo0A gene of the genus Geobacillus.  

UK PubMed Central (United Kingdom)

In this work, the variability of spo0A gene in the genus Geobacillus and applicability of this gene for the taxonomy within this genus were evaluated. The protein Spo0A is the master regulator of the endospore-forming process in the all endospore-forming bacteria. Geobacillus genus-specific primers GEOSPO were designed based on the sequences of Geobacillus spo0A gene available through the public databases. Inter and intraspecific variability of Geobacillus spo0A gene was determined after sequencing of the GEOSPO-PCR products. Geobacillus spo0A sequence analysis showed that three species--Geobacillus thermodenitrificans, G. stearothermophilus, and G. jurassicus--could be easily identified. Similarity between the sequences of these species and the other species were in the range of 83.3%-92.0%. In contrast, intraspecific similarity of G. thermodenitrificans and G. stearothermophilus was high--above 99.0%. Similarity of spo0A sequences of G. subterraneus-G. uzenensis species cluster also matched this interval. Intercluster similarity between G. lituanicus-G. thermoleovorans-G. kaustophilus-G. vulcani and G. thermocatenulatus-G. gargensis-G. caldoxylosilyticus-G. toebii-G. thermoglucosidasius species clusters, as well as interspecific similarity within these two clusters was in the range of the intraspecific similarity determined for G. thermodenitrificans and G. stearothermophilus. It was also determined that spo0A cannot be used as the phylogenetic marker for the genus Geobacillus.

Kuisiene N; Raugalas J; Chitavichius D

2009-06-01

83

Genetics of thermophilic bacteria. [Bacillus stearothermophilus:a2  

Energy Technology Data Exchange (ETDEWEB)

Organisms adapted to high temperature have evolved a variety of unique solutions to the biochemical problems imposed by this environment. Adaptation is commonly used to describe the biochemical properties of organisms which have become adapted to their environment (genetic adaptation). It can also mean the direct response-at the cellular level-of an organism to changes in temperature (physiological adaptation). Thermophilic bacilli (strains of Bacillus stearothermophilus) can exhibit a variety of biochemical adaptations in response to changes in temperature. These include changes in the composition and stability of the membrane, metabolic potential, the transport of amino acids, regulatory mechanisms, ribose methylation of tRNA, protein thermostability, and nutritional requirements. The objectives of the research were to develop efficient and reliable genetic systems to analyze and manipulate B. Stearothermophilus, and to use these systems initiate a biochemical, molecular, and genetic investigations of genes that are required for growth at high temperature.

Welker, N.E.

1991-01-01

84

Crystal structure of an inverting GH 43 1,5-alpha-L-arabinanase from Geobacillus stearothermophilus complexed with its substrate.  

UK PubMed Central (United Kingdom)

Arabinanases are glycosidases that hydrolyse alpha-(1-->5)- arabinofuranosidic linkages found in the backbone of the pectic polysaccharide arabinan. Here we describe the biochemical characterization and the enzyme-substrate crystal structure of an inverting family 43 arabinanase from Geobacillus stearothermophilus T-6 (AbnB). Based on viscosity and reducing power measurements, and based on product analysis for the hydrolysis of linear arabinan by AbnB, the enzyme works in an endo mode of action. Isothermal titration calorimetry studies of a catalytic mutant with various arabino-oligosaccharides suggested that the enzyme active site can accommodate at least five arabinose units. The crystal structure of AbnB was determined at 1.06 A (1 A=0.1 nm) resolution, revealing a single five-bladed-beta-propeller fold domain. Co-crystallization of catalytic mutants of the enzyme with different substrates allowed us to obtain complex structures of AbnBE201A with arabinotriose and AbnBD147A with arabinobiose. Based on the crystal structures of AbnB together with its substrates, the position of the three catalytic carboxylates: Asp27, the general base; Glu201, the general acid; and Asp147, the pKa modulator, is in agreement with their putative catalytic roles. In the complex structure of AbnBE201A with arabinotriose, a single water molecule is located 2.8 A from Asp27 and 3.7 A from the anomeric carbon. The position of this water molecule is kept via hydrogen bonding with a conserved tyrosine (Tyr229) that is 2.6 A distant from it. The location of this molecule suggests that it can function as the catalytic water molecule in the hydrolysis reaction, resulting in the inversion of the anomeric configuration of the product.

Alhassid A; Ben-David A; Tabachnikov O; Libster D; Naveh E; Zolotnitsky G; Shoham Y; Shoham G

2009-08-01

85

Crystal structure of an inverting GH 43 1,5-alpha-L-arabinanase from Geobacillus stearothermophilus complexed with its substrate.  

Science.gov (United States)

Arabinanases are glycosidases that hydrolyse alpha-(1-->5)- arabinofuranosidic linkages found in the backbone of the pectic polysaccharide arabinan. Here we describe the biochemical characterization and the enzyme-substrate crystal structure of an inverting family 43 arabinanase from Geobacillus stearothermophilus T-6 (AbnB). Based on viscosity and reducing power measurements, and based on product analysis for the hydrolysis of linear arabinan by AbnB, the enzyme works in an endo mode of action. Isothermal titration calorimetry studies of a catalytic mutant with various arabino-oligosaccharides suggested that the enzyme active site can accommodate at least five arabinose units. The crystal structure of AbnB was determined at 1.06 A (1 A=0.1 nm) resolution, revealing a single five-bladed-beta-propeller fold domain. Co-crystallization of catalytic mutants of the enzyme with different substrates allowed us to obtain complex structures of AbnBE201A with arabinotriose and AbnBD147A with arabinobiose. Based on the crystal structures of AbnB together with its substrates, the position of the three catalytic carboxylates: Asp27, the general base; Glu201, the general acid; and Asp147, the pKa modulator, is in agreement with their putative catalytic roles. In the complex structure of AbnBE201A with arabinotriose, a single water molecule is located 2.8 A from Asp27 and 3.7 A from the anomeric carbon. The position of this water molecule is kept via hydrogen bonding with a conserved tyrosine (Tyr229) that is 2.6 A distant from it. The location of this molecule suggests that it can function as the catalytic water molecule in the hydrolysis reaction, resulting in the inversion of the anomeric configuration of the product. PMID:19505290

Alhassid, Anat; Ben-David, Alon; Tabachnikov, Orly; Libster, Dima; Naveh, Einat; Zolotnitsky, Gennady; Shoham, Yuval; Shoham, Gil

2009-07-29

86

Emended descriptions of Geobacillus thermoleovorans and Geobacillus thermocatenulatus.  

Science.gov (United States)

Nineteen thermophilic, aerobic, endospore-forming bacterial strains were subjected to 16S rRNA gene sequence analysis. Eight of these strains had been received as cultures of Geobacillus kaustophilus, G. lituanicus, G. stearothermophilus, 'G. thermoleovorans subsp. stromboliensis', G. vulcani, 'Bacillus caldolyticus', 'B. caldotenax' and 'B. caldovelox', but they showed close relationships with the type strain of G. thermoleovorans, as did two other strains received as G. thermoleovorans. All strains underwent further taxonomic analysis by API and other phenotypic tests and fatty acid methyl ester analysis, and selected strains were analysed for their polar lipids and for DNA relatedness. The 11 strains that formed the G. thermoleovorans 16S rRNA cluster also showed some phenotypic similarities, and DNA relatedness data support the reassignment of the strains received as G. kaustophilus, G. lituanicus, 'G. thermoleovorans subsp. stromboliensis', G. vulcani, 'B. caldolyticus', 'B. caldotenax' and 'B. caldovelox', and one of the G. stearothermophilus strains, as members of the species G. thermoleovorans. Four other strains received as G. kaustophilus were misnamed; two were identified as G. stearothermophilus and two appeared to be closely related to Anoxybacillus rupiensis. One strain received as G. stearothermophilus remained unidentified. On the basis of a single strain, Geobacillus thermocatenulatus was shown to represent a distinct species, but study of the type strain of Geobacillus gargensis showed this species to be a later heterotypic synonym of Geobacillus thermocatenulatus. Emended descriptions of Geobacillus thermoleovorans and Geobacillus thermocatenulatus are therefore presented. PMID:20817844

Dinsdale, Anna E; Halket, Gillian; Coorevits, An; Van Landschoot, Anita; Busse, Hans-Jürgen; De Vos, Paul; Logan, Niall A

2010-09-03

87

Emended descriptions of Geobacillus thermoleovorans and Geobacillus thermocatenulatus.  

UK PubMed Central (United Kingdom)

Nineteen thermophilic, aerobic, endospore-forming bacterial strains were subjected to 16S rRNA gene sequence analysis. Eight of these strains had been received as cultures of Geobacillus kaustophilus, G. lituanicus, G. stearothermophilus, 'G. thermoleovorans subsp. stromboliensis', G. vulcani, 'Bacillus caldolyticus', 'B. caldotenax' and 'B. caldovelox', but they showed close relationships with the type strain of G. thermoleovorans, as did two other strains received as G. thermoleovorans. All strains underwent further taxonomic analysis by API and other phenotypic tests and fatty acid methyl ester analysis, and selected strains were analysed for their polar lipids and for DNA relatedness. The 11 strains that formed the G. thermoleovorans 16S rRNA cluster also showed some phenotypic similarities, and DNA relatedness data support the reassignment of the strains received as G. kaustophilus, G. lituanicus, 'G. thermoleovorans subsp. stromboliensis', G. vulcani, 'B. caldolyticus', 'B. caldotenax' and 'B. caldovelox', and one of the G. stearothermophilus strains, as members of the species G. thermoleovorans. Four other strains received as G. kaustophilus were misnamed; two were identified as G. stearothermophilus and two appeared to be closely related to Anoxybacillus rupiensis. One strain received as G. stearothermophilus remained unidentified. On the basis of a single strain, Geobacillus thermocatenulatus was shown to represent a distinct species, but study of the type strain of Geobacillus gargensis showed this species to be a later heterotypic synonym of Geobacillus thermocatenulatus. Emended descriptions of Geobacillus thermoleovorans and Geobacillus thermocatenulatus are therefore presented.

Dinsdale AE; Halket G; Coorevits A; Van Landschoot A; Busse HJ; De Vos P; Logan NA

2011-08-01

88

Effect of dimer dissociation on activity and thermostability of the alpha-glucuronidase from Geobacillus stearothermophilus: dissecting the different oligomeric forms of family 67 glycoside hydrolases.  

UK PubMed Central (United Kingdom)

The oligomeric organization of enzymes plays an important role in many biological processes, such as allosteric regulation, conformational stability and thermal stability. alpha-Glucuronidases are family 67 glycosidases that cleave the alpha-1,2-glycosidic bond between 4-O-methyl-D-glucuronic acid and xylose units as part of an array of hemicellulose-hydrolyzing enzymes. Currently, two crystal structures of alpha-glucuronidases are available, those from Geobacillus stearothermophilus (AguA) and from Cellvibrio japonicus (GlcA67A). Both enzymes are homodimeric, but surprisingly their dimeric organization is different, raising questions regarding the significance of dimerization for the enzymes' activity and stability. Structural comparison of the two enzymes suggests several elements that are responsible for the different dimerization organization. Phylogenetic analysis shows that the alpha-glucuronidases AguA and GlcA67A can be classified into two distinct subfamilies of bacterial alpha-glucuronidases, where the dimer-forming residues of each enzyme are conserved only within its own subfamily. It seems that the different dimeric forms of AguA and GlcA67A represent the two alternative dimeric organizations of these subfamilies. To study the biological significance of the dimerization in alpha-glucuronidases, we have constructed a monomeric form of AguA by mutating three of its interface residues (W328E, R329T, and R665N). The activity of the monomer was significantly lower than the activity of the wild-type dimeric AguA, and the optimal temperature for activity of the monomer was around 35 degrees C, compared to 65 degrees C of the wild-type enzyme. Nevertheless, the melting temperature of the monomeric protein, 72.9 degrees C, was almost identical to that of the wild-type, 73.4 degrees C. It appears that the dimerization of AguA is essential for efficient catalysis and that the dissociation into monomers results in subtle conformational changes in the structure which indirectly influence the active site region and reduce the activity. Structural and mechanistic explanations for these effects are discussed.

Shallom D; Golan G; Shoham G; Shoham Y

2004-10-01

89

HIV-1 Gag p17 presented as virus-like particles on the E2 scaffold from Geobacillus stearothermophilus induces sustained humoral and cellular immune responses in the absence of IFN? production by CD4+ T cells  

International Nuclear Information System (INIS)

We have constructed stable virus-like particles displaying the HIV-1 Gag(p17) protein as an N-terminal fusion with an engineered protein domain from the Geobacillus stearothermophilus pyruvate dehydrogenase subunit E2. Mice immunized with the Gag(p17)-E2 60-mer scaffold particles mounted a strong and sustained antibody response. Antibodies directed to Gag(p17) were boosted significantly with additional immunizations, while anti-E2 responses reached a plateau. The isotype of the induced antibodies was biased towards IgG1, and the E2-primed CD4+ T cells did not secrete IFN?. Using transgenic mouse model systems, we demonstrated that CD8+ T cells primed with E2 particles were able to exert lytic activity and produce IFN?. These results show that the E2 scaffold represents a powerful vaccine delivery system for whole antigenic proteins or polyepitope engineered proteins, evoking antibody production and antigen specific CTL activity even in the absence of IFN?-producing CD4+ T cells.

2010-11-25

90

Isolation of lipase producing thermophilic bacteria: optimization of production and reaction conditions for lipase from Geobacillus sp.  

Science.gov (United States)

Lipases catalyze the hydrolysis and the synthesis of esters formed from glycerol and long chain fatty acids. Lipases occur widely in nature, but only microbial lipases are commercially significant. In the present study, thirty-two bacterial strains, isolated from soil sample of a hot spring were screened for lipase production. The strain TS-4, which gave maximum activity, was identified as Geobacillus sp. at MTCC, IMTECH, Chandigarh. The isolated lipase producing bacteria were grown on minimal salt medium containing olive oil. Maximal quantities of lipase were produced when 30 h old inoculum was used at 10% (v/v) in production medium and incubated in shaking conditions (150 rpm) for 72 h. The optimal temperature and pH for the bacterial growth and lipase production were found to be 60°C and 9.5, respectively. Maximal enzyme production resulted when mustard oil was used as carbon source and yeast extract as sole nitrogen source at a concentration of 1% (v/v) and 0.15% (w/v), respectively. The different optimized reaction parameters were temperature 65°C, pH 8.5, incubation time 10 min and substrate p-nitrophenyl palmitate. The Km and Vmax values of enzyme were found to be 14 mM and 17.86 ?mol ml-1min-1, respectively, with p-nitrophenyl palmitate as substrate. All metal ions studied (1 mM) increased the lipase activity. PMID:23195552

Mehta, Akshita; Kumar, Rakesh; Gupta, Reena

2012-12-01

91

CHARACTERIZATION OF CARBOXYMETHYLCELLULASE ACTIVITY FROM GEOBACILLUS STEAROTHERMOPHILUS  

Science.gov (United States)

One of the technological impediments to widespread utilization of lignocellulosic biomass as a fermentation feedstock is the efficient and economical depolymerization of the polysaccharides found in cellulose and hemicellulose. A rational strategy toward overcoming this hurdle is the isolation of h...

92

Application of pheB as a reporter gene for Geobacillus spp., enabling qualitative colony screening and quantitative analysis of promoter strength.  

Science.gov (United States)

The pheB gene from Geobacillus stearothermophilus DSM6285 has been exploited as a reporter gene for Geobacillus spp. The gene product, catechol 2,3-dioxygenase (C23O), catalyzes the formation of 2-hydroxymuconic semialdehyde, which can be readily assayed. The reporter was used to examine expression from the ldh promoter associated with fermentative metabolism. PMID:22685159

Bartosiak-Jentys, Jeremy; Eley, Kirstin; Leak, David J

2012-06-08

93

Application of pheB as a reporter gene for Geobacillus spp., enabling qualitative colony screening and quantitative analysis of promoter strength.  

UK PubMed Central (United Kingdom)

The pheB gene from Geobacillus stearothermophilus DSM6285 has been exploited as a reporter gene for Geobacillus spp. The gene product, catechol 2,3-dioxygenase (C23O), catalyzes the formation of 2-hydroxymuconic semialdehyde, which can be readily assayed. The reporter was used to examine expression from the ldh promoter associated with fermentative metabolism.

Bartosiak-Jentys J; Eley K; Leak DJ

2012-08-01

94

Enhancing the Cellulose-Degrading Activity of Cellulolytic Bacteria CTL-6 (Clostridium thermocellum) by Co-Culture with Non-cellulolytic Bacteria W2-10 (Geobacillus sp.).  

UK PubMed Central (United Kingdom)

The effect of a non-cellulolytic bacterium W2-10 (Geobacillus sp.) on the cellulose-degrading activity of a cellulolytic bacterium CTL-6 (Clostridium thermocellum) was determined using cellulose materials (paper and straw) in peptone cellulose solution (PCS) medium under aerobic conditions. The results indicated that in the co-culture, addition of W2-10 resulted in a balanced medium pH, and may provide the required anaerobic environment for CTL-6. Overall, addition of W2-10 was beneficial to CTL-6 growth in the adverse environment of the PCS medium. In co-culture with W2-10, the CTL-6 cellulose degradation efficiency of filter paper and alkaline-treated wheat straw significantly increased up to 72.45 and 37.79 %, respectively. The CMCase activity and biomass of CTL-6 also increased from 0.23 U ml(-1) and 45.1 ?g ml(-1) (DNA content) up to 0.47 U ml(-1) and 112.2 ?g ml(-1), respectively. In addition, co-culture resulted in accumulation of acetate and propionate up to 4.26 and 2.76 mg ml(-1). This was a respective increase of 2.58 and 4.45 times, in comparison to the monoculture with CTL-6.

Lü Y; Li N; Yuan X; Hua B; Wang J; Ishii M; Igarashi Y; Cui Z

2013-08-01

95

Effect of mercaptoethylamine on DNA degradation in thermophilic bacteria Bac. stearothermophilus exposed to. gamma. -, UV-radiation or methylnitrosourea  

Energy Technology Data Exchange (ETDEWEB)

The effect of mercaptoethylamine (MEA) on degradation of DNA in thermophilic bacteria Bac. stear. exposed to ..gamma..-, UV-rays or methylnitrosourea (MNU) was studied. Using centrifugation on alkaline and neutral sucrose gradients, it was shown that MEA inhibits the accumulation of breaks in the DNA of Bac. stear. It also lowers the level of DNA degradation in toluene-treated cells of Bac. stear. under the action of the intrinsic nuclease, reduces the activity of the endonuclease specific for apurinic DNA, as well as that of S/sub 1/-nuclease and DNase-I in vitro. The inhibition in the accumulation of DNA breaks is assumed to be due to a decrease of the endonuclease activity in the cells of thermophilic bacteria.

Fomenko, L.A.; Kuznetsovea, E.A.; Gaziev, A.I.

1984-07-01

96

Bacillus stearothermophilus from Saudi Arabian Soils.  

UK PubMed Central (United Kingdom)

Ten thermophilic Gram-positive bacteria were isolated from various soils of Saudi Arabia. The strains are spore-forming rods belonging to the species Bacillus stearothermophilus. The cells are motile, strictly aerobic, catalase and oxidase positive. The sporangia appear to be swollen and their position varies from terminal in some to sub-terminal in others. The thermal stability of some enzymes of these bacteria was investigated; extracellular alpha-amylase appears to be very sensitive to pH and temperature. The ultrastructure of these bacteria shows specific changes in the cell wall when grown at the maximum, minimum and optimum growth temperatures, respectively.

Abu-Zinada AH; Hossain A; Yonis HI; Elwan SH

1981-01-01

97

Bacillus stearothermophilus from Saudi Arabian Soils.  

Science.gov (United States)

Ten thermophilic Gram-positive bacteria were isolated from various soils of Saudi Arabia. The strains are spore-forming rods belonging to the species Bacillus stearothermophilus. The cells are motile, strictly aerobic, catalase and oxidase positive. The sporangia appear to be swollen and their position varies from terminal in some to sub-terminal in others. The thermal stability of some enzymes of these bacteria was investigated; extracellular alpha-amylase appears to be very sensitive to pH and temperature. The ultrastructure of these bacteria shows specific changes in the cell wall when grown at the maximum, minimum and optimum growth temperatures, respectively. PMID:7319400

Abu-Zinada, A H; Hossain, A; Yonis, H I; Elwan, S H

1981-01-01

98

Efficacy of high hydrostatic pressure and mild heat to reduce Geobacillus stearothermophillus as 1.1923 spores in model food systems  

UK PubMed Central (United Kingdom)

The objective of this investigation was to evaluate the efficacy of high hydrostatic pressure and mild heat against spores of Geobacillus stearothermophilus AS 1.1923 in model food systems. The pressure-processing conditions were fixed at 625.0 MPa and 86C for 14 min, which have been determined as the optimum processing conditions considering six-log-cycle reductions of G. stearothermophilus spores. Based on the results, response surface methodology was performed in the present investigation, the effects from food ingredients, such as soybean protein, soybean oil and sucrose, as well as pH of the food matrix on the inactivation of G. stearothermophilus spores by high pressure and mild heat was explored, and a quadratic predictive model for the effects of food ingredients and pH on the reduction levels of G. stearothermophilus spores by high-pressure processing was built. The predictive model is significant because the level of significance was P < 0.0001 and the calculated F value is much greater than the tabulated F value. Moreover, the adequacy of the model equation for predicting the reduction of G. stearothermophilus spores was verified effectively. This paper clearly demonstrated the contributions to pressure resistance from food ingredients such as soybean protein, soybean oil, sucrose and pH of the food matrix to the inactivation of Geobacillus stearothermophilus spores by high hydrostatic pressure and mild heat. The predictive model for predicting the reduction of G. stearothermophilus spores in model food systems was built which can be beneficial to targeted process development toward high-pressure sterilization of foods.

Gao YL

2010-02-01

99

Intracellular Proteases of Bacillus stearothermophilus  

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Cell-free extracts of Bacillus stearothermophilus have been shown to exhibit proteolytic activity toward casein as well as specific activity to catalyze the hydrolysis of furylacryloylglycyl-l-leucine amide, furylacryloylglycine, and carbobenzoxyl-glycine-p-nitrophenyl ester, indicating the presence...

Feder, Joseph; Ladenburg, Kurt; Delente, Jacques; Wildi, B. S.

100

Isolation of the phe-operon from G. stearothermophilus comprising the phenol degradative meta-pathway genes and a novel transcriptional regulator.  

UK PubMed Central (United Kingdom)

BACKGROUND: Geobacillus stearothermophilus is able to utilize phenol as a sole carbon source. A DNA fragment encoding a phenol hydroxylase catalyzing the first step in the meta-pathway has been isolated previously. Based on these findings a PCR-based DNA walk was performed initially to isolate a catechol 2,3-dioxygenase for biosensoric applications but was continued to elucidate the organisation of the genes encoding the proteins for the metabolization of phenol. RESULTS: A 20.2 kb DNA fragment was isolated as a result of the DNA walk. Fifteen open reading frames residing on a low-copy megaplasmid were identified. Eleven genes are co-transcribed in one polycistronic mRNA as shown by reverse transcription-PCR. Ten genes encode proteins, that are directly linked with the meta-cleavage pathway. The deduced amino acid sequences display similarities to a two-component phenol hydroxylase, a catechol 2,3-dioxygenase, a 4-oxalocrotonate tautomerase, a 2-oxopent-4-dienoate hydratase, a 4-oxalocrotonate decarboxylase, a 4-hydroxy-2-oxovalerate aldolase, an acetaldehyde dehydrogenase, a plant-type ferredoxin involved in the reactivation of extradiol dioxygenases and a novel regulatory protein. The only enzymes missing for the complete mineralization of phenol are a 2-hydroxymuconic acid-6-semialdehyde hydrolase and/or 2-hydroxymuconic acid-6-semialdehyde dehydrogenase. CONCLUSION: Research on the bacterial degradation of aromatic compounds on a sub-cellular level has been more intensively studied in gram-negative organisms than in gram-positive bacteria. Especially regulatory mechanisms in gram-positive (thermophilic) prokaryotes remain mostly unknown. We isolated the first complete sequence of an operon from a thermophilic bacterium encoding the meta-pathway genes and analyzed the genetic organization. Moreover, the first transcriptional regulator of the phenol metabolism in gram-positive bacteria was identified. This is a first step to elucidate regulatory mechanisms that are likely to be distinct from modes described for gram-negative bacteria.

Omokoko B; Jäntges UK; Zimmermann M; Reiss M; Hartmeier W

2008-01-01

 
 
 
 
101

Cloning and Expression of Thermostable ?-Amylase Gene from Bacillus stearothermophilus in Bacillus stearothermophilus and Bacillus subtilis  

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The structural gene for a thermostable ?-amylase from Bacillus stearothermophilus was cloned in plasmids pTB90 and pTB53. It was expressed in both B. stearothermophilus and Bacillus subtilis. B. stearothermophilus carrying the recombinant plasmid produced about fivefold more ?-amylase (20.9 U/mg of ...

Aiba, Shuichi; Kitai, Kazuo; Imanaka, Tadayuki

102

Tracking spore-forming bacteria in food: from natural biodiversity to selection by processes.  

UK PubMed Central (United Kingdom)

Sporeforming bacteria are ubiquitous in the environment and exhibit a wide range of diversity leading to their natural prevalence in foodstuff. The state of the art of sporeformer prevalence in ingredients and food was investigated using a multiparametric PCR-based tool that enables simultaneous detection and identification of various genera and species mostly encountered in food, i.e., Alicyclobacillus, Anoxybacillus flavithermus, Bacillus, B. cereus group, B. licheniformis, B. pumilus, B. sporothermodurans, B. subtilis, Brevibacillus laterosporus, Clostridium, Geobacillus stearothermophilus, Moorella and Paenibacillus species. In addition, 16S rDNA sequencing was used to extend identification to other possibly present contaminants. A total of 90 food products, with or without visible trace of spoilage were analysed, i.e., 30 egg-based products, 30 milk and dairy products and 30 canned food and ingredients. Results indicated that most samples contained one or several of the targeted genera and species. For all three tested food categories, 30 to 40% of products were contaminated with both Bacillus and Clostridium. The percentage of contaminations associated with Clostridium or Bacillus represented 100% in raw materials, 72% in dehydrated ingredients and 80% in processed foods. In the last two product types, additional thermophilic contaminants were identified (A. flavithermus, Geobacillus spp., Thermoanaerobacterium spp. and Moorella spp.). These results suggest that selection, and therefore the observed (re)-emergence of unexpected sporeforming contaminants in food might be favoured by the use of given food ingredients and food processing technologies.

Postollec F; Mathot AG; Bernard M; Divanac'h ML; Pavan S; Sohier D

2012-08-01

103

Tracking spore-forming bacteria in food: from natural biodiversity to selection by processes.  

Science.gov (United States)

Sporeforming bacteria are ubiquitous in the environment and exhibit a wide range of diversity leading to their natural prevalence in foodstuff. The state of the art of sporeformer prevalence in ingredients and food was investigated using a multiparametric PCR-based tool that enables simultaneous detection and identification of various genera and species mostly encountered in food, i.e., Alicyclobacillus, Anoxybacillus flavithermus, Bacillus, B. cereus group, B. licheniformis, B. pumilus, B. sporothermodurans, B. subtilis, Brevibacillus laterosporus, Clostridium, Geobacillus stearothermophilus, Moorella and Paenibacillus species. In addition, 16S rDNA sequencing was used to extend identification to other possibly present contaminants. A total of 90 food products, with or without visible trace of spoilage were analysed, i.e., 30 egg-based products, 30 milk and dairy products and 30 canned food and ingredients. Results indicated that most samples contained one or several of the targeted genera and species. For all three tested food categories, 30 to 40% of products were contaminated with both Bacillus and Clostridium. The percentage of contaminations associated with Clostridium or Bacillus represented 100% in raw materials, 72% in dehydrated ingredients and 80% in processed foods. In the last two product types, additional thermophilic contaminants were identified (A. flavithermus, Geobacillus spp., Thermoanaerobacterium spp. and Moorella spp.). These results suggest that selection, and therefore the observed (re)-emergence of unexpected sporeforming contaminants in food might be favoured by the use of given food ingredients and food processing technologies. PMID:22795797

Postollec, Florence; Mathot, Anne-Gabrielle; Bernard, Muriel; Divanac'h, Marie-Laure; Pavan, Sonia; Sohier, Danièle

2012-03-09

104

ELECTRON TRANSPORT PARTICLES FROM BACILLUS STEAROTHERMOPHILUS1  

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Downey, Ronald J. (University of Nebraska, Lincoln), Carl E. Georgi, and Walter E. Militzer. Electron transport particles from Bacillus stearothermophilus. J. Bacteriol. 83:1140–1146. 1962—Electron-transport particles (ETP) have been isolated from Bacillus stearothermophilus. They are capable of oxi...

Downey, Ronald J.; Georgi, Carl E.; Militzer, Walter E.

105

ISOLATION AND CHARACTERIZATION OF MANNANOLYTIC THERMOPHILIC BACTERIA FROM PALM OIL SHELL AND THEIR MANNANASE ENZYME PRODUCTION PROPERTIES  

Directory of Open Access Journals (Sweden)

Full Text Available A mannanolytic thermophilic bacterium (L-07) was isolated from palm oil shell after 2 days of enrichment in liquid medium supplemented with 1% palm kernel meal as mannan source. Sequence analysis of 16S-rRNA indicated that L-07 was similar (98%) to Geobacillus stearothermophilus, a species of thermophilic aerobi c bacteria. We found that G. stearothermophilus L-07 produced extracellular ? -1,4-mannanases, but no ? -manosidase and ? -galactosidase activities. The growth of L-07 reached its maximum (3.0 x 106 cell/ml) at 12-20 hours, while the highest ? -mannanase activity (0.52 U/ml) was observed in culture medium after 36 hours of cultivation at 60oC. The medium containing locust bean gum was the best for producing extracellular ? -1,4-mannanases compared with kolang kaling , konjak , and palm kernel meal. SDS-PAGE and zymogram analysis demonstrated that crude mannanase complex of L-07 from locust bean gum containing medium comprised three active bands with molecular weight of 85, 73 and 50 kDa.

SUMARDI; ANTONIUS SUWANTO; MAGGY T HENAWIDJAJA; T RESNAWATI P URWADARIA

2005-01-01

106

Intracellular proteases of Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Cell-free extracts of Bacillus stearothermophilus have been shown to exhibit proteolytic activity toward casein as well as specific activity to catalyze the hydrolysis of furylacryloylglycyl-l-leucine amide, furylacryloylglycine, and carbobenzoxyl-glycine-p-nitrophenyl ester, indicating the presence of a neutral proteinase, a carboxypeptidase-like enzyme, and an alkaline proteinase. The neutral proteinase and carboxypeptidase-like activities were separated by gel filtration over Bio-Gel P-60, and both were reversibly inhibited by 1, 10-phenanthroline. The esterase activity was inhibited by diisopropylfluorophosphate, which did not affect other enzymatic activities and was insensitive to 1, 10-phenanthroline and ethylenediaminetetra-acetic acid.

Feder J; Ladenburg K; Delente J; Wildi BS

1971-12-01

107

Intracellular proteases of Bacillus stearothermophilus.  

Science.gov (United States)

Cell-free extracts of Bacillus stearothermophilus have been shown to exhibit proteolytic activity toward casein as well as specific activity to catalyze the hydrolysis of furylacryloylglycyl-l-leucine amide, furylacryloylglycine, and carbobenzoxyl-glycine-p-nitrophenyl ester, indicating the presence of a neutral proteinase, a carboxypeptidase-like enzyme, and an alkaline proteinase. The neutral proteinase and carboxypeptidase-like activities were separated by gel filtration over Bio-Gel P-60, and both were reversibly inhibited by 1, 10-phenanthroline. The esterase activity was inhibited by diisopropylfluorophosphate, which did not affect other enzymatic activities and was insensitive to 1, 10-phenanthroline and ethylenediaminetetra-acetic acid. PMID:5002894

Feder, J; Ladenburg, K; Delente, J; Wildi, B S

1971-12-01

108

Inactivation of Geobacillus stearothermophilus Spores by High-Pressure Carbon Dioxide Treatment  

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High-pressure CO2 treatment has been studied as a promising method for inactivating bacterial spores. In the present study, we compared this method with other sterilization techniques, including heat and pressure treatment. Spores of Bacillus coagulans, Bacillus subtilis, Bacillus cereus, Bacillus l...

Watanabe, Taisuke; Furukawa, Soichi; Hirata, Junichi; Koyama, Tetsuya; Ogihara, Hirokazu; Yamasaki, Makari

109

Isolation of the phe-operon from G. stearothermophilus comprising the phenol degradative meta-pathway genes and a novel transcriptional regulator  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Geobacillus stearothermophilus is able to utilize phenol as a sole carbon source. A DNA fragment encoding a phenol hydroxylase catalyzing the first step in the meta-pathway has been isolated previously. Based on these findings a PCR-based DNA walk was performed initially to isolate a catechol 2,3-dioxygenase for biosensoric applications but was continued to elucidate the organisation of the genes encoding the proteins for the metabolization of phenol. Results A 20.2 kb DNA fragment was isolated as a result of the DNA walk. Fifteen open reading frames residing on a low-copy megaplasmid were identified. Eleven genes are co-transcribed in one polycistronic mRNA as shown by reverse transcription-PCR. Ten genes encode proteins, that are directly linked with the meta-cleavage pathway. The deduced amino acid sequences display similarities to a two-component phenol hydroxylase, a catechol 2,3-dioxygenase, a 4-oxalocrotonate tautomerase, a 2-oxopent-4-dienoate hydratase, a 4-oxalocrotonate decarboxylase, a 4-hydroxy-2-oxovalerate aldolase, an acetaldehyde dehydrogenase, a plant-type ferredoxin involved in the reactivation of extradiol dioxygenases and a novel regulatory protein. The only enzymes missing for the complete mineralization of phenol are a 2-hydroxymuconic acid-6-semialdehyde hydrolase and/or 2-hydroxymuconic acid-6-semialdehyde dehydrogenase. Conclusion Research on the bacterial degradation of aromatic compounds on a sub-cellular level has been more intensively studied in gram-negative organisms than in gram-positive bacteria. Especially regulatory mechanisms in gram-positive (thermophilic) prokaryotes remain mostly unknown. We isolated the first complete sequence of an operon from a thermophilic bacterium encoding the meta-pathway genes and analyzed the genetic organization. Moreover, the first transcriptional regulator of the phenol metabolism in gram-positive bacteria was identified. This is a first step to elucidate regulatory mechanisms that are likely to be distinct from modes described for gram-negative bacteria.

Omokoko Bastian; Jäntges Uwe K; Zimmermann Martin; Reiss Monika; Hartmeier Winfried

2008-01-01

110

Effect of temperature on the viability of Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

One of the obligate thermophilic bacteria, Bacillus stearothermophilus, was unable to grow at temperatures below 35 degrees C. About 80% of the population in the bacterial culture died at the temperatures, and the same extent of loss in either of the activities of oxygen consumption or synthesis of protein or nucleic acid of the organisms was observed. With the progress of death of the organisms, reduced nicotinamide-adenine dinucleotide came to be oxidized by the organisms, enzymes such as fructose-1,6-diphosphate aldolase, when the organisms were washed with phosphate buffer, were leaked out of the organisms, and an increasing amount of ribonucleoprotein was released into the culture medium. The change of the membrane state was then suggested to be one of the possible causes for the death of the organisms at the temperatures.

Hashizume S; Sekiguchi T; Nosoh Y

1976-02-01

111

Effect of temperature on the viability of Bacillus stearothermophilus.  

Science.gov (United States)

One of the obligate thermophilic bacteria, Bacillus stearothermophilus, was unable to grow at temperatures below 35 degrees C. About 80% of the population in the bacterial culture died at the temperatures, and the same extent of loss in either of the activities of oxygen consumption or synthesis of protein or nucleic acid of the organisms was observed. With the progress of death of the organisms, reduced nicotinamide-adenine dinucleotide came to be oxidized by the organisms, enzymes such as fructose-1,6-diphosphate aldolase, when the organisms were washed with phosphate buffer, were leaked out of the organisms, and an increasing amount of ribonucleoprotein was released into the culture medium. The change of the membrane state was then suggested to be one of the possible causes for the death of the organisms at the temperatures. PMID:175753

Hashizume, S; Sekiguchi, T; Nosoh, Y

1976-02-01

112

Construction and expression of an ethanol production operon in Gram-positive bacteria.  

Science.gov (United States)

Pyruvate decarboxylase (PDC), an enzyme central to homoethanol fermentation, catalyses the non-oxidative decarboxylation of pyruvate to acetaldehyde with release of carbon dioxide. PDC enzymes from diverse organisms have different kinetic properties, thermal stability and codon usage that are likely to offer unique advantages for the development of desirable Gram-positive biocatalysts for use in the ethanol industry. To examine this further, pdc genes from bacteria to yeast were expressed in the Gram-positive host Bacillus megaterium. The PDC activity and protein levels were determined for each strain. In addition, the levels of pdc-specific mRNA transcripts and stability of recombinant proteins were assessed. From this analysis, the pdc gene of Gram-positive Sarcina ventriculi was found to be the most advantageous for engineering high-level synthesis of PDC in a Gram-positive host. This gene was thus selected for transcriptional coupling to the alcohol dehydrogenase gene (adh) of Geobacillus stearothermophilus. The resulting Gram-positive ethanol production operon was expressed at high levels in B. megaterium. Extracts from this recombinant were shown to catalyse the production of ethanol from pyruvate. PMID:16339947

Talarico, Lee A; Gil, Malgorzata A; Yomano, Lorraine P; Ingram, Lonnie O; Maupin-Furlow, Julie A

2005-12-01

113

Molecular cloning of a thermostable neutral protease gene from Bacillus stearothermophilus in a vector plasmid and its expression in Bacillus stearothermophilus and Bacillus subtilis.  

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The structural gene for a thermostable protease from Bacillus stearothermophilus was cloned in plasmid pTB90. It is expressed in both B. stearothermophilus and Bacillus subtilis. B. stearothermophilus carrying the recombinant plasmid produced about 15-fold more protease (310 U/mg of cell dry weight)...

Fujii, M; Takagi, M; Imanaka, T; Aiba, S

114

Identifying critical unrecognized sugar-protein interactions in GH10 xylanases from Geobacillus stearothermophilus using STD NMR.  

UK PubMed Central (United Kingdom)

(1) H solution NMR spectroscopy is used synergistically with 3D crystallographic structures to map experimentally significant hydrophobic interactions upon substrate binding in solution under thermodynamic equilibrium. Using saturation transfer difference spectroscopy (STD NMR), a comparison is made between wild-type xylanase XT6 and its acid/base catalytic mutant E159Q - a non-active, single-heteroatom alteration that has been previously utilized to measure binding thermodynamics across a series of xylooligosaccharide-xylanase complexes [Zolotnitsky et al. (2004) Proc Natl Acad Sci USA 101, 11275-11280). In this study, performing STD NMR of one substrate screens binding interactions to two proteins, avoiding many disadvantages inherent to the technique and clearly revealing subtle changes in binding induced upon mutation of the catalytic Glu. To visualize and compare the binding epitopes of xylobiose-xylanase complexes, a 'SASSY' plot (saturation difference transfer spectroscopy) is used. Two extraordinarily strong, but previously unrecognized, non-covalent interactions with H2 -5 of xylobiose were observed in the wild-type enzyme but not in the E159Q mutant. Based on the crystal structure, these interactions were assigned to tryptophan residues at the -1 subsite. The mutant selectively binds only the ?-xylobiose anomer. The (1) H solution NMR spectrum of a xylotriose-E159Q complex displays non-uniform broadening of the NMR signals. Differential broadening provides a unique subsite assignment tool based on structural knowledge of face-to-face stacking with a conserved tyrosine residue at the +1 subsite. The results obtained herein by substrate-observed NMR spectroscopy are discussed further in terms of methodological contributions and mechanistic understanding of substrate-binding adjustments upon a charge change in the E159Q construct.

Balazs YS; Lisitsin E; Carmiel O; Shoham G; Shoham Y; Schmidt A

2013-09-01

115

Use of soybean vinasses as a germinant medium for a Geobacillus stearothermophilus ATCC 7953 sterilization biological indicator.  

UK PubMed Central (United Kingdom)

A novel low-cost medium was developed from by-products and wastes from the ethanol agro-industry to replace commercial media in the production of a steam sterilization biological indicator (BI). Various recovery media were developed using soybean or sugarcane molasses and vinasse to prepare a self-contained BI. Media performance was evaluated by viability and heat resistance (D(121 °C) value) according to regulatory standards. A medium produced with a soybean vinasse ratio of 1:70 (1.4%) (w/v) produced the results, with D(121 °C)=2.9±0.5 min and Usk=12.7±2.1 min. The addition of 0.8% (w/v) yeast extract improved the germination of heat-damaged spores. The pH variation from 6.0 to 7.3 resulted in a gradual increase in the D(121 °C) value. The absence of calcium chloride resulted in a decrease in germination, while no significant differences were observed with starch addition. Soybean vinasses may thus be used as the main component of a culture medium to substitute for commercial media in the production of self-contained biological indicators. The use of ethanol production waste in this biotechnological process realized a reliable performance, minimized the environmental impact, and decreased BI production costs while producing a high quality product.

Dlugokenski RE; Sella SR; Guizelini BP; Vandenberghe LP; Woiciechowski AL; Soccol CR; Minozzo JC

2011-04-01

116

Identifying critical unrecognized sugar-protein interactions in GH10 xylanases from Geobacillus stearothermophilus using STD NMR.  

Science.gov (United States)

(1) H solution NMR spectroscopy is used synergistically with 3D crystallographic structures to map experimentally significant hydrophobic interactions upon substrate binding in solution under thermodynamic equilibrium. Using saturation transfer difference spectroscopy (STD NMR), a comparison is made between wild-type xylanase XT6 and its acid/base catalytic mutant E159Q - a non-active, single-heteroatom alteration that has been previously utilized to measure binding thermodynamics across a series of xylooligosaccharide-xylanase complexes [Zolotnitsky et al. (2004) Proc Natl Acad Sci USA 101, 11275-11280). In this study, performing STD NMR of one substrate screens binding interactions to two proteins, avoiding many disadvantages inherent to the technique and clearly revealing subtle changes in binding induced upon mutation of the catalytic Glu. To visualize and compare the binding epitopes of xylobiose-xylanase complexes, a 'SASSY' plot (saturation difference transfer spectroscopy) is used. Two extraordinarily strong, but previously unrecognized, non-covalent interactions with H2 -5 of xylobiose were observed in the wild-type enzyme but not in the E159Q mutant. Based on the crystal structure, these interactions were assigned to tryptophan residues at the -1 subsite. The mutant selectively binds only the ?-xylobiose anomer. The (1) H solution NMR spectrum of a xylotriose-E159Q complex displays non-uniform broadening of the NMR signals. Differential broadening provides a unique subsite assignment tool based on structural knowledge of face-to-face stacking with a conserved tyrosine residue at the +1 subsite. The results obtained herein by substrate-observed NMR spectroscopy are discussed further in terms of methodological contributions and mechanistic understanding of substrate-binding adjustments upon a charge change in the E159Q construct. PMID:23863045

Balazs, Yael S; Lisitsin, Elina; Carmiel, Oshrat; Shoham, Gil; Shoham, Yuval; Schmidt, Asher

2013-08-05

117

Surface topography of the Bacillus stearothermophilus ribosome  

International Nuclear Information System (INIS)

[en] The surface topography of the intact 70S ribosome and free 30S and 50S subunits from Bacillus stearothermophilus strain 2,184 was investigated by lactoperoxidase-catalyzed iodination. Two-dimensional polyacrylamide gel electrophoresis was employed to separate ribosomal proteins for analysis of their reactivity. Free 50S subunits incorporated about 18% more 125I than did 50S subunits derived from 70S ribosomes, whereas free 30S subunits and 30S subunits derived from 70S ribosomes incorporated similar amounts of 125I. Iodinated 70S ribosomes and subunits retained 62-78% of the protein synthesis activity of untreated particles and sedimentation profiles showed no gross conformational changes due to iodination. The proteins most reactive to enzymatic iodination were S4, S7, S10 and Sa of the small subunit and L2, L4, L5/9, L6 and L36 of the large subunit. Proteins S2, S3, S7, S13, Sa, L5/9, L10, L11 and L24/25 were labeled substantially more in the free subunits than in the 70S ribosome. Other proteins, including S5, S9, S12, S15/16, S18 and L36 were more extensively iodinated in the 70S ribosome than in the free subunits. The locations of tyrosine residues in some homologus ribosomal proteins from B. stearothermophilus and E. coli are compared. (orig.)[de] Die Oberflaechen-Topographie des intakten 70S-Ribosoms und der freien 30S- und 50S-Untereinheiten des Bacillus stearothermophilus-Stamms 2.184 wurde mit Hilfe der Laktoperoxidase-katalysierten Jodierung untersucht. Die zweidimensionale Polyakrylamid-Gelelektrophorese wurde benutzt, um die ribosomalen Proteine zur Bestimmung ihrer Reaktivitaet abzutrennen. Die freien 50S-Untereinheiten inkorporierten ungefaehr 18% mehr 125J als die von 70S-Ribosomen stammenden 50S-Partikel. Dagegen inkorporierten frei und von 70S-Ribosomen stammende 30S-Partikel etwa die gleiche Menge an 125J. Jodierte 70S-Ribosome und ihre Untereinheiten zeigten 62-78% der Proteinsyntheseaktivitaet unbehandelter Partikel und das Sedimentationsverhalten liess auf keine wesentliche Konformationsaenderung durch die Jodierung schliessen. Die Proteine, die am meisten auf die enzymatische Jodierung ansprachen, waren S4, S7, S10 und Sa auf den 30S-Partikeln und L2, L4, L5/9, L6 und L36 auf den 50S-Partikeln. Die Proteine S2, S3, S7, S13, Sa, L5/9, L10, L11 und L24/25 wurden in den freien Untereinheiten staerker markiert als in dem 70S-Ribosom. Andere Proteine wie S5, S9, S12, S15/16, S18 und L36 waren im 70S-Ribosom staerker markiert. Die Lage der Tyrosin-Abbauprodukte in einigen homologen ribosomalen Proteinen aus B. stearothermophilus und E. coli werden verglichen. (orig./MG)

1976-01-01

118

Cloning and sequence analysis of the heat-stable acrylamidase from a newly isolated thermophilic bacterium, Geobacillus thermoglucosidasius AUT-01.  

UK PubMed Central (United Kingdom)

A thermophilic bacterium capable of degrading acrylamide, AUT-01, was isolated from soil collected from a hot spring area in Montana, USA. The thermophilic strain grew with 0.2 % glucose as the sole carbon source and 1.4 mM acrylamide as the sole nitrogen source. The isolate AUT-01 was identified as Geobacillus thermoglucosidasius based on 16S rDNA sequence. An enzyme from the strain capable of transforming acrylamide to acrylic acid was purified by a series of chromatographic columns. The molecular weight of the enzyme was estimated to be 38 kDa by SDS-PAGE. The enzyme activity had pH and temperature optima of 6.2 and 70 ºC, respectively. The influence of different metals and amino acids on the ability of the purified protein to transform acrylamide to acrylic acid was evaluated. The gene from G. thermoglucosidasius encoding the acrylamidase was cloned, sequenced, and compared to aliphatic amidases from other bacterial strains. The G. thermoglucosidasius gene, amiE, encoded a 38 kDa, monomeric, heat-stable amidase that catalysed the cleavage of carbon-nitrogen bonds in acrylamide. Comparison of the amino acid sequence to other bacterial amidases revealed 99 and 82 % similarity to the amino acid sequences of Bacillus stearothermophilus and Pseudomonas aeruginosa, respectively.

Cha M; Chambliss GH

2013-02-01

119

Isolation and Characterization of a Bacteriocin-Like Substance Produced by Geobacillus toebii Strain HBB-247.  

UK PubMed Central (United Kingdom)

A total of 201 thermophilic bacteria isolated from various thermal spring, mud and soil were tested for their antibacterial activity. Among the mostly active isolates, Geobacillus toebii HBB-247 was further examined. Bacteriocin-like inhibitory substance (BLIS) produced by strain HBB-247 was found to be stable up to 60°C, sensitive to proteolytic enzymes and effective against Enterococcus faecalis, Listeria sp., E. avium, Clostridium pasteurianum, Cellulomonas fimi and some thermophilic strains isolated and identified in this study. As a result of Tricine-SDS-PAGE molecular weight of BLIS was estimated about 38 kDa. Production studies showed that G. toebii HBB-247 starts to produce antibacterial substance at early logarithmic phase of growth and maximum production was detected at the end of the logarithmic phase.

Ba?bülbül Özdemir G; Biyik HH

2012-03-01

120

Isolation and Characterization of a Bacteriocin-Like Substance Produced by Geobacillus toebii Strain HBB-247.  

Science.gov (United States)

A total of 201 thermophilic bacteria isolated from various thermal spring, mud and soil were tested for their antibacterial activity. Among the mostly active isolates, Geobacillus toebii HBB-247 was further examined. Bacteriocin-like inhibitory substance (BLIS) produced by strain HBB-247 was found to be stable up to 60°C, sensitive to proteolytic enzymes and effective against Enterococcus faecalis, Listeria sp., E. avium, Clostridium pasteurianum, Cellulomonas fimi and some thermophilic strains isolated and identified in this study. As a result of Tricine-SDS-PAGE molecular weight of BLIS was estimated about 38 kDa. Production studies showed that G. toebii HBB-247 starts to produce antibacterial substance at early logarithmic phase of growth and maximum production was detected at the end of the logarithmic phase. PMID:23448995

Ba?bülbül Özdemir, Gamze; Biyik, Haci Halil

2011-09-14

 
 
 
 
121

Characterization of aerobic spore-forming bacteria associated with industrial dairy processing environments and product spoilage.  

Science.gov (United States)

Due to changes in the design of industrial food processing and increasing international trade, highly thermoresistant spore-forming bacteria are an emerging problem in food production. Minimally processed foods and products with extended shelf life, such as milk products, are at special risk for contamination and subsequent product damages, but information about origin and food quality related properties of highly heat-resistant spore-formers is still limited. Therefore, the aim of this study was to determine the biodiversity, heat resistance, and food quality and safety affecting characteristics of aerobic spore-formers in the dairy sector. Thus, a comprehensive panel of strains (n=467), which originated from dairy processing environments, raw materials and processed foods, was compiled. The set included isolates associated with recent food spoilage cases and product damages as well as isolates not linked to product spoilage. Identification of the isolates by means of Fourier-transform infrared spectroscopy and molecular methods revealed a large biodiversity of spore-formers, especially among the spoilage associated isolates. These could be assigned to 43 species, representing 11 genera, with Bacillus cereus s.l. and Bacillus licheniformis being predominant. A screening for isolates forming thermoresistant spores (TRS, surviving 100°C, 20min) showed that about one third of the tested spore-formers was heat-resistant, with Bacillus subtilis and Geobacillus stearothermophilus being the prevalent species. Strains producing highly thermoresistant spores (HTRS, surviving 125°C, 30min) were found among mesophilic as well as among thermophilic species. B. subtilis and Bacillus amyloliquefaciens were dominating the group of mesophilic HTRS, while Bacillus smithii and Geobacillus pallidus were dominating the group of thermophilic HTRS. Analysis of spoilage-related enzymes of the TRS isolates showed that mesophilic strains, belonging to the B. subtilis and B. cereus groups, were strongly proteolytic, whereas thermophilic strains displayed generally a low enzymatic activity and thus spoilage potential. Cytotoxicity was only detected in B. cereus, suggesting that the risk of food poisoning by aerobic, thermoresistant spore-formers outside of the B. cereus group is rather low. PMID:23973839

Lücking, Genia; Stoeckel, Marina; Atamer, Zeynep; Hinrichs, Jörg; Ehling-Schulz, Monika

2013-07-16

122

Characterization of aerobic spore-forming bacteria associated with industrial dairy processing environments and product spoilage.  

UK PubMed Central (United Kingdom)

Due to changes in the design of industrial food processing and increasing international trade, highly thermoresistant spore-forming bacteria are an emerging problem in food production. Minimally processed foods and products with extended shelf life, such as milk products, are at special risk for contamination and subsequent product damages, but information about origin and food quality related properties of highly heat-resistant spore-formers is still limited. Therefore, the aim of this study was to determine the biodiversity, heat resistance, and food quality and safety affecting characteristics of aerobic spore-formers in the dairy sector. Thus, a comprehensive panel of strains (n=467), which originated from dairy processing environments, raw materials and processed foods, was compiled. The set included isolates associated with recent food spoilage cases and product damages as well as isolates not linked to product spoilage. Identification of the isolates by means of Fourier-transform infrared spectroscopy and molecular methods revealed a large biodiversity of spore-formers, especially among the spoilage associated isolates. These could be assigned to 43 species, representing 11 genera, with Bacillus cereus s.l. and Bacillus licheniformis being predominant. A screening for isolates forming thermoresistant spores (TRS, surviving 100°C, 20min) showed that about one third of the tested spore-formers was heat-resistant, with Bacillus subtilis and Geobacillus stearothermophilus being the prevalent species. Strains producing highly thermoresistant spores (HTRS, surviving 125°C, 30min) were found among mesophilic as well as among thermophilic species. B. subtilis and Bacillus amyloliquefaciens were dominating the group of mesophilic HTRS, while Bacillus smithii and Geobacillus pallidus were dominating the group of thermophilic HTRS. Analysis of spoilage-related enzymes of the TRS isolates showed that mesophilic strains, belonging to the B. subtilis and B. cereus groups, were strongly proteolytic, whereas thermophilic strains displayed generally a low enzymatic activity and thus spoilage potential. Cytotoxicity was only detected in B. cereus, suggesting that the risk of food poisoning by aerobic, thermoresistant spore-formers outside of the B. cereus group is rather low.

Lücking G; Stoeckel M; Atamer Z; Hinrichs J; Ehling-Schulz M

2013-09-01

123

Strong and consistently synergistic inactivation of spores of spoilage-associated Bacillus and Geobacillus spp. by high pressure and heat compared with inactivation by heat alone.  

UK PubMed Central (United Kingdom)

The inactivation of spores of four low-acid food spoilage organisms by high pressure thermal (HPT) and thermal-only processing was compared on the basis of equivalent thermal lethality calculated at a reference temperature of 121.1°C (F(z)(121.1)(°)(C, 0.1 MPa or 600 MPa)) and characterized as synergistic, not different or protective. In addition, the relative resistances of spores of the different spoilage microorganisms to HPT processing were compared. Processing was performed and inactivation was compared in both laboratory and pilot scale systems and in model (diluted) and actual food products. Where statistical comparisons could be made, at least 4 times and up to around 190 times more inactivation (log(10) reduction/minute at F(T)(z)(121.1)(°)(C)) of spores of Bacillus amyloliquefaciens, Bacillus sporothermodurans, and Geobacillus stearothermophilus was achieved using HPT, indicating a strong synergistic effect of high pressure and heat. Bacillus coagulans spores were also synergistically inactivated in diluted and undiluted Bolognese sauce but were protected by pressure against thermal inactivation in undiluted cream sauce. Irrespective of the response characterization, B. coagulans and B. sporothermodurans were identified as the most HPT-resistant isolates in the pilot scale and laboratory scale studies, respectively, and G. stearothermophilus as the least in both studies and all products. This is the first study to comprehensively quantitatively characterize the responses of a range of spores of spoilage microorganisms as synergistic (or otherwise) using an integrated thermal-lethality approach (F(T)(z)). The use of the F(T)(z) approach is ultimately important for the translation of commercial minimum microbiologically safe and stable thermal processes to HPT processes.

Olivier SA; Bull MK; Stone G; van Diepenbeek RJ; Kormelink F; Jacops L; Chapman B

2011-04-01

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Influence of temperature and organic load on chemical disinfection of Geobacillus steareothermophilus spores, a surrogate for Bacillus anthracis.  

Science.gov (United States)

This study evaluated the influence of temperature and organic load on the effectiveness of domestic bleach (DB), Surface Decontamination Foam (SDF), and Virkon in inactivating Geobacillus stearothermophilus spores, which are a surrogate for Bacillus anthracis spores. The spores were suspended in light or heavy organic preparations and the suspension was applied to stainless steel carrier disks. The dried spore inoculum was covered with the disinfectants and the disks were then incubated at various temperatures. At -20°C, the 3 disinfectants caused less than a 2.0 log10 reduction of spores in both organic preparations during a 24-h test period. At 4°C, the DB caused a 4.4 log10 reduction of spores in light organic preparations within 2 h, which was about 3 log10 higher than what was achieved with SDF or Virkon. In heavy organic preparations, after 24 h at 4°C the SDF had reduced the spore count by 4.5 log10, which was about 2 log10 higher than for DB or Virkon. In general, the disinfectants were most effective at 23°C but a 24-h contact time was required for SDF and Virkon to reduce spore counts in both organic preparations by at least 5.5 log10. Comparable disinfecting activity with DB only occurred with the light organic load. In summary, at temperatures as low as 4°C, DB was the most effective disinfectant, inactivating spores within 2 h on surfaces with a light organic load, whereas SDF produced the greatest reduction of spores within 24 h on surfaces with a heavy organic load. PMID:24082400

Guan, Jiewen; Chan, Maria; Brooks, Brian W; Rohonczy, Liz

2013-04-01

125

Influence of temperature and organic load on chemical disinfection of Geobacillus steareothermophilus spores, a surrogate for Bacillus anthracis.  

UK PubMed Central (United Kingdom)

This study evaluated the influence of temperature and organic load on the effectiveness of domestic bleach (DB), Surface Decontamination Foam (SDF), and Virkon in inactivating Geobacillus stearothermophilus spores, which are a surrogate for Bacillus anthracis spores. The spores were suspended in light or heavy organic preparations and the suspension was applied to stainless steel carrier disks. The dried spore inoculum was covered with the disinfectants and the disks were then incubated at various temperatures. At -20°C, the 3 disinfectants caused less than a 2.0 log10 reduction of spores in both organic preparations during a 24-h test period. At 4°C, the DB caused a 4.4 log10 reduction of spores in light organic preparations within 2 h, which was about 3 log10 higher than what was achieved with SDF or Virkon. In heavy organic preparations, after 24 h at 4°C the SDF had reduced the spore count by 4.5 log10, which was about 2 log10 higher than for DB or Virkon. In general, the disinfectants were most effective at 23°C but a 24-h contact time was required for SDF and Virkon to reduce spore counts in both organic preparations by at least 5.5 log10. Comparable disinfecting activity with DB only occurred with the light organic load. In summary, at temperatures as low as 4°C, DB was the most effective disinfectant, inactivating spores within 2 h on surfaces with a light organic load, whereas SDF produced the greatest reduction of spores within 24 h on surfaces with a heavy organic load.

Guan J; Chan M; Brooks BW; Rohonczy L

2013-04-01

126

Complete genome sequence of the thermophilic bacterium Geobacillus thermoleovorans CCB_US3_UF5.  

UK PubMed Central (United Kingdom)

Geobacillus thermoleovorans CCB_US3_UF5 is a thermophilic bacterium isolated from a hot spring in Malaysia. Here, we report the complete genome of G. thermoleovorans CCB_US3_UF5, which shows high similarity to the genome of Geobacillus kaustophilus HTA 426 in terms of synteny and orthologous genes.

Muhd Sakaff MK; Abdul Rahman AY; Saito JA; Hou S; Alam M

2012-03-01

127

Small-angle X-ray scattering for imaging of surface layers on intact bacteria in the native environment.  

UK PubMed Central (United Kingdom)

Crystalline cell surface layers (S-layers) represent a natural two-dimensional (2D) protein self-assembly system with nanometer-scale periodicity that decorate many prokaryotic cells. Here, we analyze the S-layer on intact bacterial cells of the Gram-positive organism Geobacillus stearothermophilus ATCC 12980 and the Gram-negative organism Aquaspirillum serpens MW5 by small-angle X-ray scattering (SAXS) and relate it to the structure obtained by transmission electron microscopy (TEM) after platinum/carbon shadowing. By measuring the scattering pattern of X rays obtained from a suspension of bacterial cells, integral information on structural elements such as the thickness and lattice parameters of the S-layers on intact, hydrated cells can be obtained nondestructively. In contrast, TEM of whole mounts is used to analyze the S-layer lattice type and parameters as well as the physical structure in a nonaqueous environment and local information on the structure is delivered. Application of SAXS to S-layer research on intact bacteria is a challenging task, as the scattering volume of the generally thin (3- to 30-nm) bacterial S-layers is low in comparison to the scattering volume of the bacterium itself. For enhancement of the scattering contrast of the S-layer in SAXS measurement, either silicification (treatment with tetraethyl orthosilicate) is used, or the difference between SAXS signals from an S-layer-deficient mutant and the corresponding S-layer-carrying bacterium is used for determination of the scattering signal. The good agreement of the SAXS and TEM data shows that S-layers on the bacterial cell surface are remarkably stable.

Sekot G; Schuster D; Messner P; Pum D; Peterlik H; Schäffer C

2013-05-01

128

Small-angle X-ray scattering for imaging of surface layers on intact bacteria in the native environment.  

Science.gov (United States)

Crystalline cell surface layers (S-layers) represent a natural two-dimensional (2D) protein self-assembly system with nanometer-scale periodicity that decorate many prokaryotic cells. Here, we analyze the S-layer on intact bacterial cells of the Gram-positive organism Geobacillus stearothermophilus ATCC 12980 and the Gram-negative organism Aquaspirillum serpens MW5 by small-angle X-ray scattering (SAXS) and relate it to the structure obtained by transmission electron microscopy (TEM) after platinum/carbon shadowing. By measuring the scattering pattern of X rays obtained from a suspension of bacterial cells, integral information on structural elements such as the thickness and lattice parameters of the S-layers on intact, hydrated cells can be obtained nondestructively. In contrast, TEM of whole mounts is used to analyze the S-layer lattice type and parameters as well as the physical structure in a nonaqueous environment and local information on the structure is delivered. Application of SAXS to S-layer research on intact bacteria is a challenging task, as the scattering volume of the generally thin (3- to 30-nm) bacterial S-layers is low in comparison to the scattering volume of the bacterium itself. For enhancement of the scattering contrast of the S-layer in SAXS measurement, either silicification (treatment with tetraethyl orthosilicate) is used, or the difference between SAXS signals from an S-layer-deficient mutant and the corresponding S-layer-carrying bacterium is used for determination of the scattering signal. The good agreement of the SAXS and TEM data shows that S-layers on the bacterial cell surface are remarkably stable. PMID:23504021

Sekot, Gerhard; Schuster, David; Messner, Paul; Pum, Dietmar; Peterlik, Herwig; Schäffer, Christina

2013-03-15

129

Genetic map of the Bacillus stearothermophilus NUB36 chromosome  

Energy Technology Data Exchange (ETDEWEB)

A circular genetic map of Bacillus stearothermophilus NUB36 was constructed by transduction with bacteriophage TP-42C and protoplast fusion. Sixty-four genes were tentatively assigned a cognate Bacillus subtilis gene based on growth response to intermediates or end products of metabolism, cross-feeding, accumulation of intermediates, or their relative order in a linkage group. Although the relative position of many genes on the Bacillus subtilis genetic map appears to be similar, some differences were detected. The tentative order of the genes in the Bacillus stearothermophilus aro region is aspB-aroBAFEC-tyra-hisH-(trp), whereas it is aspB-aroE-tyrA-hisH-(trp)-aroHBF in Bacillus subtilis. The aroA, aroC, and aroG genes in Bacillus subtilis are located in another region. The tentative order of genes in the trp operon of Bacillus stearothermophilus is trpFCDABE, whereas it is trpABFCDE in Bacillus subtilis.

Vallier, H.; Welker, N.E. (Northwestern Univ., Evanston, IL (USA))

1990-02-01

130

ANTIBACTERIAL ACTIVITY OF PAPAYA LEAF EXTRACTS AGAINST PATHOGENIC BACTERIA  

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Full Text Available It was reported that the extracts of papaya leaves could inhibit the growth of Rhizopus stolonifer. Antibacterial activity of Carica papaya leaf extracts on pathogenic bacteria was observed in this study. Papaya leaves were extracted by using maceration method and three kinds of solvents: ethanol, ethyl acetate, and hexane. Papaya leaf extracts were tested against Bacillus stearothermophilus, Listeria monocytogenes, Pseudomonas sp., and Escherichia coli by agar diffusion method. The objectives of this study were to determine extract ability against pathogenic bacteria, to observe the influence of pH, NaCl, and heat on extracts ability, and to observe extract ability against B. stearothermophilus spores. The data showed that ethyl acetate extract could inhibit B. stearothermophilus, L. monocytogenes, Pseudomonas sp., and E. coli. The extract activity was influenced by pH, and it was more effective in low pH. The extract activity was influenced by NaCl against B. stearothermophillus and E. coli. However, it was not influenced by NaCl in bioassay against L. monocytogenes and Pseudomonas sp. The extract activity was influenced by heating process against all the bacteria tested. The extracts inhibited B. stearothermophilus spores as well. Papaya leaves are potential natural anti-bacteria, which might be used in certain kinds of food.

Elisa Friska Romasi; Jessica Karina; Adolf Jan Nexson Parhusip

2011-01-01

131

Transglycosylation of neohesperidin dihydrochalcone by Bacillus stearothermophilus maltogenic amylase.  

UK PubMed Central (United Kingdom)

Neohesperidin dihydrochalcone (NHDC), a sweet compound derived from citrus fruits, was modified to a series of its oligosaccharides by transglycosylation activity of Bacillus stearothermophilus maltogenic amylase (BSMA). Maltotriose as a donor was reacted with NHDC as an acceptor to glycosylate for the purpose of increasing the solubility of NHDC. Maltosyl-NHDC was a major transglycosylation product among the several transfer products by TLC analysis. The structure of the major transglycosylation product was determined to be maltosyl-alpha-(1,6)-neohesperidin dihydrochalcone by MALDI-TOF/MS and (1)H and (13)C NMR. Maltosyl-NHDC was 700 times more soluble in water and 7 times less sweet than NHDC.

Cho JS; Yoo SS; Cheong TK; Kim MJ; Kim Y; Park KH

2000-02-01

132

A commensal symbiotic interrelationship for the growth of Symbiobacterium toebii with its partner bacterium, Geobacillus toebii  

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Full Text Available Abstract Background Symbiobacterium toebii is a commensal symbiotic thermophile that absolutely requires its partner bacterium Geobacillus toebii for growth. Despite development of an independent cultivation method using cell-free extracts, the growth of Symbiobacterium remains unknown due to our poor understanding of the symbiotic relationship with its partner bacterium. Here, we investigated the interrelationship between these two bacteria for growth of S. toebii using different cell-free extracts of G. toebii. Results Symbiobacterium toebii growth-supporting factors were constitutively produced through almost all growth phases and under different oxygen tensions in G. toebii, indicating that the factor may be essential components for growth of G. toebii as well as S. toebii. The growing conditions of G. toebii under different oxygen tension dramatically affected to the initial growth of S. toebii and the retarded lag phase was completely shortened by reducing agent, L-cysteine indicating an evidence of commensal interaction of microaerobic and anaerobic bacterium S. toebii with a facultative aerobic bacterium G. toebii. In addition, the growth curve of S. toebii showed a dependency on the protein concentration of cell-free extracts of G. toebii, demonstrating that the G. toebii-derived factors have nutrient-like characters but not quorum-sensing characters. Conclusions Not only the consistent existence of the factor in G. toebii during all growth stages and under different oxygen tensions but also the concentration dependency of the factor for proliferation and optimal growth of S. toebii, suggests that an important biosynthetic machinery lacks in S. toebii during evolution. The commensal symbiotic bacterium, S. toebii uptakes certain ubiquitous and essential compound for its growth from environment or neighboring bacteria that shares the equivalent compounds. Moreover, G. toebii grown under aerobic condition shortened the lag phase of S. toebii under anaerobic and microaerobic conditions, suggests a possible commensal interaction that G. toebii scavengers ROS/RNS species and helps the initial growth of S. toebii.

Kim Kwang; Kim Joong-Jae; Masui Ryoji; Kuramitsu Seiki; Sung Moon-Hee

2011-01-01

133

Geobacillus zalihae sp. nov., a thermophilic lipolytic bacterium isolated from palm oil mill effluent in Malaysia  

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Full Text Available Abstract Background Thermophilic Bacillus strains of phylogenetic Bacillus rRNA group 5 were described as a new genus Geobacillus. Their geographical distribution included oilfields, hay compost, hydrothermal vent or soils. The members from the genus Geobacillus have a growth temperatures ranging from 35 to 78°C and contained iso-branched saturated fatty acids (iso-15:0, iso-16:0 and iso-17:0) as the major fatty acids. The members of Geobacillus have similarity in their 16S rRNA gene sequences (96.5–99.2%). Thermophiles harboring intrinsically stable enzymes are suitable for industrial applications. The quest for intrinsically thermostable lipases from thermophiles is a prominent task due to the laborious processes via genetic modification. Results Twenty-nine putative lipase producers were screened and isolated from palm oil mill effluent in Malaysia. Of these, isolate T1T was chosen for further study as relatively higher lipase activity was detected quantitatively. The crude T1 lipase showed high optimum temperature of 70°C and was also stable up to 60°C without significant loss of crude enzyme activity. Strain T1T was a Gram-positive, rod-shaped, endospore forming bacterium. On the basic of 16S rDNA analysis, strain T1T was shown to belong to the Bacillus rRNA group 5 related to Geobacillus thermoleovorans (DSM 5366T) and Geobacillus kaustophilus (DSM 7263T). Chemotaxonomic data of cellular fatty acids supported the affiliation of strain T1T to the genus Geobacillus. The results of physiological and biochemical tests, DNA/DNA hybridization, RiboPrint analysis, the length of lipase gene and protein pattern allowed genotypic and phenotypic differentiation of strain T1T from its validly published closest phylogenetic neighbors. Strain T1T therefore represents a novel species, for which the name Geobacillus zalihae sp. nov. is proposed, with the type strain T1T (=DSM 18318T; NBRC 101842T). Conclusion Strain T1T was able to secrete extracellular thermostable lipase into culture medium. The strain T1T was identified as Geobacillus zalihae T1T as it differs from its type strains Geobacillus kaustophilus (DSM 7263T) and Geobacillus thermoleovorans (DSM 5366T) on some physiological studies, cellular fatty acids composition, RiboPrint analysis, length of lipase gene and protein profile.

Rahman Raja; Leow Thean; Salleh Abu; Basri Mahiran

2007-01-01

134

Gene cloning, sequence analysis, purification, and characterization of a thermostable aminoacylase from Bacillus stearothermophilus.  

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A genomic DNA fragment encoding aminoacylase activity of the eubacterium Bacillus stearothermophilus was cloned into Escherichia coli. Transformants expressing aminoacylase activity were selected by their ability to complement E. coli mutants defective in acetylornithine deacetylase activity, the en...

Sakanyan, V; Desmarez, L; Legrain, C; Charlier, D; Mett, I; Kochikyan, A; Savchenko, A; Boyen, A; Falmagne, P; Pierard, A

135

Genetic analysis of Bacillus stearothermophilus by protoplast fusion  

Energy Technology Data Exchange (ETDEWEB)

Efficient and reliable protoplasting, regeneration, and fusion techniques were established for the prototrophic strain Bacillus stearothermophilus NUB36. Auxotrophic mutants were isolated, and protoplast fusion was used to construct isogenic mutant strains and for chromosomal mapping. Markers were mapped using two-, three-, and four-factor crosses. The order of the markers was hom-1-thr-1-his-1-(gly-1 or gly-2)-pur-1-pur-2. These markers may be analogous to hom, thrA, hisA, glyC, and purA markers on the Bacillus subtilis chromosome. No analogous pur-1 marker has been reported in B. subtilis. The relative order of three of the markers (hom-1-thr-1-gly-1) was independently confirmed by transduction.

Chen, Z.; Wojcik, S.F.; Welker, N.E.

1986-03-01

136

1H, 13C, and 15N backbone and side chain resonance assignments of thermophilic Geobacillus kaustophilus cyclophilin-A.  

UK PubMed Central (United Kingdom)

Cyclophilins catalyze the reversible peptidyl-prolyl isomerization of their substrates and are present across all kingdoms of life from humans to bacteria. Although numerous biological roles have now been discovered for cyclophilins, their function was initially ascribed to their chaperone-like activity in protein folding where they catalyze the often rate-limiting step of proline isomerization. This chaperone-like activity may be especially important under extreme conditions where cyclophilins are often over expressed, such as in tumors for human cyclophilins (Lee Archiv Pharm Res 33(2): 181-187, 2010), but also in organisms that thrive under extreme conditions, such as theromophilic bacteria. Moreover, the reversible nature of the peptidyl-prolyl isomerization reaction catalyzed by cyclophilins has allowed these enzymes to serve as model systems for probing the role of conformational changes during catalytic turnover (Eisenmesser et al. Science 295(5559): 1520-1523, 2002; Eisenmesser et al. Nature 438(7064): 117-121, 2005). Thus, we present here the resonance assignments of a thermophilic cyclophilin from Geobacillus kaustophilus derived from deep-sea sediment (Takami et al. Extremophiles 8(5): 351-356, 2004). This thermophilic cyclophilin may now be studied at a variety of temperatures to provide insight into the comparative structure, dynamics, and catalytic mechanism of cyclophilins.

Holliday MJ; Zhang F; Isern NG; Armstrong GS; Eisenmesser EZ

2012-11-01

137

Molecular characterization of the Bacillus stearothermophilus PV72 S-layer gene sbsB induced by oxidative stress.  

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S-layer protein variation from a hexagonally ordered (SbsA; 130 kDa) to a obliquely ordered (SbsB; 98 kDa) protein in Bacillus stearothermophilus PV72 is mediated by an increased oxygen supply. To elucidate the molecular basis of S-layer protein variation in B. stearothermophilus PV72, the sbsB gene...

Kuen, B; Koch, A; Asenbauer, E; Sará, M; Lubitz, W

138

Isozymes of alpha-galactosidase from Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Two molecular forms of alpha-galactosidase (EC 3.2.1.22) synthesized constitutively by Bacillus stearothermophilus, strain AT-7, have been purified. alpha-Galactosidase I (with the substrate p-nitrophenyl alpha-D-galactopyranoside (PNPG)) has a pH optimum of 6 and half-life at 65 degrees C of > 2 h at low protein concentration. alpha-Galactosidase II has a pH optimum of 7 with PNPG and a half-life at 65 degrees C of about 3 min. The isozymes also differ with respect to their Km with PNPG and melibiose. Both enzymes are inhibited competitively by D-galactose, melibiose, and Tris. With the beta-glycosides cellobiose and lactose either noncompetitive or mixed-type inhibition is observed, with the pattern dependent on both the pH and the isozyme. The two isozymes have similar Arrhenius activation energies (about 20 kcal/mol, 1 kcal = 4.184 kJ). Their molecular weights, estimated by disc gel electrophoresis, are alpha-galactosidase I, 280 000 +/- 30 000 and alpha-galactosidase II, 325 000 +/- 15 000. Dodecyl sulfate gel electrophoresis gave a single band for each enzyme. The respective molecular weights, 81 000 +/- 500 for alpha-galactosidase I and 84 000 +/- 500 for alpha-galactosidase II, suggest that both enzymes consist of four subunits.

Pederson DM; Goodman RE

1980-08-01

139

Fluorescent labelling of 6-phosphogluconate dehydrogenase from Bacillus stearothermophilus.  

Science.gov (United States)

The thermophilic 6-phosphogluconate dehydrogenase from Bacillus stearothermophilus was inhibited upon specific modification of the -SH group of cysteine residues by 7-chloro-4-nitrobenzo-2-oxa-1, 3-diazole (NBD-Cl) at pH 7.0. By using 20-100-fold molar excess of NBD-CL the reaction occurs slowly at pH 7.0 as a first order process. Partial protection from inactivation was observed when the substrate 6-phosphogluconate or the coenzyme NADP was added to the reaction mixture. Complete inactivation was achieved upon modification of 1.9 of the six cysteine residues per mole of enzyme, which corresponds to nearly one residue per enzyme subunit. Circular dichroism measurements suggest that the gross structure of the protein molecule is practically unchanged upon reaction of the enzyme with NBD-Cl. Melting profile experiments revealed a single transition occurring at about 65 degrees C. Analogously, the profile of intensity of the fluorescence emission at 520 nm of the enzyme-bound S-NBD groups versus temperature indicated a midpoint of transition near 65 degrees C. Since this melting temperature corresponds closely to that observed with the native enzyme, these results would indicate that the molecular organizations of the native and modified enzyme are similar and stabilized by similar interactions within the polypeptide chain. PMID:19368

Fontana, A; Mantovanelli, L; Boccu, E; Veronese, F M

1977-01-01

140

Detection and characterization of chlorinated-dioxin ether cleavage function in the bacterium geobacillus midousuji SH2B-J2  

Energy Technology Data Exchange (ETDEWEB)

As of now, there are no dioxin degrading microorganism reported that can be applied to bioremediation. The reasons for this are that degrading function acquired from comprehensive screening of bacteria that can be grown with a single carbon source using non-chlorinated dioxin does not function against highly chlorinated dioxins, and that although white rot fungus capable of degrading lignin, a plant polyphenol substance, have been reported to reduce chlorinated dioxins, degrading enzyme remain unclear. Geobacillus midousuji SH2B-J2 (J2 strain) that have been separated by Hoshina et al. have shown to reduce highly chlorinated dioxins in incineration fly ash, as well as octa-chlorinated dioxins (OCDD). However, details of its degrading mechanisms remain unclear. Since the J2 strain is capable of reducing even OCDD, it was hypothesized that the initial degradation reaction is intramolecular ether bond cleavage, so J2 strain dioxin degradation mechanism was analyzed for verification.

Otsuka, Y.; Hoshina, S. [Jikei Univ. School of Medicine, Tokyo (Japan). Dept. of Laboratory Medicine; Nakamura, M.; Hishiyama, S. [Forestry and Forest Products Research Institute, Ibaraki (Japan); Katayama, Y. [Tokyo Univ. of Agriculture and Technology, Koganei (Japan)

2004-09-15

 
 
 
 
141

Catalytic Biomineralization of Fluorescent Calcite by the Thermophilic Bacterium Geobacillus thermoglucosidasius?  

Science.gov (United States)

The thermophilic Geobacillus bacterium catalyzed the formation of 100-?m hexagonal crystals at 60°C in a hydrogel containing sodium acetate, calcium chloride, and magnesium sulfate. Under fluorescence microscopy, crystals fluoresced upon excitation at 365 ± 5, 480 ± 20, or 545 ± 15 nm. X-ray diffraction indicated that the crystals were magnesium-calcite in calcite-type calcium carbonate.

Yoshida, Naoto; Higashimura, Eiji; Saeki, Yuichi

2010-01-01

142

Evidence for an S-layer protein pool in the peptidoglycan of Bacillus stearothermophilus.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Intact cells of Bacillus stearothermophilus PV72 revealed, after conventional thin-sectioning procedures, the typical cell wall profile of S-layer-carrying gram-positive eubacteria consisting of a ca. 10-nm-thick peptidoglycan-containing layer and a ca. 10-nm-thick S layer. Cell wall preparations ob...

Breitwieser, A; Gruber, K; Sleytr, U B

143

Relief of Casein Inhibition of Bacillus stearothermophilus by Iron, Calcium, and Magnesium1  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Growth of Bacillus stearothermophilus strain NCA 1518 Smooth in Dextrose Tryptone Agar (DTA) was inhibited by sodium caseinate. Binding studies indicated that sodium caseinate, when present in DTA, had the capacity to effect an iron deficiency which could cause inhibition of growth. Additions of ess...

Ashton, D. H.; Busta, F. F.; Warren, J. A.

144

Lipid composition and dynamics of cell membranes of Bacillus stearothermophilus adapted to amiodarone  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Bacillus stearothermophilus, a useful model to evaluate membrane interactions of lipophilic drugs, adapts to the presence of amiodarone in the growth medium. Drug concentrations in the range of 1-2 [mu]M depress growth and 3 [mu]M completely suppresses growth. Adaptation to the presence of amiodaron...

Rosa, Sónia M. L. J.; Antunes-Madeira, Maria do Carmo; Matos, Manuel J.; Jurado, Amália S.; Madeira, Vítor M. C.

145

Draft Genome Sequence of Lignocellulose-Degrading Thermophilic Bacterium Geobacillus sp. Strain WSUCF1.  

Science.gov (United States)

Geobacillus sp. strain WSUCF1 is a thermophilic spore-forming member of the phylum Firmicutes, isolated from a soil sample collected from the compost facility. We report the draft genome sequence of this isolate with an estimated genome size of 3.4 Mb. The genome sequence of this isolate revealed several genes encoding glycoside hydrolases, making it a potential candidate for plant biomass degradation. PMID:23950119

Bhalla, Aditya; Kainth, Amoldeep Singh; Sani, Rajesh K

2013-08-15

146

Draft Genome Sequence of Lignocellulose-Degrading Thermophilic Bacterium Geobacillus sp. Strain WSUCF1.  

UK PubMed Central (United Kingdom)

Geobacillus sp. strain WSUCF1 is a thermophilic spore-forming member of the phylum Firmicutes, isolated from a soil sample collected from the compost facility. We report the draft genome sequence of this isolate with an estimated genome size of 3.4 Mb. The genome sequence of this isolate revealed several genes encoding glycoside hydrolases, making it a potential candidate for plant biomass degradation.

Bhalla A; Kainth AS; Sani RK

2013-01-01

147

Gene structure and quinol oxidase activity of a cytochrome bd-type oxidase from Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Gram-positive thermophilic Bacillus species contain cytochrome caa3-type cytochrome c oxidase as their main terminal oxidase in the respiratory chain. We previously identified and purified an alternative oxidase, cytochrome bd-type quinol oxidase, from a mutant of Bacillus stearothermophilus defective in the caa3-type oxidase activity (J. Sakamoto et al., FEMS Microbiol. Lett. 143 (1996) 151-158). Compared with proteobacterial counterparts, B. stearothermophilus cytochrome bd showed lower molecular weights of the two subunits, shorter wavelength of alpha-band absorption maximum due to heme D, and lower quinol oxidase activity. Preincubation with menaquinone-2 enhanced the enzyme activity up to 40 times, suggesting that, besides the catalytic site, there is another quinone-binding site which largely affects the enzyme activity. In order to clarify the molecular basis of the differences of cytochromes bd between B. stearothermophilus and proteobacteria, the genes encoding for the B. stearothermophilus bd was cloned based on its partial peptide sequences. The gene for subunit I (cbdA) encodes 448 amino acid residues with a molecular weight of 50195 Da, which is 14 and 17% shorter than those of Escherichia coli and Azotobacter vinelandii, respectively, and CbdA lacks the C-terminal half of the long hydrophilic loop between the putative transmembrane segments V and VI (Q loop), which has been suggested to include the substrate quinone-binding site for the E. coli enzyme. The gene for subunit II (cbdB) encodes 342 residues with a molecular weight of 38992 Da. Homology search indicated that the B. stearothermophilus cbdAB has the highest sequence similarity to ythAB in B. subtilis genome rather than to cydAB, the first set of cytochrome bd genes identified in the genome. Sequence comparison of cytochromes bd and their homologs from various organisms demonstrates that the proteins can be classified into two subfamilies, a proteobacterial type including E. coli bd and a more widely distributed type including the B. stearothermophilus enzyme, suggesting that the latter type is evolutionarily older.

Sakamoto J; Koga E; Mizuta T; Sato C; Noguchi S; Sone N

1999-04-01

148

Production and Characterization of a Mesophilic Lipase Isolated from Bacillus stearothermophilus AB-1  

Directory of Open Access Journals (Sweden)

Full Text Available Using Bacillus stearothermophilus AB-1 isolated from air, the production of lipase was attempted along with its purification and characterization studies. When different carbon and nitrogen sources were supplemented in the culture medium, xylose, tryptophan, alanine, phenylalanine and potassium nitrate were found to be the best. During cultivation, the strain secreted most of its lipase content after 48 h. In particular, the lipase produced in the culture broth showed 300 U mL-1 when cultivated at optimal temperature and pH of 35 °C and 7.5, respectively. The enzyme was purified using 60% ammonium sulfate precipitation and sephadex G200 column chromatography. The enzyme was stable up to 40 °C and in the range of pH 7-8. This research reports for the first time the characterization of mesophilic lipase from Bacillus stearothermophilus AB-1 isolated from air.

Emad Abd El-moniem Abada

2008-01-01

149

Structure based protein engineering of Bacillus stearothermophilus {alpha}-amylase: toward a new substrate specificity  

Energy Technology Data Exchange (ETDEWEB)

Full text. Structural similarity is observed in all members of {alpha}-amylase family but different products are generated during hydrolysis of starch due to different affinities for intermediate dextrins. In order to understand the structural determinants for this property and to introduce different specificity to {alpha}-amylase of Bacillus stearothermophilus we intend to solve the three dimensional structure by X-ray crystallography of the native protein by using synchrotron radiation at Brazilian National Synchrotron Light Laboratory (LNLS). Protein was over expressed in E. coli, purified and crystallization experiments were carried out by using sparse matrix Crystal Screen and Crystal Screen II from Hampton Research (Laguna Hills, CA, USA). Several condition have produced crystals with some defined characteristic: MDP seems to be important to the crystallization: the preferential pH is around 7.5 with organic buffer (HEPES); organic solvent as 2-propanol seems to be also important for the crystallization. On those condition crystals appeared as cluster of needles or small crystals with high number of nucleation. New conditions are being prepared to improve the site and quality of crystals. Data collection of native of Bacillus stearothermophilus {alpha}-amylase will e done at Protein Crystallography Station at LNLS. Crystal structure of mutated {alpha}-amylase bu site direct mutagenesis of residues suggested by the native crystal structure will be obtained. Co-crystallization of Bacillus stearothermophilus {alpha}-amylase and oligosaccharide will be carried out to identify residues involved in the binding site to plan new mutation. In another series of mutation the putative binding site residues, once identified, will be mutated to residues observed in TAKA amylase to confer different specificity to {alpha}-amylase. Based on the available TAKA amylase structure, in the primary sequence homology and in the three dimensional model of Bacillus stearothermophilus {alpha}-amylase (using Bacillus licheniformis crystal structure as initial model) it seems that Bacillus stearothermophilus {alpha}-amylase binding site is more complex with and insertion of 40 residues. Therefore the three dimensional structure is crucial to understand the specificity of the substrate of this enzyme which will be used to drive the design of mutation to introduce new properties for industrial purpose. (author)

Rasera, Ana Claudia [Sao Paulo Univ., SP (Brazil). Inst. de Ciencias Biomedicas; Iulek, Jorge [Universidade Estadual de Ponta Grossa, PR (Brazil). Inst. de Quimica; Delboni, Luis Fernando; Barbosa, Valma Martins Barbosa [Parana Univ., Curitiba, PR (Brazil). Dept. de Bioquimica

1997-12-31

150

Structure based protein engineering of Bacillus stearothermophilus ?-amylase: toward a new substrate specificity  

International Nuclear Information System (INIS)

Full text. Structural similarity is observed in all members of ?-amylase family but different products are generated during hydrolysis of starch due to different affinities for intermediate dextrins. In order to understand the structural determinants for this property and to introduce different specificity to ?-amylase of Bacillus stearothermophilus we intend to solve the three dimensional structure by X-ray crystallography of the native protein by using synchrotron radiation at Brazilian National Synchrotron Light Laboratory (LNLS). Protein was over expressed in E. coli, purified and crystallization experiments were carried out by using sparse matrix Crystal Screen and Crystal Screen II from Hampton Research (Laguna Hills, CA, USA). Several condition have produced crystals with some defined characteristic: MDP seems to be important to the crystallization: the preferential pH is around 7.5 with organic buffer (HEPES); organic solvent as 2-propanol seems to be also important for the crystallization. On those condition crystals appeared as cluster of needles or small crystals with high number of nucleation. New conditions are being prepared to improve the site and quality of crystals. Data collection of native of Bacillus stearothermophilus ?-amylase will e done at Protein Crystallography Station at LNLS. Crystal structure of mutated ?-amylase bu site direct mutagenesis of residues suggested by the native crystal structure will be obtained. Co-crystallization of Bacillus stearothermophilus ?-amylase and oligosaccharide will be carried out to identify residues involved in the binding site to plan new mutation. In another series of mutation the putative binding site residues, once identified, will be mutated to residues observed in TAKA amylase to confer different specificity to ?-amylase. Based on the available TAKA amylase structure, in the primary sequence homology and in the three dimensional model of Bacillus stearothermophilus ?-amylase (using Bacillus licheniformis crystal structure as initial model) it seems that Bacillus stearothermophilus ?-amylase binding site is more complex with and insertion of 40 residues. Therefore the three dimensional structure is crucial to understand the specificity of the substrate of this enzyme which will be used to drive the design of mutation to introduce new properties for industrial purpose. (author)

1997-01-01

151

Cloning, expression, purification, crystallization and preliminary X-ray crystallographic study of DHNA synthetase from Geobacillus kaustophilus  

Science.gov (United States)

The aerobic Gram-positive bacterium Geobacillus kaustophilus is a bacillus species that was isolated from deep-sea sediment from the Mariana Trench. 1,4-­Dihydroxy-2-naphthoate (DHNA) synthetase plays a vital role in the biosynthesis of menaquinone (vitamin K2) in this bacterium. DHNA synthetase from Geobacillus kaustophilus was crystallized in the orthorhombic space group C2221, with unit-cell parameters a = 77.01, b = 130.66, c = 131.69?Å. The crystal diffracted to a resolution of 2.2?Å. Preliminary studies and molecular-replacement calculations reveal the presence of three monomers in the asymmetric unit.

Kanaujia, Shankar Prasad; Ranjani, Chellamuthu Vasuki; Jeyakanthan, Jeyaraman; Baba, Seiki; Kuroishi, Chizu; Ebihara, Akio; Shinkai, Akeo; Kuramitsu, Seiki; Shiro, Yoshitsugu; Sekar, Kanagaraj; Yokoyama, Shigeyuki

2007-01-01

152

Direct correlationship between proton translocation and growth yield: an analysis of the respiratory chain of Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Thermophilic bacilli contain cytochrome caa3-type cytochrome c oxidase as the main terminal oxidase in the respiratory chain. A mutant strain, named K-17, lacking cytochrome caa3 and exhibiting very low N,N,N',N'-tetramethyl-p-phenylene diamine oxidase activity, was isolated by random mutation from Bacillus stearothermophilus K1041 (Sakamoto, J. et al., FEMS Microbiol. Lett., 143, 151-158, 1996). Comparing this mutant with the parent strain K1041, we observed the following differences in energy-yielding properties. (i) K-17 gave an cell yield less than one half of that of the wild type, although the doubling time of K-17 was only a little slower than that of the parent strain. (ii) In cellular respiration, the H+/O ratio of K-17 was 2.9-3.1, while that of the wild type was 6.1-6.5. (iii) A low concentration of cyanide inhibited endogenous respiration of the wild-type cells partly with a concomitant reduction of the H+/O ratio to around 3, while it did not significantly affect the respiration rate and the H+/O ratio of the K-17 cells. (iv) Cytochrome bd-type quinol oxidase seemed to operate in the wild-type cells when a low concentration (below 0.5 mM) of cyanide was added, while this enzyme is the main terminal oxidase in K-17. The K-17 cells also contained cytochrome b(o/a)3-type cytochrome c-551 oxidase. These results demonstrated that the combination of the enzymes involved in the respiratory chain determines the H+/O ratios of the cell and consequently the growth yield of the bacteria.

Sone N; Tsukita S; Sakamoto J

1999-01-01

153

An effective iodide formulation for killing Bacillus and Geobacillus spores over a wide temperature range.  

UK PubMed Central (United Kingdom)

AIMS: To develop a sporicidal reagent which shows potent activity against bacterial spores not only at ambient temperatures but also at low temperatures. METHODS AND RESULTS: Suspension tests on spores of Bacillus and Geobacillus were conducted with the reagent based on a previously reported agent (N. Kida, Y. Mochizuki and F. Taguchi, Microbiology and Immunology 2003; 47: 279-283). The modified reagent (tentatively designated as the KMT reagent) was composed of 50 mmol l(-1) EDTA-2Na, 50 mmol l(-1) ferric chloride hexahydrate (FeCl(3).6H(2)O), 50 mmol l(-1) potassium iodide (KI) and 50% ethanol in 0.85% NaCl solution at pH 0.3. The KMT reagent showed significant sporicidal activity against three species of Bacillus and Geobacillus spores over a wide range of temperature. The KMT reagent had many practical advantages, i.e. activity was much less affected by organic substances than was sodium hypochlorite, it did not generate any harmful gas and it was stable for a long period at ambient temperatures. The mechanism(s) of sporicidal activity of the KMT reagent was considered to be based on active iodine species penetrating the spores with enhanced permeability of the spore cortex by a synergistic effect of acid, ethanol and generated active oxygen. CONCLUSIONS: The data suggest that the KMT reagent shows potent sporicidal activity over a wide range temperatures and possesses many advantages for practical applications. SIGNIFICANCE AND IMPACT OF THE STUDY: The results indicate development of a highly applicable sporicidal reagent against Bacillus and Geobacillus spores.

Kida N; Mochizuki Y; Taguchi F

2004-01-01

154

Structure of the hypothetical DUF1811-family protein GK0453 from Geobacillus kaustophilus HTA426.  

UK PubMed Central (United Kingdom)

The crystal structure of a conserved hypothetical protein, GK0453, from Geobacillus kaustophilus has been determined to 2.2 Å resolution. The crystal belonged to space group P4(3)2(1)2, with unit-cell parameters a = b = 75.69, c = 64.18 Å. The structure was determined by the molecular-replacement method and was refined to a final R factor of 22.6% (R(free) = 26.3%). Based on structural homology, the GK0453 protein possesses two independent binding sites and hence it may simultaneously interact with two proteins or with a protein and a nucleic acid.

Padmanabhan B; Nakamura Y; Antonyuk SV; Strange RW; Hasnain SS; Yokoyama S; Bessho Y

2013-04-01

155

Structure of the hypothetical DUF1811-family protein GK0453 from Geobacillus kaustophilus HTA426.  

Science.gov (United States)

The crystal structure of a conserved hypothetical protein, GK0453, from Geobacillus kaustophilus has been determined to 2.2 Å resolution. The crystal belonged to space group P4(3)2(1)2, with unit-cell parameters a = b = 75.69, c = 64.18 Å. The structure was determined by the molecular-replacement method and was refined to a final R factor of 22.6% (R(free) = 26.3%). Based on structural homology, the GK0453 protein possesses two independent binding sites and hence it may simultaneously interact with two proteins or with a protein and a nucleic acid. PMID:23545635

Padmanabhan, Balasundaram; Nakamura, Yoshihiro; Antonyuk, Svetlana V; Strange, Richard W; Hasnain, S Samar; Yokoyama, Shigeyuki; Bessho, Yoshitaka

2013-03-28

156

Structure of the hypothetical DUF1811-family protein GK0453 from Geobacillus kaustophilus HTA426  

Science.gov (United States)

The crystal structure of a conserved hypothetical protein, GK0453, from Geobacillus kaustophilus has been determined to 2.2?Å resolution. The crystal belonged to space group P43212, with unit-cell parameters a = b = 75.69, c = 64.18?Å. The structure was determined by the molecular-replacement method and was refined to a final R factor of 22.6% (R free = 26.3%). Based on structural homology, the GK0453 protein possesses two independent binding sites and hence it may simultaneously interact with two proteins or with a protein and a nucleic acid.

Padmanabhan, Balasundaram; Nakamura, Yoshihiro; Antonyuk, Svetlana V.; Strange, Richard W.; Hasnain, S. Samar; Yokoyama, Shigeyuki; Bessho, Yoshitaka

2013-01-01

157

Isolation and characterization of a thermophilic bacterium, Geobacillus thermocatenulatus, degrading nylon 12 and nylon 66.  

Science.gov (United States)

A thermophilic bacterium, identified as a neighboring species to Geobacillus thermocatenulatus, having a growth optimum at 55 degrees C and, capable of degrading nylon 12, was isolated from soil by enrichment culture technique at 60 degrees C. At this temperature, the strain grew on 5 g nylon 12 l(-1) with a decrease in its molecular weight from 41000 to 11000 over 20 d. The degradation was assumed to be due to endogenous hydrolysis of amide bond in nylon 12. The strain degraded also nylon 66 with a decrease in its molecular weight from 43000 to 17000 in 20 d at 60 degrees C. Nylon 6 was not degraded. PMID:14626419

Tomita, Kosuke; Ikeda, Noritoshi; Ueno, Ayumi

2003-10-01

158

Isolation and characterization of a thermophilic bacterium, Geobacillus thermocatenulatus, degrading nylon 12 and nylon 66.  

UK PubMed Central (United Kingdom)

A thermophilic bacterium, identified as a neighboring species to Geobacillus thermocatenulatus, having a growth optimum at 55 degrees C and, capable of degrading nylon 12, was isolated from soil by enrichment culture technique at 60 degrees C. At this temperature, the strain grew on 5 g nylon 12 l(-1) with a decrease in its molecular weight from 41000 to 11000 over 20 d. The degradation was assumed to be due to endogenous hydrolysis of amide bond in nylon 12. The strain degraded also nylon 66 with a decrease in its molecular weight from 43000 to 17000 in 20 d at 60 degrees C. Nylon 6 was not degraded.

Tomita K; Ikeda N; Ueno A

2003-10-01

159

Isolation and characterization of a thermophilic bacterium, Geobacillus thermocatenulatus, degrading nylon 12 and nylon 66.  

UK PubMed Central (United Kingdom)

A thermophilic bacterium, identified as a neighboring species to Geobacillus thermocatenulatus, having a growth optimum at 55°C and, capable of degrading nylon 12, was isolated from soil by enrichment culture technique at 60°C. At this temperature, the strain grew on 5 g nylon 12 l-1 with a decrease in its molecular weight from 41 000 to 11 000 over 20 d. The degradation was assumed to be due to endogenous hydrolysis of amide bond in nylon 12. The strain degraded also nylon 66 with a decrease in its molecular weight from 43 000 to 17 000 in 20 d at 60°C. Nylon 6 was not degraded.

Tomita K; Ikeda N; Ueno A

2003-10-01

160

Production of alpha-amylase in batch and chemostat culture by bacillus stearothermophilus  

Energy Technology Data Exchange (ETDEWEB)

The production of alpha-amylase by a strain of B.stearothermophilus isolated from leaf litter was investigated in a tryptone-maltose medium at 55 degrees in batch and chemostat culture. Amylase production was growth-limited and restricted to the exponential phase in batch culture. The enzyme yield was reduced by 40% when the culture pH was maintained at pH 7.2. Amylase production in chemostat culture was influenced by the growth rate throughout the dilution rate range used.

Davis, P.E.; Cohen, D.L.; Whitaker, A.

1980-01-01

 
 
 
 
161

Modular system for assessment of glycosyl hydrolase secretion in Geobacillus thermoglucosidasius.  

UK PubMed Central (United Kingdom)

The facultatively anaerobic, thermophilic bacterium Geobacillus thermoglucosidasius is being developed as an industrial micro-organism for cellulosic bioethanol production. Process improvement would be gained by enhanced secretion of glycosyl hydrolases. Here we report the construction of a modular system for combining promoters, signal peptide encoding regions and glycosyl hydrolase genes to facilitate selection of the optimal combination in G. thermoglucosidasius. Initially, a minimal three-part E. coli-Geobacillus sp. shuttle vector pUCG3.8 was constructed using Gibson isothermal DNA assembly. The three PCR amplicons contained the pMB1 E. coli origin of replication and multiple cloning site (MCS) of pUC18, the Geobacillus sp. origin of replication pBST1 and the thermostable kanamycin nucleotidyltransferase gene (knt), respectively. G. thermoglucosidasius could be transformed with pUCG3.8 at an increased efficiency [2.8×10(5) c.f.u. (µg DNA)(-1)] compared to a previously reported shuttle vector, pUCG18. A modular cassette for the inducible expression and secretion of proteins in G. thermoglucosidasius, designed to allow the simple interchange of parts, was demonstrated using the endoglucanase Cel5A from Thermotoga maritima as a secretion target. Expression of cel5A was placed under the control of a cellobiose-inducible promoter (P?glu) together with a signal peptide encoding sequence from a G. thermoglucosidasius C56-YS93 endo-?-1,4-xylanase. The interchange of parts was demonstrated by exchanging the cel5A gene with the 3' region of a gene with homology to celA from Caldicellulosiruptor saccharolyticus and substituting P?glu for the synthetic, constitutive promoter PUp2n38, which increased Cel5A activity five-fold. Cel5A and CelA activities were detected in culture supernatants indicating successful expression and secretion. N-terminal protein sequencing of Cel5A carrying a C-terminal FLAG epitope confirmed processing of the signal peptide sequence.

Bartosiak-Jentys J; Hussein AH; Lewis CJ; Leak DJ

2013-07-01

162

Effect of ionization and nisin on the Bacillus strains and Salmonella Enteritidis inoculated Stearothermophilus  

International Nuclear Information System (INIS)

[en] The antimicrobial effect of nisin (at 1000UI/g), and irradiation (at 1, 3 and 5kGy), against the growth of Salmonella enteritidis (106 ufc/ml) and Bacillus Stearothermophilus (106 ufc/ml), inoculated in turkey salami, was studied during storage at 4 degree for 21 days. Treatment of turkey salami with nisin at 1000UI/g did not show any antimicrobial activity against S. Enteritidis with 6.7 pour cent and 0.8 pour cent of reduction after 0 and 21 days of storage respectively, and seems to be insufficient to inhibit B. Stearothermophilus with 23 pour cent and 21 pour cent of reduction after 0 and 21 days of storage respectively. Antimicrobial activities of irradiation were better and proportional to irradiation doses; it shows a reduction of 27 pour cent, 55 pour cent and 67 pour cent by D1, D2 and D3 respectively. The combination of nisin with irradiation at 5kGy showed stronger antimicrobial activities than those obtained by its combination with the first and the second irradiation dose.

2010-01-01

163

Use ofBacillus stearothermophilus as a model to study tamoxifen-membrane interactions.  

UK PubMed Central (United Kingdom)

A strain ofBacillus stearothermophilus was used as a model to study the interaction of tamoxifen (TAM) with the cell membrane and the cytostatic antiproliferative effects not related to oestrogen binding. The bacterial growth in the presence of TAM was evaluated turbidimetrically and by viable cell counting. In parallel, partition coefficients of TAM in bacterial polar lipid bilayers were determined. Additionally, studies with fluorescent probes were carried out to investigate TAM effects on the physical state of the membrane lipid bilayer. TAM inhibits growth ofB. stearothermophilus and induces loss of cell viability as a function of concentration and the growth temperature. High partitioning of this drug in the bacterial lipid membranes was observed, reaching maximal values in the temperature range of the phase transition. Fluorescence polarizations of 1,6-diphenyl-1,3,5-hexatriene (DPH) and of its propionic acid derivative (DPH-PA) report significant structural disorder of the lipid bilayer induced by the cytostatic, particularly in the phase transition range. A putative relationship between growth impairment by TAM and the TAM-induced perturbation of the physical behaviour of bacterial membrane lipids is suggested.

Luxo C; Jurado AS; Custo Dio JB; Madeira VM

1996-08-01

164

Refolding of the non-specific neutral protease from Bacillus stearothermophilus proceeds via an autoproteolytically sensitive intermediate  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Abstract A very thermostable variant of the thermolysin-like protease from Bacillus stearothermophilus (G8C/N60C) was previously created by introduction of a disulfide bond into the cysteine-free pseudo-wild type variant (pWT) and thus fixing the unfolding region 56-69. In the present paper,...

165

Structure and dynamics of the anticodon arm binding domain of Bacillus stearothermophilus Tyrosyl-tRNA synthetase.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The structure of a recombinant protein, TyrRS(delta4), corresponding to the anticodon arm binding domain of Bacillus stearothermophilus tyrosyl-tRNA synthetase, has been solved, and its dynamics have been studied by nuclear magnetic resonance (NMR). It is the first structure described for such a dom...

Guijarro, J Iñaki; Pintar, Alessandro; Prochnicka-Chalufour, Ada; Guez, Valérie; Gilquin, Bernard; Bedouelle, Hugues

166

Molecular cloning and mapping of phenol degradation genes from Bacillus stearothermophilus FDTP-3 and their expression in Escherichia coli.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Two genes of the meta pathway of phenol degradation were cloned from a phenol-utilizing strain of Bacillus stearothermophilus and were mapped by subcloning and by use of a Tn5 insertion mutation. They code for phenol hydroxylase and catechol 2,3-dioxygenase, respectively. The gene encoding catechol ...

Dong, F M; Wang, L L; Wang, C M; Cheng, J P; He, Z Q; Sheng, Z J; Shen, R Q

167

Determination of thermobacteriological parameters and size of Bacillus stearothermophilus ATCC 7953 spores Determinação dos parâmetros de destruição térmica e dimensões de esporos de Bacillus stearothermophilus ATCC 7953  

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Full Text Available In order to determine thermobacteriological parameters for B. stearothermophilus spores, they were diluted in a saline solution medium and in ground corn-soybean mix, distributed in TDT tube, and submitted to heat for a specific period of time. The D value (time to reduce 1 log cycle of microbial count under a certain temperature) and z value (variation of temperature to cause 10-fold change in D value) were estimated. To estimate their dimensions, the spores were visualized by using a scanning electron microscope. D121.1 ºC and z values for these spores, as determined in the saline solution, were 8.8 minutes and 12.8 ºC, respectively. D121,1 ºC and z values determined in the corn-soy mix were 14.2 minutes and 23.7 ºC, respectively. The micrographs indicated that the spores have homogeneous shape and size, with length and diameter of 2 and 1 µm, respectively. These results confirm that the spore is highly thermal-resistant, and it is a good biological indicator to evaluate the extrusion process as a feed sterilizer.Este experimento objetivou determinar os parâmetros de destruição térmica de esporos de Bacillus stearothermophilus ATCC 7953 e a estimativa de suas dimensões. Os esporos foram suspensos em solução salina e em mistura de grãos de milho e soja moídos, distribuídos em tubos TDT, e submetidos ao calor por tempo variável, seguido de incubação e contagem direta. Determinou-se o valor D (tempo necessário para redução da viabilidade do micro-organismo em 1 ciclo logarítmico sob determinada temperatura) e o valor z (intervalo de temperatura que ocasiona variação de 10 vezes no valor D). Os esporos suspensos em solução salina foram observados em microscópio eletrônico de varredura, para estimativa das dimensões. Os valores de D121,1 ºC e z para os esporos suspensos em solução salina foram 8,8 minutos e 12,8 ºC, respectivamente. Para aqueles suspensos em mistura milho e soja, D121,1 ºC e z foram 14,2 minutos e 23,7 ºC, respectivamente. As micrografias indicaram que os esporos apresentam-se como bastonetes, homogêneos em forma e dimensão, com comprimento e diâmetro estimados em 2 e 1 µm, respectivamente. Os resultados confirmam a elevada resistência térmica do esporo e indicam que este é um bom indicador biológico para avaliação do processo de extrusão como esterilizante de alimentos.

Marcos Fraiha; Antonio Carlos de Oliveira Ferraz; João Domingos Biagi

2010-01-01

168

Biochemical synthesis of novel, self-assembling glycolipids from ricinoleic acid by a recombinant ?-glucosidase from Geobacillus sp  

UK PubMed Central (United Kingdom)

Purpose of work To explore a novel glycolipid, we performed biochemical reactions using a recombinant ?-glucosidase from Geobacillus sp. which shows excellent transglycosylation reaction to hydroxyl groups in a variety of compounds. Two different glycolipids (GL-1 and GL-2) were prepared from ricinoleic acid using a recombinant ?-glucosidase from Geobacillus sp. The molecular structure of GL-1 was confirmed as 12-O-?-d-glucopyranosyl-9-hexadecenoic acid by 1D and 2D NMR analyses. According to MALDI-TOF/MS, GL-1 and GL-2 showed single major peaks at m/z 483.82 and 645.97, respectively. The peaks corresponded to the [M + Na]? ions of the glycolipids. GL-2 was estimated as 12-O-?-d-glucopyranosyl-(4?-O-?-glucopyranosyl)-9-hexadecenoic acid. Light polarization microscopy revealed that GL-2 easily formed self-assembled vesicles in aqueous solution.

Konishi Masa-aki; Fukuoka Tokuma; Shimane Yasuhiro; Mori Kozue; Nagano Yuriko; Ohta Yukari; Kitamoto Dai; Hatada Yuji

2011-01-01

169

Biochemical synthesis of novel, self-assembling glycolipids from ricinoleic acid by a recombinant ?-glucosidase from Geobacillus sp.  

UK PubMed Central (United Kingdom)

PURPOSE OF WORK: To explore a novel glycolipid, we performed biochemical reactions using a recombinant ?-glucosidase from Geobacillus sp. which shows excellent transglycosylation reaction to hydroxyl groups in a variety of compounds. Two different glycolipids (GL-1 and GL-2) were prepared from ricinoleic acid using a recombinant ?-glucosidase from Geobacillus sp. The molecular structure of GL-1 was confirmed as 12-O-?-D-glucopyranosyl-9-hexadecenoic acid by 1D and 2D NMR analyses. According to MALDI-TOF/MS, GL-1 and GL-2 showed single major peaks at m/z 483.82 and 645.97, respectively. The peaks corresponded to the [M + Na](+) ions of the glycolipids. GL-2 was estimated as 12-O-?-D-glucopyranosyl-(4'-O-?-glucopyranosyl)-9-hexadecenoic acid. Light polarization microscopy revealed that GL-2 easily formed self-assembled vesicles in aqueous solution.

Konishi MA; Fukuoka T; Shimane Y; Mori K; Nagano Y; Ohta Y; Kitamoto D; Hatada Y

2011-01-01

170

BOGUS BACTERIA...  

Science.gov (United States)

Here are some websites to get you started... Just click on the links and start searching! microbe world- bacteria Bacteria Rule Quiz! Bacteria.... Harmful Bacteria Bacteria Museum Bacteria! Microbes- all sorts of info... When you are finished looking at the sites or when you have enough information concerning bacteria, ask Mrs. Deaton for some books that can give you even more DETAIL!!! *Don\\'t forget to keep track of your information on your I-CHARTS... ...

Deaton, Mrs.

2007-01-24

171

Isolation and identification of lipase producing thermophilic Geobacillus sp. SBS-4S: cloning and characterization of the lipase.  

Science.gov (United States)

A thermophilic microorganism, SBS-4S, was isolated from a hot spring located in Gilgit, Northern Areas of Pakistan. It was found to be an aerobic, gram-positive, rod-shaped, thermophilic bacterium that grew on various sugars, carboxylic acids and hydrocarbons at temperatures between 45°C and 75°C. Complete 16S rRNA gene sequence of the microorganism exhibited homology to various species of genus Geobacillus. A highest homology of 99.8% was found with Geobacillus kaustophilus. A partial (0.7 kbp) chaperonin gene sequence also showed a highest homology of 99.4% to that of G. kaustophilus whereas biochemical characteristics of the microorganism were similar to Geobacillus uzenensis. Based on biochemical characterization, 16S rRNA and chaperonin gene sequences, we identified SBS-4S as a strain of genus Geobacillus. Strain SBS-4S produced several extracellular enzymes including amylase, protease and lipase. The lipase encoding gene was cloned, expressed in Escherichia coli and the gene product was characterized. The recombinant lipase was optimally active at 60°C with stability at wide pH range (6-12). The enzyme activity was enhanced remarkably in the presence of Ca(+2). The K(m) and the V(max) for the hydrolysis of p-nitrophenyl acetate were 3.8mM and 2273 ?mol min(-1)mg(-1), respectively. The ability of the recombinant enzyme to be stable at a wide pH range makes it a potential candidate for use in industry. PMID:21185780

Tayyab, Muhammad; Rashid, Naeem; Akhtar, Muhammad

2010-12-24

172

Isolation and identification of lipase producing thermophilic Geobacillus sp. SBS-4S: cloning and characterization of the lipase.  

UK PubMed Central (United Kingdom)

A thermophilic microorganism, SBS-4S, was isolated from a hot spring located in Gilgit, Northern Areas of Pakistan. It was found to be an aerobic, gram-positive, rod-shaped, thermophilic bacterium that grew on various sugars, carboxylic acids and hydrocarbons at temperatures between 45°C and 75°C. Complete 16S rRNA gene sequence of the microorganism exhibited homology to various species of genus Geobacillus. A highest homology of 99.8% was found with Geobacillus kaustophilus. A partial (0.7 kbp) chaperonin gene sequence also showed a highest homology of 99.4% to that of G. kaustophilus whereas biochemical characteristics of the microorganism were similar to Geobacillus uzenensis. Based on biochemical characterization, 16S rRNA and chaperonin gene sequences, we identified SBS-4S as a strain of genus Geobacillus. Strain SBS-4S produced several extracellular enzymes including amylase, protease and lipase. The lipase encoding gene was cloned, expressed in Escherichia coli and the gene product was characterized. The recombinant lipase was optimally active at 60°C with stability at wide pH range (6-12). The enzyme activity was enhanced remarkably in the presence of Ca(+2). The K(m) and the V(max) for the hydrolysis of p-nitrophenyl acetate were 3.8mM and 2273 ?mol min(-1)mg(-1), respectively. The ability of the recombinant enzyme to be stable at a wide pH range makes it a potential candidate for use in industry.

Tayyab M; Rashid N; Akhtar M

2011-03-01

173

Thermal resistance of Bacillus stearothermophilus spores in different heating systems containing some approved food additives.  

Science.gov (United States)

The effects of different heating systems on the heat resistance of Bacillus stearothermophilus spores (ATCC 7953, 12980, 15951 and 15952) were investigated. Spores were heated in distilled water, Sorensen buffer (0.18 mol 1-1), McIlvaine buffer (0.0025-0.18 mol 1-1), and several solutions containing sodium chloride (0.06-12%), sodium nitrite (125 ppm), potassium sorbate (0.1%) and sodium benzoate (0.1%) over a wide range of temperatures (115-140 degrees C). D-values obtained for McIlvaine and Sorensen buffers, at the same molarities, were not significantly different (P > 0.05), but decimal reduction times increased as phosphate concentrations in the solutions decreased. The concentrations, in which statistically significant differences (P 0.05). PMID:8862025

López, M; Mazas, M; González, I; González, J; Bernardo, A

1996-09-01

174

Graphical procedure for comparing thermal death of Bacillus stearothermophilus spores in saturated and superheated steam.  

Science.gov (United States)

The thermal death curve of dried spores of Bacillus stearothermophilus in saturated steam was characterized by three phases: (i) a sharp initial rise in viable count; (ii) a low rate of death which gradually increased; and (iii) logarithmic death at maximal rate. The first phase was a reflection of inadequate heat activation of the spore population. The second and third phases represented the characteristic thermal death curve of the spores in saturated steam. A jacketed steam sterilizer, equipped with a system for initial evacuation of the chamber, was examined for superheat during normal operation. Measurements of spore inactivation and temperature revealed superheat in surface layers of fabrics being processed in steam at 121 C. The high temperature of the fabric surfaces was attributed to absorption of excess heat energy from superheated steam. The superheated steam was produced at the beginning of the normal sterilizing cycle by transfer of heat from the steam-heated jacket to saturated steam entering the vessel. PMID:13988774

SHULL, J J; ERNST, R R

1962-09-01

175

Graphical procedure for comparing thermal death of Bacillus stearothermophilus spores in saturated and superheated steam.  

UK PubMed Central (United Kingdom)

The thermal death curve of dried spores of Bacillus stearothermophilus in saturated steam was characterized by three phases: (i) a sharp initial rise in viable count; (ii) a low rate of death which gradually increased; and (iii) logarithmic death at maximal rate. The first phase was a reflection of inadequate heat activation of the spore population. The second and third phases represented the characteristic thermal death curve of the spores in saturated steam. A jacketed steam sterilizer, equipped with a system for initial evacuation of the chamber, was examined for superheat during normal operation. Measurements of spore inactivation and temperature revealed superheat in surface layers of fabrics being processed in steam at 121 C. The high temperature of the fabric surfaces was attributed to absorption of excess heat energy from superheated steam. The superheated steam was produced at the beginning of the normal sterilizing cycle by transfer of heat from the steam-heated jacket to saturated steam entering the vessel.

SHULL JJ; ERNST RR

1962-09-01

176

Absorption, steady-state fluorescence, fluorescence lifetime, and 2D self-assembly properties of engineered fluorescent S-layer fusion proteins of Geobacillus stearothermophilus NRS 2004/3a.  

Science.gov (United States)

S-layer fusion protein technology was used to design four different fluorescent fusion proteins with three different GFP mutants and the red fluorescent protein mRFP1. Their absorption spectra, steady-state fluorescence, and fluorescence lifetime were investigated as a function of pH. It was found that fluorescence intensities and lifetime of the GFP mutant S-layer fusion proteins decreased about 50% between pH 6 and pH 5. The spectral properties of the red S-layer fusion protein were minimally affected by pH variations. These results were compared with His-tagged reference fluorescent proteins, demonstrating that the S-layer protein did not change the general spectral properties of the whole fusion protein. In addition, the pK(a) values of the fluorescent S-layer fusion proteins were calculated. Finally, it was shown that the S-layer fusion proteins were able to self-assemble forming 2D nanostructures of oblique p2 symmetry with lattice parameters of about a = 11 nm, b = 14 nm, and gamma = 80 degrees . The fluorescence tag did not hinder the natural self-assembly process of the S-layer protein. The combination of the fluorescence properties and the self-assembly ability of the engineered fusion proteins make them a promising tool to generate biomimetic surfaces for future applications in nanobiotechnology at a wide range of pH. PMID:19954211

Kainz, Birgit; Steiner, Kerstin; Möller, Marco; Pum, Dietmar; Schäffer, Christina; Sleytr, Uwe B; Toca-Herrera, José L

2010-01-11

177

Screening and distributing features of bacteria with hydantoinase and carbamoylase.  

UK PubMed Central (United Kingdom)

Hydantoinase and carbamoylase are key biocatalysts for the production of optically pure amino acids from dl-5-substituted hydantoins (SSH). Out of 364 isolated strains with hydantoinase and carbamoylase at 45 degrees C, 24 strains showed efficient hydantoinase and carbamoylase activities. Among them, 17 produced l-amino acids, and 7 produced d-amino acids from both aromatic dl-5-benzylhydantoin and aliphatic dl-5-isopropylhydantoin. Most of the strains that were able to form l-amino acid belonged to genera Bacillus, Geobacillus, Brevibacillus, Aneurinibacillus, Microbacterium, and Kurthia. Phylogenetic relationships were investigated based on 16S rRNA from the hydantoinase-producing bacteria. Distinct tendencies toward certain genera were observed between most of the strains forming l-amino acids and d-amino acids from SSH. The results from this study can be utilized to develop new isolation technology of hydantoinase-producing microorganisms, and to understand metabolism and evolutionary origins of hydantoinase and carbamoylase among different bacteria.

Mei Y; He B; Liu N; Ouyang P

2009-01-01

178

Geobacillus thermodenitrificans YjbH recognizes the C-terminal end of Bacillus subtilis Spx to accelerate Spx proteolysis by ClpXP.  

UK PubMed Central (United Kingdom)

Proteolytic control can govern the levels of specific regulatory factors, such as Spx, a transcriptional regulator of the oxidative stress response in Gram-positive bacteria. Under oxidative stress, Spx concentration is elevated and upregulates transcription of genes that function in the stress response. When stress is alleviated, proteolysis of Spx catalysed by ClpXP reduces Spx concentration. Proteolysis is enhanced by the substrate recognition factor YjbH, which possesses a His-Cys-rich region at its N terminus. However, mutations that generate H12A, C13A, H14A, H16A and C31/34A residue substitutions in the N terminus of Bacillus subtilis YjbH (BsYjbH) do not affect functionality in Spx proteolytic control in vivo and in vitro. Because of difficulties in obtaining soluble BsYjbH, the Geobacillus thermodenitrificans yjbH gene was cloned, which yielded soluble GtYjbH protein. Despite its lack of a His-Cys-rich region, GtYjbH complements a B. subtilis yjbH null mutant, and shows high activity in vitro when combined with ClpXP and Spx in an approximately 30 : 1 (ClpXP/Spx : GtYjbH) molar ratio. In vitro interaction experiments showed that Spx and the protease-resistant Spx(DD) (in which the last two residues of Spx are replaced with two Asp residues) bind to GtYjbH, but deletion of 12 residues from the Spx C terminus (Spx?C) significantly diminished interaction and proteolytic degradation, indicating that the C terminus of Spx is important for YjbH recognition. These experiments also showed that Spx, but not GtYjbH, interacts with ClpX. Kinetic measurements for Spx proteolysis by ClpXP in the presence and absence of GtYjbH suggest that YjbH overcomes non-productive Spx-ClpX interaction, resulting in rapid degradation.

Chan CM; Garg S; Lin AA; Zuber P

2012-05-01

179

Thermophilic fermentation of acetoin and 2,3-butanediol by a novel Geobacillus strain  

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Full Text Available Abstract Background Acetoin and 2,3-butanediol are two important biorefinery platform chemicals. They are currently fermented below 40°C using mesophilic strains, but the processes often suffer from bacterial contamination. Results This work reports the isolation and identification of a novel aerobic Geobacillus strain XT15 capable of producing both of these chemicals under elevated temperatures, thus reducing the risk of bacterial contamination. The optimum growth temperature was found to be between 45 and 55°C and the medium initial pH to be 8.0. In addition to glucose, galactose, mannitol, arabionose, and xylose were all acceptable substrates, enabling the potential use of cellulosic biomass as the feedstock. XT15 preferred organic nitrogen sources including corn steep liquor powder, a cheap by-product from corn wet-milling. At 55°C, 7.7?g/L of acetoin and 14.5?g/L of 2,3-butanediol could be obtained using corn steep liquor powder as a nitrogen source. Thirteen volatile products from the cultivation broth of XT15 were identified by gas chromatography–mass spectrometry. Acetoin, 2,3-butanediol, and their derivatives including a novel metabolite 2,3-dihydroxy-3-methylheptan-4-one, accounted for a total of about 96% of all the volatile products. In contrast, organic acids and other products were minor by-products. ?-Acetolactate decarboxylase and acetoin:2,6-dichlorophenolindophenol oxidoreductase in XT15, the two key enzymes in acetoin metabolic pathway, were found to be both moderately thermophilic with the identical optimum temperature of 45°C. Conclusions Geobacillus sp. XT15 is the first naturally occurring thermophile excreting acetoin and/or 2,3-butanediol. This work has demonstrated the attractive prospect of developing it as an industrial strain in the thermophilic fermentation of acetoin and 2,3-butanediol with improved anti-contamination performance. The novel metabolites and enzymes identified in XT15 also indicated its strong promise as a precious biological resource. Thermophilic fermentation also offers great prospect for improving its yields and efficiencies. This remains a core aim for future work.

Xiao Zijun; Wang Xiangming; Huang Yunling; Huo Fangfang; Zhu Xiankun; Xi Lijun; Lu Jian R

2012-01-01

180

Thermophilic fermentation of acetoin and 2,3-butanediol by a novel Geobacillus strain  

Science.gov (United States)

Background Acetoin and 2,3-butanediol are two important biorefinery platform chemicals. They are currently fermented below 40°C using mesophilic strains, but the processes often suffer from bacterial contamination. Results This work reports the isolation and identification of a novel aerobic Geobacillus strain XT15 capable of producing both of these chemicals under elevated temperatures, thus reducing the risk of bacterial contamination. The optimum growth temperature was found to be between 45 and 55°C and the medium initial pH to be 8.0. In addition to glucose, galactose, mannitol, arabionose, and xylose were all acceptable substrates, enabling the potential use of cellulosic biomass as the feedstock. XT15 preferred organic nitrogen sources including corn steep liquor powder, a cheap by-product from corn wet-milling. At 55°C, 7.7?g/L of acetoin and 14.5?g/L of 2,3-butanediol could be obtained using corn steep liquor powder as a nitrogen source. Thirteen volatile products from the cultivation broth of XT15 were identified by gas chromatography–mass spectrometry. Acetoin, 2,3-butanediol, and their derivatives including a novel metabolite 2,3-dihydroxy-3-methylheptan-4-one, accounted for a total of about 96% of all the volatile products. In contrast, organic acids and other products were minor by-products. ?-Acetolactate decarboxylase and acetoin:2,6-dichlorophenolindophenol oxidoreductase in XT15, the two key enzymes in acetoin metabolic pathway, were found to be both moderately thermophilic with the identical optimum temperature of 45°C. Conclusions Geobacillus sp. XT15 is the first naturally occurring thermophile excreting acetoin and/or 2,3-butanediol. This work has demonstrated the attractive prospect of developing it as an industrial strain in the thermophilic fermentation of acetoin and 2,3-butanediol with improved anti-contamination performance. The novel metabolites and enzymes identified in XT15 also indicated its strong promise as a precious biological resource. Thermophilic fermentation also offers great prospect for improving its yields and efficiencies. This remains a core aim for future work.

2012-01-01

 
 
 
 
181

Effect of acidification and oil on the thermal resistance of Bacillus stearothermophilus spores heated in food substrate.  

UK PubMed Central (United Kingdom)

The effect of the addition of vinegar and/or oil to a food homogenate (tomato sauce, tuna and vegetables) on the thermal resistance of Bacillus stearothermophilus spores was studied. The results indicated that the food substrate without the addition of vinegar and oil and a pH value of 5.28 reduced the thermal resistance of B. stearothermophilus spores compared with that obtained in double-distilled water, (D121 = 1.41 and 3.08 min respectively). The addition of vinegar reduced the pH of the substrate (4.81) and consequently the D values were reduced (D121 = 1.28 min). The addition of soya oil and vinegar to substrate until a pH of 4.81, further reduced the thermal resistance of the spores, giving a D121 value of 0.93 min.

Rodrigo F; Rodrigo C; Fernández PS; Rodrigo M; Martínez A

1999-11-01

182

Production, Partial Characterization and Cloning of Thermostable ?-amylase of a Thermophile Geobacillus thermoleovorans YN  

Directory of Open Access Journals (Sweden)

Full Text Available In a molecular screening program to select a potent thermostable amylase from a previously isolated thermophiles, a locally isolated, thermophilic lipase-producing Geobacillus thermoleovorans YN (accession number AF385083), was shown to secrete a thermostable ?-amylase constitutively. The optimal enzyme activity was measured at 75°C, where 90% of the activity was retained at 80°C after one hour of incubation. A catabolite repression due to the addition of glucose to the basal salt medium was demonstrated, while 4 folds increase in volumetric production was achieved in LB and starch-supplemented basal salt media and presented in SDS-PAGE and zymogram. A blunt end PCR fragment (2146 bp) was amplified from genomic DNA using a designed set of primers and ligated to Bluescript —II KS(+) vector, transformed to E. coli DH5-? competent cells by electroporation and screened on LB-agar plates induced with IPTG. Nucleotide sequencing of selected clone revealed two ORFs, the first was (GTG) with a molecular size 1649 nucleotides encoding 549aa residues of a predicted molecular weight 62.592 kD and the second (ATG) with a molecular size 1613 nucleotides encoding 537aa residues of a predicted molecular weight 61.04 kD.

Mahmoud M. Berekaa; Nadia A. Soliman; Yasser R. Abdel-Fattah

2007-01-01

183

A moderately thermostable alkaline phosphatase from Geobacillus thermodenitrificans T2: cloning, expression and biochemical characterization.  

Science.gov (United States)

A gene-encoding alkaline phosphatase (AP) from thermophilic Geobacillus thermodenitrificans T2, termed Gtd AP, was cloned and sequenced. The deduced Gtd AP protein comprises 424 amino acids and shares a low homology with other known AP (Gtd AP protein, without a predicted signal peptide of 30 amino acids, was successfully overexpressed in E. coli and purified as a hexa-His-tagged fusion protein. The pH and temperature optima for purified enzyme are 9.0 and 65 degrees C, respectively. The enzyme retained a high activity at 45-60 degrees C, while it could be quickly inactivated by a heat treatment at 80 degrees C for 15 min, exhibiting a half-life of 8 min at 70 degrees C. The K(m) and V (max) for pNPP were determined to be 31.5 muM and 430 muM/min at optimal conditions. A divalent cation is essential, with a combination of Mg2+ and Co2+ or Zn2+ preferred. The enzyme was strongly inhibited by 10 mM ethylenediaminetetraacetic acid (EDTA) and vanadate but highly resistant to urea and dithiothreitol. The properties of Gtd AP make it suitable for application in molecular cloning or amplification. PMID:18365148

Zhang, Yong; Ji, Chaoneng; Zhang, Xiaoxiao; Yang, Zhenxing; Peng, Jing; Qiu, Rui; Xie, Yi; Mao, Yumin

2008-03-26

184

Temperature dependence of accuracy of DNA polymerase I from Geobacillus anatolicus.  

Science.gov (United States)

Klenow-like DNA polymerase I fragment from Geobacillus anatolicus (GF) was cloned and purified. The accuracy of GF was measured in vitro at three different temperatures under single turnover conditions as well as using a forward mutation assay. In pre-steady-state kinetic measurements, when temperature was raised from 22 °C to 50 °C, the rate (k(pol)) for cognate dTTP and non-cognate dATP nucleotide incorporations increased six- and four-fold, respectively, whereas the K(d) for both nucleotide incorporations changed only slightly. As a result, the error frequency was remained constant (?4 × 10(-4)) over this temperature range. The accuracy of GF was also measured using a forward mutation assay during a single cycle of DNA synthesis of the lacZ? complementation gene in M13mp2 DNA. In this assay, which scores various types of replication errors, mutant frequency of GF was 5 × 10(-3) at 72 °C which is four-fold higher than that of 37 °C. PMID:22652043

Ca?layan, Melike; Bilgin, Ne?'e

2012-05-28

185

Temperature dependence of accuracy of DNA polymerase I from Geobacillus anatolicus.  

UK PubMed Central (United Kingdom)

Klenow-like DNA polymerase I fragment from Geobacillus anatolicus (GF) was cloned and purified. The accuracy of GF was measured in vitro at three different temperatures under single turnover conditions as well as using a forward mutation assay. In pre-steady-state kinetic measurements, when temperature was raised from 22 °C to 50 °C, the rate (k(pol)) for cognate dTTP and non-cognate dATP nucleotide incorporations increased six- and four-fold, respectively, whereas the K(d) for both nucleotide incorporations changed only slightly. As a result, the error frequency was remained constant (?4 × 10(-4)) over this temperature range. The accuracy of GF was also measured using a forward mutation assay during a single cycle of DNA synthesis of the lacZ? complementation gene in M13mp2 DNA. In this assay, which scores various types of replication errors, mutant frequency of GF was 5 × 10(-3) at 72 °C which is four-fold higher than that of 37 °C.

Ca?layan M; Bilgin N

2012-09-01

186

Structure of a bifunctional alcohol dehydrogenase involved in bioethanol generation in Geobacillus thermoglucosidasius.  

Science.gov (United States)

Bifunctional alcohol/aldehyde dehydrogenase (ADHE) enzymes are found within many fermentative microorganisms. They catalyse the conversion of an acyl-coenzyme A to an alcohol via an aldehyde intermediate; this is coupled to the oxidation of two NADH molecules to maintain the NAD(+) pool during fermentative metabolism. The structure of the alcohol dehydrogenase (ADH) domain of an ADHE protein from the ethanol-producing thermophile Geobacillus thermoglucosidasius has been determined to 2.5?Å resolution. This is the first structure to be reported for such a domain. In silico modelling has been carried out to generate a homology model of the aldehyde dehydrogenase domain, and this was subsequently docked with the ADH-domain structure to model the structure of the complete ADHE protein. This model suggests, for the first time, a structural mechanism for the formation of the large multimeric assemblies or `spirosomes' that are observed for this ADHE protein and which have previously been reported for ADHEs from other organisms. PMID:24100328

Extance, Jonathan; Crennell, Susan J; Eley, Kirstin; Cripps, Roger; Hough, David W; Danson, Michael J

2013-09-20

187

Structure of a bifunctional alcohol dehydrogenase involved in bioethanol generation in Geobacillus thermoglucosidasius.  

UK PubMed Central (United Kingdom)

Bifunctional alcohol/aldehyde dehydrogenase (ADHE) enzymes are found within many fermentative microorganisms. They catalyse the conversion of an acyl-coenzyme A to an alcohol via an aldehyde intermediate; this is coupled to the oxidation of two NADH molecules to maintain the NAD(+) pool during fermentative metabolism. The structure of the alcohol dehydrogenase (ADH) domain of an ADHE protein from the ethanol-producing thermophile Geobacillus thermoglucosidasius has been determined to 2.5?Å resolution. This is the first structure to be reported for such a domain. In silico modelling has been carried out to generate a homology model of the aldehyde dehydrogenase domain, and this was subsequently docked with the ADH-domain structure to model the structure of the complete ADHE protein. This model suggests, for the first time, a structural mechanism for the formation of the large multimeric assemblies or `spirosomes' that are observed for this ADHE protein and which have previously been reported for ADHEs from other organisms.

Extance J; Crennell SJ; Eley K; Cripps R; Hough DW; Danson MJ

2013-10-01

188

Characterization of a recombinant thermostable xylanase from hot spring thermophilic Geobacillus sp. TC-W7.  

UK PubMed Central (United Kingdom)

A xylanase-producing thermophilic strain, Geobacillus sp. TC-W7, was isolated from a hot spring in Yongtai (Fuzhou, China). Subsequently, the xylanase gene that encoded 407 amino acids was cloned and expressed. The recombinant xylanase was purified by GST affinity chromatography and exhibited maximum activity at 75 degrees C and a pH of 8.2. The enzyme was active up to 95 degrees C and showed activity over a wide pH range of 5.2 to 10.2. Additionally, the recombinant xylanase showed high thermostability and pH stability. More than 85% of the enzyme's activity was retained after incubation at 70 degrees C for 90 min at a pH of 8.2. The activity of the recombinant xylanase was enhanced by treatment with 10 mM enzyme inhibitors (DDT, Tween-20, 2-Me, or TritonX-100) and was inhibited by EDTA or PMSF. Its functionality was stable in the presence of Li+, Na+, and K+, but inhibited by Hg2+, Ni2+, Co2+, Cu2+, Zn2+, Pb2+, Fe3+, and Al3+. The functionality of the crude xylanase had similar properties to the recombinant xylanase except for when it was treated with Al2+ or Fe2+. The enzyme might be a promising candidate for various industrial applications such as the biofuel, food, and paper and pulp industries.

Liu B; Zhang N; Zhao C; Lin B; Xie L; Huang Y

2012-10-01

189

Characterization of a recombinant thermostable xylanase from hot spring thermophilic Geobacillus sp. TC-W7.  

Science.gov (United States)

A xylanase-producing thermophilic strain, Geobacillus sp. TC-W7, was isolated from a hot spring in Yongtai (Fuzhou, China). Subsequently, the xylanase gene that encoded 407 amino acids was cloned and expressed. The recombinant xylanase was purified by GST affinity chromatography and exhibited maximum activity at 75 degrees C and a pH of 8.2. The enzyme was active up to 95 degrees C and showed activity over a wide pH range of 5.2 to 10.2. Additionally, the recombinant xylanase showed high thermostability and pH stability. More than 85% of the enzyme's activity was retained after incubation at 70 degrees C for 90 min at a pH of 8.2. The activity of the recombinant xylanase was enhanced by treatment with 10 mM enzyme inhibitors (DDT, Tween-20, 2-Me, or TritonX-100) and was inhibited by EDTA or PMSF. Its functionality was stable in the presence of Li+, Na+, and K+, but inhibited by Hg2+, Ni2+, Co2+, Cu2+, Zn2+, Pb2+, Fe3+, and Al3+. The functionality of the crude xylanase had similar properties to the recombinant xylanase except for when it was treated with Al2+ or Fe2+. The enzyme might be a promising candidate for various industrial applications such as the biofuel, food, and paper and pulp industries. PMID:23075790

Liu, Bin; Zhang, Ningning; Zhao, Chao; Lin, Baixue; Xie, Lianhui; Huang, Yifan

2012-10-01

190

Bacteria Museum  

Science.gov (United States)

Who knew that bacteria had their own virtual museum? Here, visitors will "learn that not all bacteria are harmful, how they are used in industry, that they belong to the oldest living creatures on Earth", and many more interesting facts to discover about the diverse world of bacteria. The "Bacterial Species Files" tab at the top of the page, allows visitors to look up information on 40 different specific bacteria, from Anthrax to Yersinia enterocolitica. The information provided for each bacterium includes photographs, consumer guides, fact sheets, and scientific links. Visitors will find that the "Main Exhibits" tab addresses the basics about bacteria, as well as "Pathogenic Bacteria", "Evolution", "How We Fight Bacteria", and "Food and Water Safety". Visitors will surely enjoy the "Good Bacteria in Food" link found in the Food and Water Safety section, as it explains how some foods benefit from good bacteria, such as Swiss cheese, sausage, sauerkraut, chocolate, and coffee.

191

The S-Layer Proteins of Two Bacillus stearothermophilus Wild-Type Strains Are Bound via Their N-Terminal Region to a Secondary Cell Wall Polymer of Identical Chemical Composition  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Two Bacillus stearothermophilus wild-type strains were investigated regarding a common recognition and binding mechanism between the S-layer protein and the underlying cell envelope layer. The S-layer protein from B. stearothermophilus PV72/p6 has a molecular weight of 130,000 and assembles into a h...

Egelseer, Eva Maria; Leitner, Karl; Jarosch, Marina; Hotzy, Christoph; Zayni, Sonja; Sleytr, Uwe B.; Sára, Margit

192

Gene cloning, DNA sequencing, and expression of thermostable beta-mannanase from Bacillus stearothermophilus.  

Science.gov (United States)

A DNA genomic library constructed from Bacillus stearothermophilus, a gram-positive, facultative thermophilic aerobe that secretes a thermostable beta-mannanase, was screened for mannan hydrolytic activity. Recombinant beta-mannanase activity was detected on the basis of the clearing of halos around Escherichia coli colonies grown on a dye-labelled substrate, Remazol brilliant blue-locust bean gum. The nucleotide sequence of the mannanase gene, manF, corresponded to an open reading frame of 2,085 bp that codes for a 32-amino-acid signal peptide and a mature protein with a molecular mass of 76,089 Da. From sequence analysis, ManF belongs to glycosyl hydrolase family 5 and exhibits higher similarity to eukaryotic than to bacterial mannanases. The manF coding sequence was subcloned into the pH6EX3 expression plasmid and expressed in E. coli as a recombinant fusion protein containing a hexahistidine N-terminal sequence. The fusion protein has thermostability similar to the native enzyme and was purified by Ni2+ affinity chromatography. The values for the kinetic parameters Vmax and Km were 384 U/mg and 2.4 mg/ml, respectively, for the recombinant mannanase and were comparable to those of the native enzyme. PMID:9797302

Ethier, N; Talbot, G; Sygusch, J

1998-11-01

193

Gene cloning, DNA sequencing, and expression of thermostable beta-mannanase from Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

A DNA genomic library constructed from Bacillus stearothermophilus, a gram-positive, facultative thermophilic aerobe that secretes a thermostable beta-mannanase, was screened for mannan hydrolytic activity. Recombinant beta-mannanase activity was detected on the basis of the clearing of halos around Escherichia coli colonies grown on a dye-labelled substrate, Remazol brilliant blue-locust bean gum. The nucleotide sequence of the mannanase gene, manF, corresponded to an open reading frame of 2,085 bp that codes for a 32-amino-acid signal peptide and a mature protein with a molecular mass of 76,089 Da. From sequence analysis, ManF belongs to glycosyl hydrolase family 5 and exhibits higher similarity to eukaryotic than to bacterial mannanases. The manF coding sequence was subcloned into the pH6EX3 expression plasmid and expressed in E. coli as a recombinant fusion protein containing a hexahistidine N-terminal sequence. The fusion protein has thermostability similar to the native enzyme and was purified by Ni2+ affinity chromatography. The values for the kinetic parameters Vmax and Km were 384 U/mg and 2.4 mg/ml, respectively, for the recombinant mannanase and were comparable to those of the native enzyme.

Ethier N; Talbot G; Sygusch J

1998-11-01

194

Gene Cloning, DNA Sequencing, and Expression of Thermostable ?-Mannanase from Bacillus stearothermophilus  

Science.gov (United States)

A DNA genomic library constructed from Bacillus stearothermophilus, a gram-positive, facultative thermophilic aerobe that secretes a thermostable ?-mannanase, was screened for mannan hydrolytic activity. Recombinant ?-mannanase activity was detected on the basis of the clearing of halos around Escherichia coli colonies grown on a dye-labelled substrate, Remazol brilliant blue-locust bean gum. The nucleotide sequence of the mannanase gene, manF, corresponded to an open reading frame of 2,085 bp that codes for a 32-amino-acid signal peptide and a mature protein with a molecular mass of 76,089 Da. From sequence analysis, ManF belongs to glycosyl hydrolase family 5 and exhibits higher similarity to eukaryotic than to bacterial mannanases. The manF coding sequence was subcloned into the pH6EX3 expression plasmid and expressed in E. coli as a recombinant fusion protein containing a hexahistidine N-terminal sequence. The fusion protein has thermostability similar to the native enzyme and was purified by Ni2+ affinity chromatography. The values for the kinetic parameters Vmax and Km were 384 U/mg and 2.4 mg/ml, respectively, for the recombinant mannanase and were comparable to those of the native enzyme.

Ethier, Nathalie; Talbot, Guylaine; Sygusch, Jurgen

1998-01-01

195

Cloning and characterization of the str operon and elongation factor Tu expression in Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

The complete primary structure of the str operon of Bacillus stearothermophilus was determined. It was established that the operon is a five-gene transcriptional unit: 5'-ybxF (unknown function; homology to eukaryotic ribosomal protein L30)-rpsL (S12)-rpsG (S7)-fus (elongation factor G [EF-G])-tuf (elongation factor Tu [EF-Tu])-3'. The main operon promoter (strp) was mapped upstream of ybxF, and its strength was compared with the strength of the tuf-specific promoter (tufp) located in the fus-tuf intergenic region. The strength of the tufp region to initiate transcription is about 20-fold higher than that of the strp region, as determined in chloramphenicol acetyltransferase assays. Deletion mapping experiments revealed that the different strengths of the promoters are the consequence of a combined effect of oppositely acting cis elements, identified upstream of strp (an inhibitory region) and tufp (a stimulatory A/T-rich block). Our results suggest that the oppositely adjusted core promoters significantly contribute to the differential expression of the str operon genes, as monitored by the expression of EF-Tu and EF-G.

Krásný L; Vacík T; Fucík V; Jonák J

2000-11-01

196

A pulse-radiolysis study of the manganese-containing superoxide dismutase from Bacillus stearothermophilus  

International Nuclear Information System (INIS)

The mechanism of catalysis of the manganese-containing superoxide dismutase from Bacillus stearothermophilus has been shown to involve a 'fast cycle' and a 'slow cycle' (McAdam, M.E., Fox, R.A., Lavelle, F., and Fielden, E.M., Biochem. J.; 165:71 (1977)). Further properties of the enzyme are now considered. Pulse-radiolysis studies, under conditions of low substrate concentration to enzyme concentration (i.e. when the fast cycle predominates), showed that enzyme activity decreases as pH increases (6.5 to 10.2). Activity was unaffected by the addition of H2O2 or NaN3 but slightly decreased by KCN. Both H2O2 and the reducing radical anion CO2sup(-.) caused a decrease in A480 of the native enzyme. The rate of the fast catalytic cycle was independent of temperature (5 to 550C), and as temperature increased the slow catalytic cycle became relatively more important. Arrhenius parameters of the rate constants were estimated. The possible identity of the various forms of the enzyme is considered. (author).

1977-01-01

197

Salt-Induced Changes in the Subunit Structure of the Bacillus stearothermophilus Lipoate Acetyltransferase.  

UK PubMed Central (United Kingdom)

The Bacillus stearothermophilus lipoate acetyltransferase (E2), composed of sixty identical, subunits is the core component of the pyruvate dehydrogenase complex (PDC). E2 polypeptide is composed of LD, PSBD, and CD domains. Most studies had focused on a truncated E2 that is deficient in LD and PSBD, because CD mainly contributes to maintaining the multimeric structure. We examined salt-induced changes in E2 without truncation and constructed reaction models. We speculate that in the presence of KCl, E2 is dissociated into a monomer and then assembled into an aggregative complex (CA) and a quasi-stable complex (CQ). CA was larger than CQ, but smaller than intact E2. CA and CQ were dominant complexes at about neutral pH and at basic pH respectively. PDC, in which PSBD is occupied by other components, and a truncated E2 undergo dissociation only. LD-PSBD region besides CD might then contribute to the partial association of dissociated E2.

Shigeoka Y; Fujisawa T; Teshiba S; Fukumori H; Yamamoto K; Banno Y; Aso Y

2013-08-01

198

Thermal resistance of Bacillus stearothermophilus spores in different heating systems containing some approved food additives.  

UK PubMed Central (United Kingdom)

The effects of different heating systems on the heat resistance of Bacillus stearothermophilus spores (ATCC 7953, 12980, 15951 and 15952) were investigated. Spores were heated in distilled water, Sorensen buffer (0.18 mol 1-1), McIlvaine buffer (0.0025-0.18 mol 1-1), and several solutions containing sodium chloride (0.06-12%), sodium nitrite (125 ppm), potassium sorbate (0.1%) and sodium benzoate (0.1%) over a wide range of temperatures (115-140 degrees C). D-values obtained for McIlvaine and Sorensen buffers, at the same molarities, were not significantly different (P > 0.05), but decimal reduction times increased as phosphate concentrations in the solutions decreased. The concentrations, in which statistically significant differences (P < 0.05) were obtained, varied among strains. Among the additives assayed, only sodium chloride reduced heat resistance, being effective at concentrations as low as 0.06%. The z-values calculated in this study ranged from 6.99 to 8.40 with a mean value of 7.60 +/- 0.45. Although z-values observed for salt and buffers (180 mol 1-1) were slightly higher than obtained in the other conditions assayed, the difference was not statistically significant (P > 0.05).

López M; Mazas M; González I; González J; Bernardo A

1996-09-01

199

The effect of lipids on the enzymatic activity of 6-phosphofructo-1-kinase from B. stearothermophilus.  

Science.gov (United States)

6-Phosphofructo-1-kinase (PFK-1), a major regulatory enzyme in the glycolysis pathway, is a cytoplasmic enzyme with complicated allosteric kinetics. Here we investigate the effects of lipids on the activity of PFK from Bacillus stearothermophilus (BsPFK), to determine whether BsPFK shares any of the membrane binding or lipid binding properties reported for some mammalian PFKs. Our results show that large unilamellar vesicles (LUVs) composed of either the phospholipid 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) or of 1:1 (mole ratio) DOPC and the fatty acid, oleic acid (OA), cause a three-fold increase in V(max), depending on the lipid concentration and vesicle composition, but no change in K(m). Further studies show lipids do not reverse the allosteric inhibitory effects of phosphoenolpyruvate (PEP) on BsPFK. SDS/PAGE studies do not show significant binding of the BsPFK tetramer to the surface of the phospholipid vesicles, suggesting that modulation of catalytic activity is due to binding of lipid monomers. By simulating the kinetics of BsPFK interaction with vesicles and lipid monomers we conclude that the change in BsPFK catalytic activity with respect to lipid concentration is consistent with monomer abstraction from vesicles rather than direct uptake of lipid monomers from solution. PMID:21871874

Tsaloglou, Maria-Nefeli; Attard, George S; Dymond, Marcus K

2011-08-17

200

The effect of lipids on the enzymatic activity of 6-phosphofructo-1-kinase from B. stearothermophilus.  

UK PubMed Central (United Kingdom)

6-Phosphofructo-1-kinase (PFK-1), a major regulatory enzyme in the glycolysis pathway, is a cytoplasmic enzyme with complicated allosteric kinetics. Here we investigate the effects of lipids on the activity of PFK from Bacillus stearothermophilus (BsPFK), to determine whether BsPFK shares any of the membrane binding or lipid binding properties reported for some mammalian PFKs. Our results show that large unilamellar vesicles (LUVs) composed of either the phospholipid 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) or of 1:1 (mole ratio) DOPC and the fatty acid, oleic acid (OA), cause a three-fold increase in V(max), depending on the lipid concentration and vesicle composition, but no change in K(m). Further studies show lipids do not reverse the allosteric inhibitory effects of phosphoenolpyruvate (PEP) on BsPFK. SDS/PAGE studies do not show significant binding of the BsPFK tetramer to the surface of the phospholipid vesicles, suggesting that modulation of catalytic activity is due to binding of lipid monomers. By simulating the kinetics of BsPFK interaction with vesicles and lipid monomers we conclude that the change in BsPFK catalytic activity with respect to lipid concentration is consistent with monomer abstraction from vesicles rather than direct uptake of lipid monomers from solution.

Tsaloglou MN; Attard GS; Dymond MK

2011-11-01

 
 
 
 
201

Exploring the unfolding mechanism of ?-glutamyltranspeptidases: the case of the thermophilic enzyme from Geobacillus thermodenitrificans.  

UK PubMed Central (United Kingdom)

?-glutamyltranspeptidases (?-GTs) are ubiquitous enzymes that catalyze the hydrolysis of ?-glutamyl bonds in glutathione and glutamine and the transfer of the released ?-glutamyl group to amino acids or short peptides. These enzymes are generally synthesized as precursor proteins, which undergo an intra-molecular autocatalytic cleavage yielding a large and a small subunit. In this study, circular dichroism and intrinsic fluorescence measurements have been used to investigate the structural features and the temperature- and guanidinium hydrochloride (GdnHCl)-induced unfolding of the mature form of the ?-GT from Geobacillus thermodenitrificans (GthGT) and that of its T353A mutant, which represents a mimic of the precursor protein. Data indicate that a) the mutant and the mature GthGT have a different secondary structure content and a slightly different exposure of hydrophobic regions, b) the thermal unfolding processes of both GthGT forms occur through a three-state model, characterized by a stable intermediate species, whereas chemical denaturations proceed through a single transition, c) both GthGT forms exhibit remarkable stability against temperature, but they do not display a strong resistance to the denaturing action of GdnHCl. These findings suggest that electrostatic interactions significantly contribute to the protein stability and that both the precursor and the mature form of GthGT assume compact and stable conformations to resist to the extreme temperatures where G. thermodenidrificans lives. Owing to its thermostability and unique catalytic properties, GthGT is an excellent candidate to be used as a glutaminase in food industry.

Pica A; Russo Krauss I; Castellano I; Rossi M; La Cara F; Graziano G; Sica F; Merlino A

2012-04-01

202

Thermoactive extracellular proteases of Geobacillus caldoproteolyticus, sp. nov., from sewage sludge.  

UK PubMed Central (United Kingdom)

A proteolytic thermophilic bacterial strain, designated as strain SF03, was isolated from sewage sludge in Singapore. Strain SF03 is a strictly aerobic, Gram stain-positive, catalase-positive, oxidase-positive, and endospore-forming rod. It grows at temperatures ranging from 35 to 65 degrees C, pH ranging from 6.0 to 9.0, and salinities ranging from 0 to 2.5%. Phylogenetic analyses revealed that strain SF03 was most similar to Saccharococcus thermophilus, Geobacillus caldoxylosilyticus, and G. thermoglucosidasius, with 16S rRNA gene sequence identities of 97.6, 97.5 and 97.2%, respectively. Based on taxonomic and 16S rRNA analyses, strain SF03 was named G. caldoproteolyticus sp. nov. Production of extracellular protease from strain SF03 was observed on a basal peptone medium supplemented with different carbon and nitrogen sources. Protease production was repressed by glucose, lactose, and casamino acids but was enhanced by sucrose and NH4Cl. The cell growth and protease production were significantly improved when strain SF03 was cultivated on a 10% skim-milk culture medium, suggesting that the presence of protein induced the synthesis of protease. The protease produced by strain SF03 remained active over a pH range of 6.0-11.0 and a temperature range of 40-90 degrees C, with an optimal pH of 8.0-9.0 and an optimal temperature of 70-80 degrees C, respectively. The protease was stable over the temperature range of 40-70 degrees C and retained 57 and 38% of its activity at 80 and 90 degrees C, respectively, after 1 h.

Chen XG; Stabnikova O; Tay JH; Wang JY; Tay ST

2004-12-01

203

Methanotrophic bacteria.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Methane-utilizing bacteria (methanotrophs) are a diverse group of gram-negative bacteria that are related to other members of the Proteobacteria. These bacteria are classified into three groups based on the pathways used for assimilation of formaldehyde, the major source of cell carbon, and other ph...

Hanson, R S; Hanson, T E

204

Production of l-Ribose from l-Ribulose by a Triple-Site Variant of Mannose-6-Phosphate Isomerase from Geobacillus thermodenitrificans  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A triple-site variant (W17Q N90A L129F) of mannose-6-phosphate isomerase from Geobacillus thermodenitrificans was obtained by combining variants with residue substitutions at different positions after random and site-directed mutagenesis. The specific activity and catalytic efficiency (kcat/Km) for ...

Lim, Yu-Ri; Yeom, Soo-Jin; Oh, Deok-Kun

205

Transglycosylation reactions of Bacillus stearothermophilus maltogenic amylase with acarbose and various acceptors.  

Science.gov (United States)

It was observed that Bacillus stearothermophilus maltogenic amylase cleaved the first glycosidic bond of acarbose to produce glucose and a pseudotrisaccharide (PTS) that was transferred to C-6 of the glucose to give an alpha-(1-->6) glycosidic linkage and the formation of isoacarbose. The addition of a number of different carbohydrates to the digest gave transfer products in which PTS was primarily attached alpha-(1-->6) to D-glucose, D-mannose, D-galactose, and methyl alpha-D-glucopyranoside. With D-fructopyranose and D-xylopyranose, PTS was linked alpha-(1-->5) and alpha-(1-->4), respectively. PTS was primarily transferred to C-6 of the nonreducing residue of maltose, cellobiose, lactose, and gentiobiose. Lesser amounts of alpha-(1-->3) and/or alpha-(1-->4) transfer products were also observed for these carbohydrate acceptors. The major transfer product to sucrose gave PTS linked alpha-(1-->4) to the glucose residue. alpha,alpha-Trehalose gave two major products with PTS linked alpha-(1-->6) and alpha-(1-->4). Maltitol gave two major products with PTS linked alpha-(1-->6) and alpha-(1-->4) to the glucopyranose residue. Raffinose gave two major products with PTS linked alpha-(1-->6) and alpha-(1-->4) to the D-galactopyranose residue. Maltotriose gave two major products with PTS linked alpha-(1-->6) and alpha-(1-->4) to the nonreducing end glucopyranose residue. Xylitol gave PTS linked alpha-(1-->5) as the major product and D-glucitol gave PTS linked alpha-(1-->6) as the only product. The structures of the transfer products were determined using thin-layer chromatography, high-performance ion chromatography, enzyme hydrolysis, methylation analysis and 13C NMR spectroscopy. The best acceptor was gentiobiose, followed closely by maltose and cellobiose, and the weakest acceptor was D-glucitol. PMID:10209866

Park, K H; Kim, M J; Lee, H S; Han, N S; Kim, D; Robyt, J F

1998-12-15

206

Transglycosylation reactions of Bacillus stearothermophilus maltogenic amylase with acarbose and various acceptors  

International Nuclear Information System (INIS)

It was observed that Bacillus stearothermophilus maltogenic amylase cleaved the first glycosidic bond of acarbose to produce glucose and a pseudotrisaccharide (PTS) that was transferred to C-6 of the glucose to give an ?-(1-6) glycosidic linkage and the formation of isoacarbose. The addition of a number of different carbohydrates to the digest gave transfer products in which PTS was primarily attached ?-(1-6) to d-glucose, d-mannose, d-galactose, and methyl ?-d-glucopyranoside. With d-fructopyranose and d-xylopyranose, PTS was linked ?-(1-5) and ?-(1-4), respectively. PTS was primarily transferred to C-6 of the nonreducing residue of maltose, cellobiose, lactose, and gentiobiose. Lesser amounts of ?-(1-3) and/or ?-(1-4) transfer products were also observed for these carbohydrate acceptors. The major transfer product to sucrose gave PTS linked ?-(1-4) to the glucose residue. ?,?-Trehalose gave two major products with PTS linked ?-(1-6) and ?-(1-4). Maltitol gave two major products with PTS linked ?-(1-6) and ?-(1-4) to the glucopyranose residue. Raffinose gave two major products with PTS linked ?-(1-6) and ?-(1-4) to the d-galactopyranose residue. Maltotriose gave two major products with PTS linked ?-(1-6) and ?-(1-4) to the nonreducing end glucopyranose residue. Xylitol gave PTS linked ?-(1-5) as the major product and d-glucitol gave PTS linked ?-(1-6) as the only product. The structures of the transfer products were determined using thin layer-chromatography, high-performance ion chromatography, enzyme hydrolysis, methylation analysis and 13C NMR spectroscopy. The best acceptor was gentiobiose, followed closely by maltose and cellobiose, and the weakest acceptor was d-glucitol. (Copyright (c) 1998 Elsevier Science B.V., Amsterdam. All rights reserved.)

1998-12-15

207

Effect on pH heating medium on the thermal resistance of Bacillus stearothermophilus spores.  

Science.gov (United States)

The influence of the pH of heating medium on heat resistance of Bacillus stearothermophilus spores (ATCC 7953, 12980, 15951 and 15952) were studied. The pH values tested were: 4.0, 5.0, 6.0 and 7.0 at temperatures of 115, 120, 125, 130 and 135 degrees C. It was found that at low treatment temperatures (115 degrees C) D-values decreased between 7- and 10-fold with 7953, 12980 and 15951 strains and about 23-fold with 15952 strain when pH dropped from 7.0 to 4.0. At highest treatment temperatures (135 degrees C) D-values obtained with pH 6.0 and 7.0 did not show any significant statistical differences (p > 0.05). z-Values appeared to be higher when the medium was acidified, ranging from 7.58 to 8.20 and 9.43 10.0 for spores suspended in McIlvaine buffer pH 7.0 and pH 4.0, respectively, although the difference was not statistically significant. Heat resistance of strain ATCC 7953 at 120, 128, and 135 degrees C in asparagus purée and tomato purée at pH 5.0 under continuous monitoring of pH was also determined. D-values obtained in asparagus purée were similar to those obtained in buffer at the same pH, whereas those observed in tomato purée were found to be lower. PMID:8652348

López, M; González, I; Condón, S; Bernardo, A

1996-01-01

208

Effect on pH heating medium on the thermal resistance of Bacillus stearothermophilus spores.  

UK PubMed Central (United Kingdom)

The influence of the pH of heating medium on heat resistance of Bacillus stearothermophilus spores (ATCC 7953, 12980, 15951 and 15952) were studied. The pH values tested were: 4.0, 5.0, 6.0 and 7.0 at temperatures of 115, 120, 125, 130 and 135 degrees C. It was found that at low treatment temperatures (115 degrees C) D-values decreased between 7- and 10-fold with 7953, 12980 and 15951 strains and about 23-fold with 15952 strain when pH dropped from 7.0 to 4.0. At highest treatment temperatures (135 degrees C) D-values obtained with pH 6.0 and 7.0 did not show any significant statistical differences (p > 0.05). z-Values appeared to be higher when the medium was acidified, ranging from 7.58 to 8.20 and 9.43 10.0 for spores suspended in McIlvaine buffer pH 7.0 and pH 4.0, respectively, although the difference was not statistically significant. Heat resistance of strain ATCC 7953 at 120, 128, and 135 degrees C in asparagus purée and tomato purée at pH 5.0 under continuous monitoring of pH was also determined. D-values obtained in asparagus purée were similar to those obtained in buffer at the same pH, whereas those observed in tomato purée were found to be lower.

López M; González I; Condón S; Bernardo A

1996-01-01

209

Efficient synthesis and secretion of a thermophilic alpha-amylase by protein-producing Bacillus brevis 47 carrying the Bacillus stearothermophilus amylase gene.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Bacillus subtilis and Bacillus brevis 47-5, carrying the Bacillus stearothermophilus alpha-amylase gene on pUB110 (pBAM101), synthesized the same alpha-amylase as the donor strain as determined by the enzyme's thermal stability and NH2-terminal amino acid sequence. Regardless of the host, the 34-ami...

Tsukagoshi, N; Iritani, S; Sasaki, T; Takemura, T; Ihara, H; Idota, Y; Yamagata, H; Udaka, S

210

Evidence that the N-terminal part of the S-layer protein from Bacillus stearothermophilus PV72/p2 recognizes a secondary cell wall polymer.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The S-layer of Bacillus stearothermophilus PV72/p2 shows oblique lattice symmetry and is composed of identical protein subunits with a molecular weight of 97,000. The isolated S-layer subunits could bind and recrystallize into the oblique lattice on native peptidoglycan-containing sacculi which cons...

Ries, W; Hotzy, C; Schocher, I; Sleytr, U B; Sára, M

211

Influence of the Secondary Cell Wall Polymer on the Reassembly, Recrystallization, and Stability Properties of the S-Layer Protein from Bacillus stearothermophilus PV72/p2  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The high-molecular-weight secondary cell wall polymer (SCWP) from Bacillus stearothermophilus PV72/p2 is mainly composed of N-acetylglucosamine (GlcNAc) and N-acetylmannosamine (ManNAc) and is involved in anchoring the S-layer protein via its N-terminal region to the rigid cell wall layer. In additi...

Sára, Margit; Dekitsch, Christine; Mayer, Harald F.; Egelseer, Eva M.; Sleytr, Uwe B.

212

Identification and characterization of clustered genes for thermostable xylan-degrading enzymes, beta-xylosidase and xylanase, of Bacillus stearothermophilus 21.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Bacillus stearothermophilus 21 is a gram-positive, facultative thermophilic aerobe that can utilize xylan as a sole source of carbon. We isolated this strain from soil, purified its extracellular xylanase and beta-xylosidase, and analyzed the two-step degradation of xylan by these enzymes (T. Nanmor...

Baba, T; Shinke, R; Nanmori, T

213

Crystallization and preliminary X-ray crystallographic analysis of Pz peptidase B from Geobacillus collagenovorans MO-1.  

UK PubMed Central (United Kingdom)

Pz peptidase B is an intracellular M3 metallopeptidase that is found together with Pz peptidase A in the thermophile Geobacillus collagenovorans MO-1 and recognizes collagen-specific tripeptide units (-Gly-Pro-X-). These peptidases have low homology in their primary structures; however, their cleavage patterns towards peptide substrates are similar. In this work, Pz peptidase B was crystallized using the counter-diffusion method. Data were collected to a resolution of 1.6?Å at 100?K from a crystal obtained in the Japanese Experiment Module (JEM; also known as `Kibo') at the International Space Station (ISS). The crystal belonged to the trigonal space group P3(1)21, with unit-cell parameters a = b = 87.64, c = 210.5?Å. A complete data set was also obtained from crystals of selenomethionine-substituted protein.

Nakano H; Hosokawa A; Tagawa R; Inaka K; Ohta K; Nakatsu T; Kato H; Watanabe K

2012-07-01

214

Transglycosylation reactions of Bacillus stearothermophilus maltogenic amylase with acarbose and various acceptors  

Energy Technology Data Exchange (ETDEWEB)

It was observed that Bacillus stearothermophilus maltogenic amylase cleaved the first glycosidic bond of acarbose to produce glucose and a pseudotrisaccharide (PTS) that was transferred to C-6 of the glucose to give an {alpha}-(1-6) glycosidic linkage and the formation of isoacarbose. The addition of a number of different carbohydrates to the digest gave transfer products in which PTS was primarily attached {alpha}-(1-6) to d-glucose, d-mannose, d-galactose, and methyl {alpha}-d-glucopyranoside. With d-fructopyranose and d-xylopyranose, PTS was linked {alpha}-(1-5) and {alpha}-(1-4), respectively. PTS was primarily transferred to C-6 of the nonreducing residue of maltose, cellobiose, lactose, and gentiobiose. Lesser amounts of {alpha}-(1-3) and/or {alpha}-(1-4) transfer products were also observed for these carbohydrate acceptors. The major transfer product to sucrose gave PTS linked {alpha}-(1-4) to the glucose residue. {alpha},{alpha}-Trehalose gave two major products with PTS linked {alpha}-(1-6) and {alpha}-(1-4). Maltitol gave two major products with PTS linked {alpha}-(1-6) and {alpha}-(1-4) to the glucopyranose residue. Raffinose gave two major products with PTS linked {alpha}-(1-6) and {alpha}-(1-4) to the d-galactopyranose residue. Maltotriose gave two major products with PTS linked {alpha}-(1-6) and {alpha}-(1-4) to the nonreducing end glucopyranose residue. Xylitol gave PTS linked {alpha}-(1-5) as the major product and d-glucitol gave PTS linked {alpha}-(1-6) as the only product. The structures of the transfer products were determined using thin layer-chromatography, high-performance ion chromatography, enzyme hydrolysis, methylation analysis and {sup 13}C NMR spectroscopy. The best acceptor was gentiobiose, followed closely by maltose and cellobiose, and the weakest acceptor was d-glucitol. (Copyright (c) 1998 Elsevier Science B.V., Amsterdam. All rights reserved.)

Hwa Park, K.; Jeong Kim, M.; Seob Lee, H.; Kim, D. [Department of Food Science and Technology and Research Center for New Bio-Materials in Agriculture, Seoul National University, Suwon (Korea, Republic of); Soo Han, N.; Robyt, J.F. [Laboratory for Carbohydrate Chemistry and Enzymology, Department of Biochemistry and Biophysics, Iowa State University, Ames, IA (United States)

1998-12-15

215

Redefining the Role of the Quaternary Shift in Bacillus stearothermophilus Phosphofructokinase.  

Science.gov (United States)

Bacillus stearothermophilus phosphofructokinase (BsPFK) is a homotetramer that is allosterically inhibited by phosphoenolpyruvate (PEP), which binds along one dimer-dimer interface. The substrate, fructose 6-phosphate (Fru-6-P), binds along the other dimer-dimer interface. Evans et al. observed that the structure with inhibitor (phosphoglycolate) bound, compared to the structure of wild-type BsPFK with substrate and activator bound, exhibits a 7° rotation about the substrate-binding interface, termed the quaternary shift [Schirmer, T., and Evans, P. R. (1990) Nature 343, 140-145]. We report that the variant D12A BsPFK exhibits a 100-fold increase in its binding affinity for PEP, a 50-fold decrease in its binding affinity for Fru-6-P, but an inhibitory coupling comparable to that of the wild type. Crystal structures of the apo and PEP-bound forms of D12A BsPFK have been determined (Protein Data Bank entries 4I36 and 4I7E , respectively), and both indicate a shifted structure similar to the inhibitor-bound structure of the wild type. D12 does not directly bind to either substrate or inhibitor and is located along the substrate-binding interface. A conserved hydrogen bond between D12 and T156 forms across the substrate-binding subunit-subunit interface in the substrate-bound form of BsPFK. The variant T156A BsPFK, when compared to the wild type, shows a 30-fold increase in PEP binding affinity, a 17-fold decrease in Fru-6-P binding affinity, and an estimated coupling that is also approximately equal to that of the wild type. In addition, the T156A BsPFK crystal structure bound to PEP is reported (Protein Data Bank entry 4I4I ), and it exhibits a shifted structure similar to that of D12A BsPFK and the inhibitor-bound structure of the wild type. The results suggest that the main role of the quaternary shift may be to influence ligand binding and not to cause the heterotropic allosteric inhibition per se. PMID:23859543

Mosser, Rockann; Reddy, Manchi C M; Bruning, John B; Sacchettini, James C; Reinhart, Gregory D

2013-07-31

216

Significance of Phe-220 and Gln-221 in the catalytic mechanism of farnesyl diphosphate synthase of Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

Farnesyl diphosphate synthase [EC 2.5.1.10] from Bacillus stearothermophilus was specifically altered at two amino acid residues by using site-directed mutagenesis. The highly conserved Phe and Gln residues at the sequential amino acid positions 220-221 in an upstream part of the putative substrate binding site were replaced with Ala and Glu, respectively. These mutageneses (F220A and Q221E) resulted in 10(-5) and 10(-3) decreases in catalytic activity of farnesyl diphosphate synthesis, respectively. Michaelis constants of the Q221E mutant for the allylic substrates (dimethylallyl- and geranyl diphosphates) increased approximately 25- and 2-folds, respectively, compared to wild type, whereas those for the homoallylic substrate (isopentenyl diphosphate) were not altered much. These results suggest that the Phe-Gln motif is involved not only in the binding of allylic substrates but also in the catalysis by farnesyl diphosphate synthase.

Koyama T; Tajima M; Nishino T; Ogura K

1995-07-01

217

Combined impact of Bacillus stearothermophilus maltogenic alpha-amylase and surfactants on starch pasting and gelation properties.  

UK PubMed Central (United Kingdom)

In baking applications involving starch gelatinisation, surfactants such as sodium stearoyl lactylate (SSL) and monoacylglycerols (MAG) and Bacillus stearothermophilus maltogenic alpha-amylase (BStA) can be used jointly. We here showed that SSL but not MAG delays wheat starch hydrolysis by BStA. The effects were explained in terms of different degrees of adsorption of the surfactants on the starch granule surface, retarded and/or decreased water uptake and delayed availability of gelatinised starch for hydrolysis by BStA. Additional experiments with waxy maize starch led to the conclusion that SSL impacts swelling power and carbohydrate leaching more by covering the starch granule surface than by forming amylose-lipid complexes. SSL postponed starch hydrolysis by BStA, but this did not influence subsequent starch gelation. Finally, when adding SSL or MAG on top of BStA to starch suspensions, the effect of the surfactants on gel strength predominated over that of BStA.

Van Steertegem B; Pareyt B; Brijs K; Delcour JA

2013-08-01

218

A mixed-species microarray for identification of food spoilage bacilli.  

Science.gov (United States)

Failure of food preservation is frequently caused by thermostable spores of members of the Bacillaceae family, which show a wide spectrum of resistance to cleaning and preservation treatments. We constructed and validated a mixed-species genotyping array for 6 Bacillus species, including Bacillus subtilis, Bacillus licheniformis, Bacillus pumilus, Bacillus sporothermodurans, Bacillus cereus and Bacillus coagulans, and 4 Geobacillus species, including Geobacillus stearothermophilus, Geobacillus thermocatenulatus, Geobacillus toebii and Geobacillus sp., in order to track food spoilage isolates from ingredient to product. The discriminating power of the array was evaluated with sets of 42 reference and 20 test strains. Bacterial isolates contain a within-species-conserved core genome comprising 68-88% of the entire genome and a non-conserved accessory genome comprising 7-22%. The majority of the core genome markers do not hybridise between species, thus they allow for efficient discrimination at the species level. The accessory genome array markers provide high-resolution discrimination at the level of individual isolates from a single species. In conclusion, the reported mixed-species microarray contains discriminating markers that allow rapid and cost-effective typing of Bacillus food spoilage bacteria in a wide variety of food products. PMID:21315980

Caspers, Martien P M; Schuren, Frank H J; van Zuijlen, Andre C M; Brul, Stanley; Montijn, Roy C; Abee, Tjakko; Kort, Remco

2010-03-24

219

A mixed-species microarray for identification of food spoilage bacilli.  

UK PubMed Central (United Kingdom)

Failure of food preservation is frequently caused by thermostable spores of members of the Bacillaceae family, which show a wide spectrum of resistance to cleaning and preservation treatments. We constructed and validated a mixed-species genotyping array for 6 Bacillus species, including Bacillus subtilis, Bacillus licheniformis, Bacillus pumilus, Bacillus sporothermodurans, Bacillus cereus and Bacillus coagulans, and 4 Geobacillus species, including Geobacillus stearothermophilus, Geobacillus thermocatenulatus, Geobacillus toebii and Geobacillus sp., in order to track food spoilage isolates from ingredient to product. The discriminating power of the array was evaluated with sets of 42 reference and 20 test strains. Bacterial isolates contain a within-species-conserved core genome comprising 68-88% of the entire genome and a non-conserved accessory genome comprising 7-22%. The majority of the core genome markers do not hybridise between species, thus they allow for efficient discrimination at the species level. The accessory genome array markers provide high-resolution discrimination at the level of individual isolates from a single species. In conclusion, the reported mixed-species microarray contains discriminating markers that allow rapid and cost-effective typing of Bacillus food spoilage bacteria in a wide variety of food products.

Caspers MP; Schuren FH; van Zuijlen AC; Brul S; Montijn RC; Abee T; Kort R

2011-04-01

220

A mixed-species microarray for identification of food spoilage bacilli  

UK PubMed Central (United Kingdom)

Failure of food preservation is frequently caused by thermostable spores of members of the Bacillaceae family, which show a wide spectrum of resistance to cleaning and preservation treatments. We constructed and validated a mixed-species genotyping array for 6 Bacillus species, including Bacillus subtilis, Bacillus licheniformis, Bacillus pumilus, Bacillus sporothermodurans, Bacillus cereus and Bacillus coagulans, and 4 Geobacillus species, including Geobacillus stearothermophilus, Geobacillus thermocatenulatus, Geobacillus toebii and Geobacillus sp., in order to track food spoilage isolates from ingredient to product. The discriminating power of the array was evaluated with sets of 42 reference and 20 test strains. Bacterial isolates contain a within-species-conserved core genome comprising 68–88% of the entire genome and a non-conserved accessory genome comprising 7–22%. The majority of the core genome markers do not hybridise between species, thus they allow for efficient discrimination at the species level. The accessory genome array markers provide high-resolution discrimination at the level of individual isolates from a single species. In conclusion, the reported mixed-species microarray contains discriminating markers that allow rapid and cost-effective typing of Bacillus food spoilage bacteria in a wide variety of food products.

Caspers MPM; Schuren FHJ; van Zuijlen ACM; Brul S; Montijn RC; Abee T; Kort R

2011-04-01

 
 
 
 
221

Key residues in the allosteric transition of Bacillus stearothermophilus pyruvate kinase identified by site-directed mutagenesis.  

UK PubMed Central (United Kingdom)

The structural gene for pyruvate kinase from Bacillus stearothermophilus has been cloned in Escherichia coli and sequenced. The open reading frame from the ATG start codon to the TAG stop codon is 1482 base-pairs and encodes a peptide of relative molecular mass 52,967. In the expression vector pKK223-3, containing the synthetic tac promoter, the gene is overexpressed in E. coli cells to an estimated level of 30% total soluble cell protein. A purification procedure for the overexpressed protein has been established. The construction and characterization of a pair of mutant proteins has given insight into the structural basis of allosteric regulation in the tetrameric enzyme. Substituting tryptophan for tyrosine at position 466 (mutant Trp466-->Tyr) resulted in an activated form of the enzyme, having a reduced K1/2 for the substrate phosphoenolpyruvate. We propose that the characteristics of this mutant might be the result of bulk removal releasing steric inhibition to the formation of an interdomain salt bridge between Asp356 and Arg444. The regulatory behaviour of the double mutant produced by making the additional substitution aspartate for glutamate at position 356 (Trp466-->Tyr/Asp356-->Glu) corroborates this. The position of the salt bridge is such that it might be pivotal to the conformation of a pocket that is proposed to open up when the active R-conformation is adopted. We suggest that the mechanism of activation of B. stearothermophilus pyruvate kinase by ribose-5-phosphate might hinge on an interaction with, or indirectly through, residue Trp466, removing it from the vicinity of the potential salt bridge between Asp356 and Arg444 and thus effecting a closing together of the protein structure concomitant with an opening up of the pocket region.

Walker D; Chia WN; Muirhead H

1992-11-01

222

[Isolation of endophytic bacteria in potato and test of antagonistic action to bacterial ring rot of potato].  

UK PubMed Central (United Kingdom)

In this study, two hundred and forty bacterial strains were isolated from inner tissue of potato tubers collected from DaTong, TaiYuan and Inner Mongolia Autonomous regions. On the basis of antagonistic examination in vitro, fifty and five bacteria strains were characterized for antagonistic bacteria to ring rot of potato. It was 22.9 percentage of all bacteria strains. The biggest radius of suppression circle was 13 mm. Nine strains were chosen for their suppression of bacterial ring rot, blackleg and dry rot of potato. These strains were bacteriologically ideatified. Strain 118 was Pseudomonas fluorescens biovar V. Strain 110 was Bacillus pumilus. Strain 085 was Bacillus stearothermophilus. Strain 069 was Erwinia herbicola. Strain 043 was Xanthomomas fragariae. Strain 116 was Curtobacterium. Strains A-10' and T3 were Bacillus. Strain H1-6 was Pseudomonas fluorescens.

Cui L; Sun Z; Tian HX; Wang LQ; Xu HY; Sun FZ; Yuan J

2002-12-01

223

Isolation and Characterization of Thermophilic Cellulase-Producing Bacteria from Empty Fruit Bunches-Palm Oil Mill Effluent Compost  

Directory of Open Access Journals (Sweden)

Full Text Available Problems statement: Lack of information on locally isolated cellulase-producing bacterium in thermophilic compost using a mixture of Empty Fruit Bunch (EFB) and Palm Oil Mill Effluent (POME) as composting materials. Approach: The isolation of microbes from compost heap was conducted at day 7 of composting process where the mixture of composting materials consisted of 45.8% cellulose, 17.1% hemicellulose and 28.3% lignin content. The temperature, pH and moisture content of the composting pile at day 7 treatment were 58.3, 8.1 and 65.5°C, respectively. The morphological analysis of the isolated microbes was conducted using Scanning Electron Microscope (SEM) and Gram stain method. The congo red test was conducted in order to detect 1% CMC agar degradation activities. Total genomic DNAs were extracted from approximately 1.0 g of mixed compost and amplified by using PCR primers. The PCR product was sequent to identify the nearest relatives of 16S rRNA genes. The localization of bacteria chromosomes was determined by Fluorescence In Situ Hybridization (FISH) analysis. Results: Single isolated bacteria species was successfully isolated from Empty Fruit Bunch (EFB)-Palm Oil Mill Effluent (POME) compost at thermophilic stage. Restriction fragment length polymorphism profiles of the DNAs coding for the 16S rRNAs with the phylogenetic analysis showed that the isolated bacteria from EFB-POME thermophilic compost gave the highest homology (99%) with similarity to Geobacillus pallidus. The strain was spore forming bacteria and able to grow at 60°C with pH 7. Conclusion: Thermophilic bacteria strain, Geobacillus pallidus was successfully isolated from Empty Fruit Bunch (EFB) and Palm Oil Mil Effluent (POME) compost and characterized.

Azhari S. Baharuddin; Mohamad N.A. Razak; Lim S. Hock; Mohd N. Ahmad; Suraini Abd-Aziz; Nor A.A. Rahman; Umi K.M Shah; Mohd A. Hassan; Kenji Sakai; Yoshihito Shirai

2010-01-01

224

Biochemical and structural properties of gamma-glutamyl transpeptidase from Geobacillus thermodenitrificans: an enzyme specialized in hydrolase activity.  

UK PubMed Central (United Kingdom)

Gamma-glutamyltranspeptidases (gamma-GTs) catalyze the transfer of the gamma-glutamyl moiety of glutathione and related gamma-glutamyl amides to water (hydrolysis) or to amino acids and peptides (transpeptidation) and play a key role in glutathione metabolism. Recently, gamma-GTs have been considered attractive pharmaceutical targets for cancer and useful tools to produce gamma-glutamyl compounds. To find out gamma-GTs with special properties we have chosen microorganisms belonging to Geobacillus species which are source of several thermostable enzymes of potential interest for biotechnology. gamma-GT from Geobacillus thermodenitrificans (GthGT) was cloned, expressed in Escherichia coli, purified to homogeneity and characterized. The enzyme, synthesized as a precursor homotetrameric protein of 61-kDa per subunit, undergoes an internal post-translational cleavage of the 61 kDa monomer into 40- and 21-kDa shorter subunits, which are then assembled into an active heterotetramer composed of two 40- and two 21-kDa subunits. The kinetic characterization of the hydrolysis reaction using L-glutamic acid gamma-(4-nitroanilide) as the substrate reveals that the active enzyme has K(m) 7.6 microM and V(max) 0.36 micromol min/mg. The optimum pH and temperature for the hydrolysis activity are 7.8 and 52 degrees C, respectively. GthGT hydrolyses the physiological antioxidant glutathione, suggesting an involvement of the enzyme in the cellular defense mechanism against oxidative stress. Unlike other gamma-GTs, the mutation of the highly conserved catalytic nucleophile, Thr353, abolishes the post-translational cleavage of the pro-enzyme, but does not completely block the hydrolytic action. Furthermore, GthGT does not show any transpeptidase activity, suggesting that the enzyme is a specialized gamma-glutamyl hydrolase. The GthGT homology-model structure reveals peculiar structural features, which should be responsible for the different functional properties of the enzyme and suggests the structural bases of protein thermostability.

Castellano I; Merlino A; Rossi M; La Cara F

2010-05-01

225

[Heat resistance of "Bacillus subtilis" and "Bacillus stearothermophilus" spores in ethylene glycol, propylene glycol and butylene glycol solutions. Criticism of the use of thermodynamic parameters (author's transl)  

UK PubMed Central (United Kingdom)

Increasing concentrations of ethylene glycol (EG), 1,2-propylene glycol (PG) or 2,3-butylene glycol (BG) lower the heat resistance of B. subtilis SJ2 and B. stearothermophilus 1518 spores, and there is a linear relationship between logarithm of decimal reduction time (D) and glycol concentration. D120 degreesc values of B. subtilis spores in 0.02M, pH 7.0 phosphate buffer containing 20 per cent (w/w) EG, PG and BG are respectively 1, 0.7 and 1.1 min compared to 1.5 min in buffer alone. Corresponding values for B. stearothermophilus spores are 2, 2.4 and 3 min compared to 3.2 min. The type of glycol has little effect upon temperature coefficient z for destruction of the B. subtilis spores (average 6.9 degrees C). On the contrary, in the case of B. stearothermophilus, z increases when the number of carbons increases in the glycol molecule (from 7 to 15 degrees). The thermodynamic parameters which characterize the activation of the spore destruction reaction cannot lead to a general conclusion about a possible mechanism of destruction in the presence of chemical compounds belonging to an homologous series: the two behave diversely, and there is no "isokinetic temperature".

Cerf O; L'Haridon R; Hermier J

1975-01-01

226

Gene cloning and characterization of an aldehyde dehydrogenase from long-chain alkane-degrading Geobacillus thermoleovorans B23.  

UK PubMed Central (United Kingdom)

Geobacillus thermoleovorans B23 is capable of degrading long-chain alkanes at 70 degrees C. Bt-aldh, an aldehyde dehydrogenase gene in B23, was located in the upstream region of p21 whose expression level was dramatically increased when alkane degradation was started (Kato et al. 2009, BMC Microbiol 9:60). Like p21, transcription level of Bt-aldh was also increased upon alkane degradation. Bt-Aldh (497 aa, MW = 53,886) was overproduced in Escherichia coli, purified, and characterized biochemically. Bt-Aldh acted as an octamer, required NAD(+) as a coenzyme, and showed high activity against aliphatic long-chain aldehydes such as tetradecanal. The optimum condition for activity was 50-55 degrees C and pH 10.0. The activity was elevated to two- to threefold in the presence of 2 mM Ba(2+), Ca(2+), or Sr(2+), while Mg(2+) and Zn(2+) inhibited the enzyme activity. Bt-Aldh represents thermophilic aldehyde dehydrogenases responsible for degradation of long-chain alkanes.

Kato T; Miyanaga A; Kanaya S; Morikawa M

2010-01-01

227

Characterization of a broad-range aldehyde dehydrogenase involved in alkane degradation in Geobacillus thermodenitrificans NG80-2.  

Science.gov (United States)

An aldehyde dehydrogenase (ALDH) involved in alkane degradation in crude oil-degrading Geobacillus thermodenitrificans NG80-2 was characterized in vitro. The ALDH was expressed heterologously in Escherichia coli and purified as a His-tagged homotetrameric protein with a subunit of 57 kDa based on SDS-PAGE and Native-PAGE analysis. The purified ALDH-oxidized alkyl aldehydes ranging from formaldehyde (C?) to eicosanoic aldehyde (C??) with the highest activity on C?. It also oxidized several aromatic aldehydes including benzaldehyde, phenylacetaldehyde, o-chloro-benzaldehyde and o-phthalaldehyde. The ALDH uses only NAD(+) as the cofactor, and has no reductive activity on acetate or hexadecanoic acid. Therefore, it is an irreversible NAD(+)-dependent aldehyde dehydrogenase. Kinetic parameters, temperature and pH optimum of the enzyme, and effects of metal ions, EDTA and Triton X-100 on the enzyme activity were investigated. Physiological roles of the ALDH for the survival of NG80-2 in oil reservoirs are discussed. PMID:20171064

Li, Xiaomin; Li, Yanxia; Wei, Dongmei; Li, Ping; Wang, Lei; Feng, Lu

2010-02-18

228

Characterization of a broad-range aldehyde dehydrogenase involved in alkane degradation in Geobacillus thermodenitrificans NG80-2.  

UK PubMed Central (United Kingdom)

An aldehyde dehydrogenase (ALDH) involved in alkane degradation in crude oil-degrading Geobacillus thermodenitrificans NG80-2 was characterized in vitro. The ALDH was expressed heterologously in Escherichia coli and purified as a His-tagged homotetrameric protein with a subunit of 57 kDa based on SDS-PAGE and Native-PAGE analysis. The purified ALDH-oxidized alkyl aldehydes ranging from formaldehyde (C?) to eicosanoic aldehyde (C??) with the highest activity on C?. It also oxidized several aromatic aldehydes including benzaldehyde, phenylacetaldehyde, o-chloro-benzaldehyde and o-phthalaldehyde. The ALDH uses only NAD(+) as the cofactor, and has no reductive activity on acetate or hexadecanoic acid. Therefore, it is an irreversible NAD(+)-dependent aldehyde dehydrogenase. Kinetic parameters, temperature and pH optimum of the enzyme, and effects of metal ions, EDTA and Triton X-100 on the enzyme activity were investigated. Physiological roles of the ALDH for the survival of NG80-2 in oil reservoirs are discussed.

Li X; Li Y; Wei D; Li P; Wang L; Feng L

2010-10-01

229

Gold nanoparticles synthesized by Geobacillus sp. strain ID17 a thermophilic bacterium isolated from deception Island, Antarctica.  

UK PubMed Central (United Kingdom)

BACKGROUND: The use of microorganisms in the synthesis of nanoparticles emerges as an eco-friendly and exciting approach, for production of nanoparticles due to its low energy requirement, environmental compatibility, reduced costs of manufacture, scalability, and nanoparticle stabilization compared with the chemical synthesis. RESULTS: The production of gold nanoparticles by the thermophilic bacterium Geobacillus sp. strain ID17 is reported in this study. Cells exposed to Au3+ turned from colourless into an intense purple colour. This change of colour indicates the accumulation of intracellular gold nanoparticles. Elemental analysis of particles composition was verified using TEM and EDX analysis. The intracellular localization and particles size were verified by TEM showing two different types of particles predominant quasi-hexagonal shape with size ranging from 5--50 nm. The most of them (66%) were between 10--20 nm. FT-IR was utilized to characterize the chemical surface of gold nanoparticles. This assay supports the idea of a protein type of compound on the surface of biosynthesized gold nanoparticles. Reductase activity involved in the synthesis of gold nanoparticles has been previously reported to be present in others microorganisms. This reduction using NADH as substrate was tested in ID17. Crude extracts of the microorganism could catalyze the NADH-dependent Au3+ reductions. CONCLUSIONS: Our results strongly suggest that the biosynthesis of gold nanoparticles by ID17 is mediated by enzymes and NADH as a cofactor for this biological transformation.

Correa-Llantén DN; Muñoz-Ibacache SA; Castro ME; Muñoz PA; Blamey JM

2013-08-01

230

Isolation and properties of 6-phosphogluconate dehydrogenase from Escherichia coli. Some comparisons with the thermophilic enzyme from Bacillus stearothermophilus.  

Science.gov (United States)

6-Phosphogluconate dehydrogenase (6-phospho-D-gluconate:NADP oxidoreductase (decarboxylating), EC 1.1.1.44) of Escherichia coli MREp 600 has been isolated with the purpose of carrying out comparative studies with the thermostable enzyme previously isolated from Bacillus stearothermophilus (Veronese, F.M., Boccù, E.,Fontana, A., Benassi,C.A., and Scoffone, E. (1974), Biochim, Biophys. Acta 334, 31). The purified enzyme appeared homogeneous by the criteria of disc gel electrophoresis with and without sodium dodecyl sulfate, ultracentrifugation, and gel filtration. The enzyme has enzymological and physiochemical properties similar to the enzyme isolated from other sources, including B. stearothermophilus. The E. coli enzyme has a mol wt of 100,000 +/- 3000 and is composed of two apparently identical subunits. The amino acid composition of both the mesophilic and thermophilic enzyme has been determined and found to present large similarities. The E. coli enzyme shows a high degree of specificity for nicotinamide adenine dinucleotide (NADP) and it is inhibited by reduced NADP (NADPH). Cysteine residues are involved in the catalytic activity, since on incubation of the enzyme with p-chloromercuribenzoate or 5.5'-dithiobis(2-nitrobenzoic acid) strong inhibition occurs, activity being restored by treatment with excess of beta-mercaptoethanol. The substrate 6-phosphogluconate protects partially the enzyme from inactivation. Both the mesophilic and thermophilic 6-phosphogluconate dehydrogenases are inactivated by Rose Bengal in the presence of light by similar kinetics and protected against photoinactivation by the enzyme substrate. The E. coli enzyme, on the other hand, showed distinct differences in stability against heat and unfolding agents in respect to the B. stearothermophilus enzyme. Heating at 50 degrees C or incubation in 8 M urea results in rapid inactivation. The gross structure of the mesophilic and thermophilic enzyme was very similar as judged by circular dichroic measurements. The far-ultraviolet circular dichroic spectrum had a negative band centered at about 220 nm. In both cases, the fluorescence emission spectrum indicates that the environment of the tryptophan residues is similar, since both enzymes show an emission maximum at 334 nm upon excitation at 295 nm. Circular dichroism measured at various temperatures between 25 and 80 degrees C showed the mesophilic enzyme to be conformationally stable below about 45 degrees C and the thermophilic enzyme below 60 degrees C. The secondary structure of the E. coli enzyme was very sensitive to the denaturing action of urea, since in 8 M urea it rapidly unfolded. Partial renaturation after urea treatment occurred on dilution with buffer or dialysis, as evidenced by spectral properties of the renatured enzyme. The results show that the mesophilic and thermophilic enzymes are very similar and that differences in thermal stability depend on subtle differences in the architectures of the proteins. PMID:786365

Veronese, F M; Boccù, E; Fontana, A

1976-09-01

231

Isolation and properties of 6-phosphogluconate dehydrogenase from Escherichia coli. Some comparisons with the thermophilic enzyme from Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

6-Phosphogluconate dehydrogenase (6-phospho-D-gluconate:NADP oxidoreductase (decarboxylating), EC 1.1.1.44) of Escherichia coli MREp 600 has been isolated with the purpose of carrying out comparative studies with the thermostable enzyme previously isolated from Bacillus stearothermophilus (Veronese, F.M., Boccù, E.,Fontana, A., Benassi,C.A., and Scoffone, E. (1974), Biochim, Biophys. Acta 334, 31). The purified enzyme appeared homogeneous by the criteria of disc gel electrophoresis with and without sodium dodecyl sulfate, ultracentrifugation, and gel filtration. The enzyme has enzymological and physiochemical properties similar to the enzyme isolated from other sources, including B. stearothermophilus. The E. coli enzyme has a mol wt of 100,000 +/- 3000 and is composed of two apparently identical subunits. The amino acid composition of both the mesophilic and thermophilic enzyme has been determined and found to present large similarities. The E. coli enzyme shows a high degree of specificity for nicotinamide adenine dinucleotide (NADP) and it is inhibited by reduced NADP (NADPH). Cysteine residues are involved in the catalytic activity, since on incubation of the enzyme with p-chloromercuribenzoate or 5.5'-dithiobis(2-nitrobenzoic acid) strong inhibition occurs, activity being restored by treatment with excess of beta-mercaptoethanol. The substrate 6-phosphogluconate protects partially the enzyme from inactivation. Both the mesophilic and thermophilic 6-phosphogluconate dehydrogenases are inactivated by Rose Bengal in the presence of light by similar kinetics and protected against photoinactivation by the enzyme substrate. The E. coli enzyme, on the other hand, showed distinct differences in stability against heat and unfolding agents in respect to the B. stearothermophilus enzyme. Heating at 50 degrees C or incubation in 8 M urea results in rapid inactivation. The gross structure of the mesophilic and thermophilic enzyme was very similar as judged by circular dichroic measurements. The far-ultraviolet circular dichroic spectrum had a negative band centered at about 220 nm. In both cases, the fluorescence emission spectrum indicates that the environment of the tryptophan residues is similar, since both enzymes show an emission maximum at 334 nm upon excitation at 295 nm. Circular dichroism measured at various temperatures between 25 and 80 degrees C showed the mesophilic enzyme to be conformationally stable below about 45 degrees C and the thermophilic enzyme below 60 degrees C. The secondary structure of the E. coli enzyme was very sensitive to the denaturing action of urea, since in 8 M urea it rapidly unfolded. Partial renaturation after urea treatment occurred on dilution with buffer or dialysis, as evidenced by spectral properties of the renatured enzyme. The results show that the mesophilic and thermophilic enzymes are very similar and that differences in thermal stability depend on subtle differences in the architectures of the proteins.

Veronese FM; Boccù E; Fontana A

1976-09-01

232

Surface plasmon resonance-enabled antibacterial digital versatile discs  

Science.gov (United States)

We report the achievement of effective sterilization of exemplary bacteria including Escherichia coli and Geobacillus stearothermophilus spores on a digital versatile disc (DVD). The spiral arrangement of aluminum-covered pits generates strong surface plasmon resonance (SPR) absorption of near-infrared light, leading to high surface temperature that could even damage the DVD plastics. Localized protein denaturation and high sterilization efficiency have been demonstrated by using a fluorescence microscope and cell cultures. Numerical simulations have also been conducted to model the SPR properties and the surface temperature distribution of DVDs under laser illumination. The theoretical predictions agree reasonably well with the experimental results.

Dou, Xuan; Chung, Pei-Yu; Jiang, Peng; Dai, Jianli

2012-02-01

233

Geobacillus subterraneus subsp. aromaticivorans subsp. nov., a novel thermophilic and alkaliphilic bacterium isolated from a hot spring in S?rnak, Turkey.  

UK PubMed Central (United Kingdom)

A new thermophilic spore-forming strain Ge1(T) was isolated from the Guclukonak hot spring in S?rnak, Turkey. The strain was identified by using a polyphasic taxonomic approach. Strain Ge1(T) was Gram-positive, spore-forming, alkaliphilic rod-shaped, motile, occurring in pairs or filamentous. Growth was observed between 30 and 65°C (optimum 60°C) and at pH 5.5-10.0 (optimum pH 9.0). It was capable of utilizing starch, growth was observed at 0-3% NaCl (w/v) and was positive for catalase and urease. The major cellular fatty acids were iso-C(15:0) and iso-C(17:0), and the predominant lipoquinone found was menaquinone MK7 type. The DNA G+C content of the genomic DNA of strain Ge1(T) was 52.0%. Comparative 16S rRNA gene sequence studies showed that the isolate belonged to the genus Geobacillus. The DNA-DNA hybridization mean values between the representative strain Ge1(T) and the closely related species G. subterraneus, G. thermodenitrificans, G. thermocatenulatus, G. vulcani and G. thermoleovorans were 69.3%, 57%, 37%, 27% and 26%, respectively. The results of DNA-DNA hybridization, physiological and biochemical tests allowed genotypic and phenotypic differentiation of strain Ge1(T). Based on these results, we propose assigning a novel subspecies of Geobacillus subterraneus, to be named as Geobacillus subterraneus subsp. aromaticivorans subsp. nov. with the type strain Ge1(T) (DSM 23066 (T)= CIP 110341(T)).

Poli A; Guven K; Romano I; Pirinccioglu H; Guven RG; Euzeby JP; Matpan F; Acer O; Orlando P; Nicolaus B

2012-01-01

234

Geobacillus subterraneus subsp. aromaticivorans subsp. nov., a novel thermophilic and alkaliphilic bacterium isolated from a hot spring in S?rnak, Turkey.  

Science.gov (United States)

A new thermophilic spore-forming strain Ge1(T) was isolated from the Guclukonak hot spring in S?rnak, Turkey. The strain was identified by using a polyphasic taxonomic approach. Strain Ge1(T) was Gram-positive, spore-forming, alkaliphilic rod-shaped, motile, occurring in pairs or filamentous. Growth was observed between 30 and 65°C (optimum 60°C) and at pH 5.5-10.0 (optimum pH 9.0). It was capable of utilizing starch, growth was observed at 0-3% NaCl (w/v) and was positive for catalase and urease. The major cellular fatty acids were iso-C(15:0) and iso-C(17:0), and the predominant lipoquinone found was menaquinone MK7 type. The DNA G+C content of the genomic DNA of strain Ge1(T) was 52.0%. Comparative 16S rRNA gene sequence studies showed that the isolate belonged to the genus Geobacillus. The DNA-DNA hybridization mean values between the representative strain Ge1(T) and the closely related species G. subterraneus, G. thermodenitrificans, G. thermocatenulatus, G. vulcani and G. thermoleovorans were 69.3%, 57%, 37%, 27% and 26%, respectively. The results of DNA-DNA hybridization, physiological and biochemical tests allowed genotypic and phenotypic differentiation of strain Ge1(T). Based on these results, we propose assigning a novel subspecies of Geobacillus subterraneus, to be named as Geobacillus subterraneus subsp. aromaticivorans subsp. nov. with the type strain Ge1(T) (DSM 23066 (T)= CIP 110341(T)). PMID:23337579

Poli, Annarita; Guven, Kemal; Romano, Ida; Pirinccioglu, Hamsi; Guven, Reyhan Gul; Euzeby, Jean Paul Marie; Matpan, Fatma; Acer, Omer; Orlando, Pierangelo; Nicolaus, Barbara

2012-01-01

235

Isolation and identification of thermophilic bacteria from sewage sludge compost  

Energy Technology Data Exchange (ETDEWEB)

Seawage sludge containing about 70%( dry basis )of organic matter and 30% of inorganic matter was used for study. Isolation was carried under various combinations of culture conditions; temperatures ( 30, 40, 60, and 80 {degree}C ), different media(for bacreria, for molds, and for actinomycetes ), various pHs( 4, 7, 8, and 10 ), and aerobicity or anaerobicity. Identification of the bacteria was carried out on the basis of their morphology, physical and biochemical tests. 12 strains of the bacteria under aerobic or anaerobic conditions at 60{degree}C has been isolated. Nine strains have been identified as Bacillus stearothermophilus and two strains as Thermus sp. Growth of these bacrteria has occured in the temperature range of 40 to 78{degree}C, with the optimum growth temperature ranging from 60 to 65{degree}C. The isolated bacilli has grown reasonably on seawage sludge in a mixed culture. However Thermus strains were identified only from aquatic area. The anaerobe was not identified. It was concluded that composting of municipal sludge could be hastened by using B.srearothermophilus. 13 refs., 3 tabs.

Fujio, Y.; Kume, S. (Kyushu Univ., Fukuoka (Japan). Faculty of Agriculture)

1991-11-25

236

Enhanced secretion of Bacillus stearothermophilus L1 lipase in Saccharomyces cerevisiae by translational fusion to cellulose-binding domain.  

Science.gov (United States)

The secretion of Bacillus stearothermophilus L1 lipase in Saccharomyces cerevisiae was investigated by employing a fusion partner, a cellulose-binding domain (CBD) from Trichoderma harzianum endoglucanase II (THEG). The CBD was connected to the N-terminal of L1 lipase through an endogenous linker peptide from THEG. The expression cassette for the fusion protein in S. cerevisiae was constructed using the alpha-amylase signal peptide and the galactose-inducible GAL10 promoter. Secretion of CBD-linker-L1 lipase by this fusion construct was dramatically 7-fold enhanced, compared with that of the mature L1 lipase without CBD-fusion. The fusion protein was secreted into the culture medium, reaching levels of approximately 1.3 g/l in high-cell-density fed-batch cultures. Insertion of a KEX2 cleavage site into the junction between CBD-linker and L1 lipase resulted in the same level of enhanced secretion, indicating that the CBD-linker fusion probably plays a critical role in secretion from endoplasmic reticulum to Golgi apparatus. Therefore, the CBD from THEG can be used both as an affinity tag and as a secretion enhancer for the secretory production of heterologous proteins in S. cerevisiae, since in vivo breakage at the linker was almost negligible. PMID:14740195

Ahn, J O; Choi, E S; Lee, H W; Hwang, S H; Kim, C S; Jang, H W; Haam, S J; Jung, J K

2004-01-23

237

Combined impact of Bacillus stearothermophilus maltogenic alpha-amylase and surfactants on starch pasting and gelation properties.  

Science.gov (United States)

In baking applications involving starch gelatinisation, surfactants such as sodium stearoyl lactylate (SSL) and monoacylglycerols (MAG) and Bacillus stearothermophilus maltogenic alpha-amylase (BStA) can be used jointly. We here showed that SSL but not MAG delays wheat starch hydrolysis by BStA. The effects were explained in terms of different degrees of adsorption of the surfactants on the starch granule surface, retarded and/or decreased water uptake and delayed availability of gelatinised starch for hydrolysis by BStA. Additional experiments with waxy maize starch led to the conclusion that SSL impacts swelling power and carbohydrate leaching more by covering the starch granule surface than by forming amylose-lipid complexes. SSL postponed starch hydrolysis by BStA, but this did not influence subsequent starch gelation. Finally, when adding SSL or MAG on top of BStA to starch suspensions, the effect of the surfactants on gel strength predominated over that of BStA. PMID:23561216

Van Steertegem, Bénédicte; Pareyt, Bram; Brijs, Kristof; Delcour, Jan A

2013-02-08

238

Rumen bacteria  

International Nuclear Information System (INIS)

The rumen is the most extensively studied gut community and is characterized by its high population density, wide diversity and complexity of interactions. This complex, mixed microbial culture is comprised of prokaryote organisms including methane-producing archaebacteria, eukaryote organisms, such as ciliate and flagellate protozoa, anaerobic phycomycete fungi and bacteriophage. Bacteria are predominant (up to 1011 viable cells per g comprising 200 species) but a variety of ciliate protozoa occur widely (104-106/g distributed over 25 genera). The anaerobic fungi are also widely distributed (zoospore population densities of 102-104/g distributed over 5 genera). The occurrence of bacteriophage is well documented (107-109 particles/g). This section focuses primarily on the widely used methods for the cultivation and the enumeration of rumen microbes, especially bacteria, which grow under anaerobic conditions. Methods that can be used to measure hydrolytic enzymes (cellulases, xylanases, amylases and proteinases) are also described, along with cell harvesting and fractionation procedures. Brief reference is also made to fungi and protozoa, but detailed explanations for culturing and enumerating these microbes is presented in Chapters 2.4 and 2.5

2005-01-01

239

High growth rate and substrate exhaustion results in rapid cell death and lysis in the thermophilic bacterium Geobacillus thermoleovorans.  

UK PubMed Central (United Kingdom)

Batch cultures of the thermophilic bacterium Geobacillus thermoleovorans T80 attained extremely high-specific glucose utilization rates leading to high specific growth rates, followed by extensive cell death and lysis with the onset of substrate exhaustion. The dramatic decrease in live cell numbers, as determined by flow cytometry, was accompanied by the release of soluble protein. Once the growth phase reached the point of commitment to lysis created by the impending exhaustion of substrate, the addition of extra carbon substrate did not halt the rapid death rate and lysis, although, towards the end of the exponential growth phase, the substrate was utilized producing only a small additional biomass concentration as a result of the net effect of cell growth and death. This lytic phenomenon was observed when a range of different carbon substrates (glucose, pyruvate, acetate, n-hexadecane, nutrient broth), as well as ammonium (the nitrogen source) in the presence of excess carbon source, reached near exhaustion. The rate and extent of cell death and the ensuing lysis depend on the culture growth rate. Cultures batch grown with a lower initial substrate concentration, or at a lower temperature, or at lower dilution rates for continuous-flow cultures, exhibited a lower rate and extent of cell death and lysis. Batch re-culture of the persister cells resulted in a behavior identical to that of the original culture indicating that these cells were not genetically modified. The glucose utilization, cell growth and death rates were mathematically described based on Monod kinetics and estimated values of pertinent biokinetic constants are reported.

Pavlostathis SG; Marchant R; Banat IM; Ternan NG; McMullan G

2006-09-01

240

Characterization of catalytic carboxylate triad in non-replicative DNA polymerase III (pol E) of Geobacillus kaustophilus HTA.  

UK PubMed Central (United Kingdom)

Three aspartic acid residues D378, D380 and D531 form the catalytic carboxylate triad in Geobacillus kaustophilus (Gka) DNA polymerase III ?-subunit homolog, pol E. We cloned, expressed and purified wild type (WT), alanine (D ? A) and glutamate (D ? E) mutant enzymes of D378, D380 and D531. The WT and mutant enzymes were biochemically characterized for DNA binding, dNTP binding and catalytic activity in the presence of two metal ions (Mg2+ and Mn2+). The polymerase activity of all mutant enzymes was lost in the presence Mg2+, whereas D378E and D531E mutant enzymes showed about 35 and 60 percent activity, with Mn2+. D380E mutant enzyme did not show noticeable activity with either metal ions suggesting its absolute requirement in polymerase reaction. Kinetic characterization of individual mutant proteins showed that the template-primer binding affinity (KD.DNA) did not change due to both D ? A or D ? E mutation. The KM.dNTP for D378E and D531E increased by about 10- and 100-fold, compared to WT enzyme implicating the function of these residues in dNTP binding. Based on these results and the analysis of the available crystal structures of the homologous enzyme species in their apo and E.DNA.dNTP ternary complex forms, we conclude that D378 and D531 are mainly responsible for the binding of metal chelated substrate dNTP, while D380 is solely responsible for the chemical step of phosphodiester bond formation.

Sandalli C; Singh K; Modak MJ

2012-01-01

 
 
 
 
241

Analysis of metabolic pathways and fluxes in a newly discovered thermophilic and ethanol-tolerant Geobacillus strain.  

Science.gov (United States)

A recently discovered thermophilic bacterium, Geobacillus thermoglucosidasius M10EXG, ferments a range of C5 (e.g., xylose) and C6 sugars (e.g., glucose) and is tolerant to high ethanol concentrations (10%, v/v). We have investigated the central metabolism of this bacterium using both in vitro enzyme assays and 13C-based flux analysis to provide insights into the physiological properties of this extremophile and explore its metabolism for bio-ethanol or other bioprocess applications. Our findings show that glucose metabolism in G. thermoglucosidasius M10EXG proceeds via glycolysis, the pentose phosphate pathway, and the TCA cycle; the Entner-Doudoroff pathway and transhydrogenase activity were not detected. Anaplerotic reactions (including the glyoxylate shunt, pyruvate carboxylase, and phosphoenolpyruvate carboxykinase) were active, but fluxes through those pathways could not be accurately determined using amino acid labeling. When growth conditions were switched from aerobic to micro-aerobic conditions, fluxes (based on a normalized glucose uptake rate of 100 units (g DCW)(-1) h(-1)) through the TCA cycle and oxidative pentose phosphate pathway were reduced from 64 +/- 3 to 25 +/- 2 and from 30 +/- 2 to 19 +/- 2, respectively. The carbon flux under micro-aerobic growth was directed to ethanol, L-lactate (> 99% optical purity), acetate, and formate. Under fully anerobic conditions, G. thermoglucosidasius M10EXG used a mixed acid fermentation process and exhibited a maximum ethanol yield of 0.38 +/- 0.07 mol mol(-1) glucose. In silico flux balance modeling demonstrates that lactate and acetate production from G. thermoglucosidasius M10EXG reduces the maximum ethanol yield by approximately threefold, thus indicating that both pathways should be modified to maximize ethanol production. PMID:19016470

Tang, Yinjie J; Sapra, Rajat; Joyner, Dominique; Hazen, Terry C; Myers, Samuel; Reichmuth, David; Blanch, Harvey; Keasling, Jay D

2009-04-01

242

Analysis of metabolic pathways and fluxes in a newly discovered thermophilic and ethanol-tolerant Geobacillus strain.  

UK PubMed Central (United Kingdom)

A recently discovered thermophilic bacterium, Geobacillus thermoglucosidasius M10EXG, ferments a range of C5 (e.g., xylose) and C6 sugars (e.g., glucose) and is tolerant to high ethanol concentrations (10%, v/v). We have investigated the central metabolism of this bacterium using both in vitro enzyme assays and 13C-based flux analysis to provide insights into the physiological properties of this extremophile and explore its metabolism for bio-ethanol or other bioprocess applications. Our findings show that glucose metabolism in G. thermoglucosidasius M10EXG proceeds via glycolysis, the pentose phosphate pathway, and the TCA cycle; the Entner-Doudoroff pathway and transhydrogenase activity were not detected. Anaplerotic reactions (including the glyoxylate shunt, pyruvate carboxylase, and phosphoenolpyruvate carboxykinase) were active, but fluxes through those pathways could not be accurately determined using amino acid labeling. When growth conditions were switched from aerobic to micro-aerobic conditions, fluxes (based on a normalized glucose uptake rate of 100 units (g DCW)(-1) h(-1)) through the TCA cycle and oxidative pentose phosphate pathway were reduced from 64 +/- 3 to 25 +/- 2 and from 30 +/- 2 to 19 +/- 2, respectively. The carbon flux under micro-aerobic growth was directed to ethanol, L-lactate (> 99% optical purity), acetate, and formate. Under fully anerobic conditions, G. thermoglucosidasius M10EXG used a mixed acid fermentation process and exhibited a maximum ethanol yield of 0.38 +/- 0.07 mol mol(-1) glucose. In silico flux balance modeling demonstrates that lactate and acetate production from G. thermoglucosidasius M10EXG reduces the maximum ethanol yield by approximately threefold, thus indicating that both pathways should be modified to maximize ethanol production.

Tang YJ; Sapra R; Joyner D; Hazen TC; Myers S; Reichmuth D; Blanch H; Keasling JD

2009-04-01

243

Analysis of Metabolic Pathways and Fluxes in a Newly Discovered Thermophilic and Ethanol-Tolerant Geobacillus Strain  

Energy Technology Data Exchange (ETDEWEB)

A recently discovered thermophilic bacterium, Geobacillus thermoglucosidasius M10EXG, ferments a range of C5 (e.g., xylose) and C6 sugars (e.g., glucose) and istolerant to high ethanol concentrations (10percent, v/v). We have investigated the central metabolism of this bacterium using both in vitro enzyme assays and 13C-based flux analysis to provide insights into the physiological properties of this extremophile and explore its metabolism for bio-ethanol or other bioprocess applications. Our findings show that glucose metabolism in G. thermoglucosidasius M10EXG proceeds via glycolysis, the pentose phosphate pathway, and the TCA cycle; the Entner?Doudoroff pathway and transhydrogenase activity were not detected. Anaplerotic reactions (including the glyoxylate shunt, pyruvate carboxylase, and phosphoenolpyruvate carboxykinase) were active, but fluxes through those pathways could not be accuratelydetermined using amino acid labeling. When growth conditions were switched from aerobic to micro-aerobic conditions, fluxes (based on a normalized glucose uptake rate of 100 units (g DCW)-1 h-1) through the TCA cycle and oxidative pentose phosphate pathway were reduced from 64+-3 to 25+-2 and from 30+-2 to 19+-2, respectively. The carbon flux under micro-aerobic growth was directed formate. Under fully anerobic conditions, G. thermoglucosidasius M10EXG used a mixed acid fermentation process and exhibited a maximum ethanol yield of 0.38+-0.07 mol mol-1 glucose. In silico flux balance modeling demonstrates that lactate and acetate production from G. thermoglucosidasius M10EXG reduces the maximum ethanol yieldby approximately threefold, thus indicating that both pathways should be modified to maximize ethanol production.

Tang, Yinjie J.; Sapra, Rajat; Joyner, Dominique; Hazen, Terry C.; Myers, Samuel; Reichmuth, David; Blanch, Harvey; Keasling, Jay D.

2009-01-20

244

Glutamic acid 160 is the acid-base catalyst of beta-xylosidase from Bacillus stearothermophilus T-6: a family 39 glycoside hydrolase.  

UK PubMed Central (United Kingdom)

A beta-xylosidase from Bacillus stearothermophilus T-6 was cloned, overexpressed in Escherichia coli and purified to homogeneity. Based on sequence alignment, the enzyme belongs to family 39 glycoside hydrolases, which itself forms part of the wider GH-A clan. The conserved Glu160 was proposed as the acid-base catalyst. An E160A mutant was constructed and subjected to steady state and pre-steady state kinetic analysis together with azide rescue and pH activity profiles. The observed results support the assignment of Glu160 as the acid-base catalytic residue.

Bravman T; Mechaly A; Shulami S; Belakhov V; Baasov T; Shoham G; Shoham Y

2001-04-01

245

A novel cytochrome b(o/a)3-type oxidase from Bacillus stearothermophilus catalyzes cytochrome c-551 oxidation.  

UK PubMed Central (United Kingdom)

Gram-positive thermophilic Bacillus species contain cytochrome caa3-type cytochrome c oxidase as their main terminal oxidase in the respiratory chain. To identify alternative oxidases, we isolated several mutants from B. stearothermophilus defective in the caa3-type oxidase activity [Sakamoto, J. et al (1996) FEMS Microbiol. Lett. 143, 151-158]. A novel oxidase was isolated from membrane preparations of one of the mutants, K17. The oxidase was composed of two subunits with molecular masses of 56 and 19 kDa, and contained protoheme IX, heme O, heme A, and Cu in a ratio of 1:0.7:0.2:3. CO difference spectra indicate that the high-spin heme is mainly heme O. These results suggest that the enzyme belongs to the heme-copper oxidase family and is a cytochrome b(o/a)3-type oxidase, whose high-spin heme is mainly heme O and partly heme A. The enzyme oxidized cytochrome c-551, which is a membrane-bound lipoprotein of thermophilic Bacillus. The turnover rate of the activity (Vmax = 190 s[-1]) and its affinity for cytochrome c-551 (Km = 0.15 microM) were much higher than those for yeast and equine heart cytochromes c. The oxidase activity was enhanced by the presence of salts and inhibited by sodium cyanide with a Ki value of 19 microM. The enzyme kinetics suggests that cytochrome c-551 is the natural substrate to this oxidase. Furthermore, the oxidase had similarity to cytochrome ba3-type oxidase from Thermus thermophilus in the subunit composition, partial amino acid sequence, and prosthetic groups, and therefore is suggested to belong to a unique subgroup of the heme-copper oxidase family together with the Thermus enzyme and archaeal oxidases such as Sulfolobus SoxABCD.

Sakamoto J; Handa Y; Sone N

1997-10-01

246

Inactivation factors of spore-forming bacteria using low-pressure microwave plasmas in an N{sub 2} and O{sub 2} gas mixture  

Energy Technology Data Exchange (ETDEWEB)

In this study, we investigated the inactivation characteristics of Geobacillus stearothermophilus spores under different plasma exposure conditions using low-pressure microwave plasma in nitrogen, oxygen and an air-simulated (N{sub 2}:O{sub 2}=4:1) gas mixture. The microwave-excited surface-wave plasma discharges were produced at low pressure by a large volume device. The directly plasma-exposed spores, up to 10{sup 6} populations, were successfully inactivated within 15, 10 and 5 min of surface-wave plasma treatment using nitrogen, oxygen and an air-simulated gas mixture, respectively, as working gases within the temperature of 75 deg. C. The contribution of different inactivation factors was evaluated by placing different filters (e.g. a LiF plate, a quartz plate and a Tyvek (registered) sheet) as indirect exposure of spores to the plasma. It was observed that optical emissions (including vacuum UV (VUV)/UV) play an important role in the inactivation process. To further evaluate the effect of VUV/UV photons, we placed an evacuated isolated chamber, inside which spores were set, into the main plasma chamber. The experimental results show that the inactivation time by VUV/UV photons alone, without working gas in the immediate vicinity of the spores, is longer than that with working gas. This suggests that the VUV/UV emission is responsible not only for direct UV inactivation of spores but also for generation of reactive neutral species by photoexcitation. The scanning electron microscopy images revealed significant changes in the morphology of directly plasma-exposed spores but no change in the spores irradiated by VUV/UV photons only.

Singh, M K; Ogino, A; Nagatsu, M [Graduate School of Science and Technology, Shizuoka University, Johoku 3-5-1, Hamamatsu 432-8561 (Japan)], E-mail: tmnagat@ipc.shizuoka.ac.jp

2009-11-15

247

Inactivation factors of spore-forming bacteria using low-pressure microwave plasmas in an N2 and O2 gas mixture  

Science.gov (United States)

In this study, we investigated the inactivation characteristics of Geobacillus stearothermophilus spores under different plasma exposure conditions using low-pressure microwave plasma in nitrogen, oxygen and an air-simulated (N2:O2=4:1) gas mixture. The microwave-excited surface-wave plasma discharges were produced at low pressure by a large volume device. The directly plasma-exposed spores, up to 106 populations, were successfully inactivated within 15, 10 and 5 min of surface-wave plasma treatment using nitrogen, oxygen and an air-simulated gas mixture, respectively, as working gases within the temperature of 75 °C. The contribution of different inactivation factors was evaluated by placing different filters (e.g. a LiF plate, a quartz plate and a Tyvek® sheet) as indirect exposure of spores to the plasma. It was observed that optical emissions (including vacuum UV (VUV)/UV) play an important role in the inactivation process. To further evaluate the effect of VUV/UV photons, we placed an evacuated isolated chamber, inside which spores were set, into the main plasma chamber. The experimental results show that the inactivation time by VUV/UV photons alone, without working gas in the immediate vicinity of the spores, is longer than that with working gas. This suggests that the VUV/UV emission is responsible not only for direct UV inactivation of spores but also for generation of reactive neutral species by photoexcitation. The scanning electron microscopy images revealed significant changes in the morphology of directly plasma-exposed spores but no change in the spores irradiated by VUV/UV photons only.

Singh, M. K.; Ogino, A.; Nagatsu, M.

2009-11-01

248

Inactivation factors of spore-forming bacteria using low-pressure microwave plasmas in an N2 and O2 gas mixture  

International Nuclear Information System (INIS)

In this study, we investigated the inactivation characteristics of Geobacillus stearothermophilus spores under different plasma exposure conditions using low-pressure microwave plasma in nitrogen, oxygen and an air-simulated (N2:O2=4:1) gas mixture. The microwave-excited surface-wave plasma discharges were produced at low pressure by a large volume device. The directly plasma-exposed spores, up to 106 populations, were successfully inactivated within 15, 10 and 5 min of surface-wave plasma treatment using nitrogen, oxygen and an air-simulated gas mixture, respectively, as working gases within the temperature of 75 deg. C. The contribution of different inactivation factors was evaluated by placing different filters (e.g. a LiF plate, a quartz plate and a Tyvek (registered) sheet) as indirect exposure of spores to the plasma. It was observed that optical emissions (including vacuum UV (VUV)/UV) play an important role in the inactivation process. To further evaluate the effect of VUV/UV photons, we placed an evacuated isolated chamber, inside which spores were set, into the main plasma chamber. The experimental results show that the inactivation time by VUV/UV photons alone, without working gas in the immediate vicinity of the spores, is longer than that with working gas. This suggests that the VUV/UV emission is responsible not only for direct UV inactivation of spores but also for generation of reactive neutral species by photoexcitation. The scanning electron microscopy images revealed significant changes in the morphology of directly plasma-exposed spores but no change in the spores irradiated by VUV/UV photons only.

2009-01-01

249

Characterization of the Bacillus stearothermophilus manganese superoxide dismutase gene and its ability to complement copper/zinc superoxide dismutase deficiency in Saccharomyces cerevisiae  

Energy Technology Data Exchange (ETDEWEB)

Recombinant clones containing the manganese superoxide dismutase (MnSOD) gene of Bacillus stearothermophilus were isolated with an oligonucleotide probe designed to match a part of the previously determined amino acid sequence. Complementation analyses, performed by introducing each plasmid into a superoxide dismutase-deficient mutant of Escherichia coli, allowed us to define the region of DNA which encodes the MnSOD structural gene and to identify a promoter region immediately upstream from the gene. These data were subsequently confirmed by DNA sequencing. Since MnSOD is normally restricted to the mitochondria in eucaryotes, we were interested (i) in determining whether B. stearothermophilus MnSOD could function in eucaryotic cytosol and (ii) in determining whether MnSOD could replace the structurally unrelated copper/zinc superoxide dismutase (Cu/ZnSOD) which is normally found there. To test this, the sequence encoding bacterial MnSOD was cloned into a yeast expression vector and subsequently introduced into a Cu/ZnSOD-deficient mutant of the yeast Saccharomyces cerevisiae. Functional expression of the protein was demonstrated, and complementation tests revealed that the protein was able to provide tolerance at wild-type levels to conditions which are normally restrictive for this mutant. Thus, in spite of the evolutionary unrelatedness of these two enzymes, Cu/ZnSOD can be functionally replaced by MnSOD in yeast cytosol.

Bowler, C.; Inze, D.; Van Camp, W.; Kaer, L.V.; Dhaese, P. (Rijksuniversiteit Gent (Belgium))

1990-03-01

250

Dynamics in oxygen-induced changes in S-layer protein synthesis from Bacillus stearothermophilus PV72 and the S-layer-deficient variant T5 in continuous culture and studies of the cell wall composition.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Stable synthesis of the hexagonally ordered (p6) S-layer protein from the wild-type strain of Bacillus stearothermophilus PV72 could be achieved in continuous culture on complex medium only under oxygen-limited conditions when glucose was used as the sole carbon source. Depending on the adaptation o...

Sára, M; Kuen, B; Mayer, H F; Mandl, F; Schuster, K C; Sleytr, U B

251

Identification of Two Binding Domains, One for Peptidoglycan and Another for a Secondary Cell Wall Polymer, on the N-Terminal Part of the S-Layer Protein SbsB from Bacillus stearothermophilus PV72/p2  

Digital Repository Infrastructure Vision for European Research (DRIVER)

First studies on the structure-function relationship of the S-layer protein from B. stearothermophilus PV72/p2 revealed the coexistence of two binding domains on its N-terminal part, one for peptidoglycan and another for a secondary cell wall polymer (SCWP). The peptidoglycan binding domain is locat...

Sára, Margit; Egelseer, Eva M.; Dekitsch, Christine; Sleytr, Uwe B.

252

Decontamination effects of low-temperature plasma generated by corona discharge. Part II: new insights.  

UK PubMed Central (United Kingdom)

The second part of our paper presents the results of experiments with the decontamination of surfaces by low-temperature plasma generated by corona discharge in air at atmospheric pressure. A simple device is described and the effects of the corona discharge on model microorganisms, viz. the yeast Candida albicans, Gram-negative bacteria Escherichia coli, Enterobacter aerogenes, Neisseria sicca, Stenotrophomonas maltophilia, Gram-positive bacteria Deinococcus radiodurans, Enterococcus faecium, Staphylococcus epidermidis, Streptococcus sanguinis, and vegetative and spore forms of Geobacillus stearothermophilus are discussed. A similar microbicidal effect after about one-minute exposure was observed in all vegetative forms of the microorganisms. Measurement in growth inhibition zones on a semisolid medium was used to determine the dependence of the microbicidal effect on exposure time and the distance between electrodes. Counting of colonies served to assess the microbicidal effect of the discharge on contaminated inert surfaces observable after more than 1 min exposure. Geobacillus stearothermophilus spores were found to have several times lower susceptibility to the action of the discharge and the microbicidal effect was observed only after an 8 min exposure. Reaction with the iodide reagent did not unambiguously demonstrate the difference between ozone and singlet oxygen as presumed active components of the corona. The area distribution of reactive oxygen species was determined; it was found to differ from the Wartburg law depending on exposure time. Qualitative evidence was obtained on the penetration of the reactive oxygen species into the semisolid medium.

Scholtz V; Julák J; Kríha V; Mosinger J; Kopecká S

2007-01-01

253

Decontamination effects of low-temperature plasma generated by corona discharge. Part II: new insights.  

Science.gov (United States)

The second part of our paper presents the results of experiments with the decontamination of surfaces by low-temperature plasma generated by corona discharge in air at atmospheric pressure. A simple device is described and the effects of the corona discharge on model microorganisms, viz. the yeast Candida albicans, Gram-negative bacteria Escherichia coli, Enterobacter aerogenes, Neisseria sicca, Stenotrophomonas maltophilia, Gram-positive bacteria Deinococcus radiodurans, Enterococcus faecium, Staphylococcus epidermidis, Streptococcus sanguinis, and vegetative and spore forms of Geobacillus stearothermophilus are discussed. A similar microbicidal effect after about one-minute exposure was observed in all vegetative forms of the microorganisms. Measurement in growth inhibition zones on a semisolid medium was used to determine the dependence of the microbicidal effect on exposure time and the distance between electrodes. Counting of colonies served to assess the microbicidal effect of the discharge on contaminated inert surfaces observable after more than 1 min exposure. Geobacillus stearothermophilus spores were found to have several times lower susceptibility to the action of the discharge and the microbicidal effect was observed only after an 8 min exposure. Reaction with the iodide reagent did not unambiguously demonstrate the difference between ozone and singlet oxygen as presumed active components of the corona. The area distribution of reactive oxygen species was determined; it was found to differ from the Wartburg law depending on exposure time. Qualitative evidence was obtained on the penetration of the reactive oxygen species into the semisolid medium. PMID:18225640

Scholtz, V; Julák, J; Kríha, V; Mosinger, J; Kopecká, S

2007-01-01

254

Thermostable and alkalistable endoxylanase of the extremely thermophilic bacterium Geobacillus thermodenitrificans TSAA1: cloning, expression, characteristics and its applicability in generating xylooligosaccharides and fermentable sugars.  

UK PubMed Central (United Kingdom)

Xylanase encoding gene (1,224 bp) from Geobacillus thermodenitrificans was cloned in pET28a (+) vector and successfully expressed in Escherichia coli BL21 (DE3). The deduced amino acid sequence analysis revealed homology with that of glycosyl hydrolase (GH) 10 family with a high molecular mass (50 kDa). The purified recombinant xylanase is optimally active at pH 9.0 and 70 °C with T(1/2) of 10 min at 80 °C, and retains greater than 85 % activity after exposure to 70 °C for 180 min. The enzyme liberates xylose as well as xylooligosaccharides from birchwood xylan and agro-residues, and therefore, this is an endoxylanase. The xylan hydrolytic products (xylooligosaccharides, xylose, and xylobiose) find application as prebiotics and in the production of bioethanol. The xylanase being thermostable and alkalistable, it has released chromophores and phenolics from the residual lignin of pulps, suggesting its utility in mitigating chlorine requirement in pulp bleaching.

Verma D; Anand A; Satyanarayana T

2013-05-01

255

The Museum of Bacteria  

Science.gov (United States)

The Museum of Bacteria serves as a clearinghouse of Web links on bacteria and bacteriology and also provides "crystal-clear information about many aspects of bacteria." The Museum of Bacteria is provided by the Foundation of Bacteria, a non-profit organization dedicated to promoting the field of bacteriology. Links are selected for a general audience, although one section is geared toward professionals in the field. Some of the latest features of the Museum are an "exhibit" on the good bacteria found in food and a Student Hall where students can present their own bacteria-related projects.

256

Translational initiation factor IF2 from Bacillus stearothermophilus: a spectroscopic and microcalorimetric study of the C-domain.  

UK PubMed Central (United Kingdom)

Conformation and stability of the C-terminal domain of initiation factor IF2 from Bacillus stearothermophilus were analyzed by circular dichroism, fluorescence and Raman spectroscopy, and microcalorimetry under different solvent conditions. From circular dichroism and Raman measurements, IF2C at neutral pH can be classified as an alpha + beta protein. Solvent perturbation and Raman spectroscopy indicate a high accessibility of the tyrosine residues in the native protein. The Gdn/HCl-induced unfolding of IF2C was monitored by circular dichroism. IF2C unfolding at neutral pH proceeds in two discrete steps. The midpoints (c(m)) and the free energy of unfolding (deltaG(u)H2O) of the first and second transition are 2.05 M and 6.2 kcal x mol(-1) and 4.1 M and 12.9 kcal x mol(-1), respectively. ANS does not bind to the stable intermediate formed at 3 M Gdn/HCl. It seems likely that IF2C is composed of two subdomains which unfold in a stepwise process. Melting experiments at pH 7.0 are impaired by irreversible aggregation at higher temperatures. However, in Gdn/HCl containing buffer at denaturant concentrations up to 1.5 M the melting becomes a reversible process and can be analyzed by differential scanning calorimetry. At Gdn/HCl concentrations between 1.0 and 1.5 M, IF2C seems to be composed of two folding units with Tm values of about 60 and 78 degrees C and folding enthalpy values (deltaHm) of about 37 and 58 kcal x mol(-1). At pH values below pH 3.0, IF2C can adopt a new acid-induced conformation, which is characterized by a high secondary structure content and a strong ANS binding. The Gdn/HCl-induced unfolding of IF2C at pH 2.6 takes place only in one discrete step with a midpoint c(m) of 3.3 M and a deltaG(AUa)H2O of 11.9 kcal x mol(-1).

Misselwitz R; Welfe K; Krafft C; Gualerzi CO; Welfle H

1997-03-01

257

Thermodynamic analysis of the binding of component enzymes in the assembly of the pyruvate dehydrogenase multienzyme complex of Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

The peripheral subunit-binding domain (PSBD) of the dihydrolipoyl acetyltransferase (E2, EC 2.3.1.12) binds tightly but mutually exclusively to dihydrolipoyl dehydrogenase (E3, EC 1.8.1.4) and pyruvate decarboxylase (E1, EC 1.2.4.1) in the pyruvate dehydrogenase multienzyme complex of Bacillus stearothermophilus. Isothermal titration calorimetry (ITC) experiments demonstrated that the enthalpies of binding (DeltaH degrees ) of both E3 and E1 with the PSBD varied with salt concentration, temperature, pH, and buffer composition. There is little significant difference in the free energies of binding (DeltaG degrees = -12.6 kcal/mol for E3 and = -12.9 kcal/mol for E1 at pH 7.4 and 25 degrees C). However, the association with E3 was characterized by a small, unfavorable enthalpy change (DeltaH degrees = +2.2 kcal/mol) and a large, positive entropy change (TDeltaS degrees = +14.8 kcal/mol), whereas that with E1 was accompanied by a favorable enthalpy change (DeltaH degrees = -8.4 kcal/mol) and a less positive entropy change (TDeltaS degrees = +4.5 kcal/mol). Values of DeltaC(p) of -316 cal/molK and -470 cal/molK were obtained for the binding of E3 and E1, respectively. The value for E3 was not compatible with the DeltaC(p) calculated from the nonpolar surface area buried in the crystal structure of the E3-PSBD complex. In this instance, a large negative DeltaC(p) is not indicative of a classical hydrophobic interaction. In differential scanning calorimetry experiments, the midpoint melting temperature (T(m)) of E3 increased from 91 degrees C to 97.1 degrees C when it was bound to PSBD, and that of E1 increased from 65.2 degrees C to 70.0 degrees C. These high T(m) values eliminate unfolding as a major source of the anomalous DeltaC(p) effects at the temperatures (10-37 degrees C) used for the ITC experiments.

Jung HI; Bowden SJ; Cooper A; Perham RN

2002-05-01

258

Abiotic and microbiotic factors controlling biofilm formation by thermophilic sporeformers.  

UK PubMed Central (United Kingdom)

One of the major concerns in the production of dairy concentrates is the risk of contamination by heat-resistant spores from thermophilic bacteria. In order to acquire more insight in the composition of microbial communities occurring in the dairy concentrate industry, a bar-coded 16S amplicon sequencing analysis was carried out on milk, final products, and fouling samples taken from dairy concentrate production lines. The analysis of these samples revealed the presence of DNA from a broad range of bacterial taxa, including a majority of mesophiles and a minority of (thermophilic) spore-forming bacteria. Enrichments of fouling samples at 55°C showed the accumulation of predominantly Brevibacillus and Bacillus, whereas enrichments at 65°C led to the accumulation of Anoxybacillus and Geobacillus species. Bacterial population analysis of biofilms grown using fouling samples as an inoculum indicated that both Anoxybacillus and Geobacillus preferentially form biofilms on surfaces at air-liquid interfaces rather than on submerged surfaces. Three of the most potent biofilm-forming strains isolated from the dairy factory industrial samples, including Geobacillus thermoglucosidans, Geobacillus stearothermophilus, and Anoxybacillus flavithermus, have been characterized in detail with respect to their growth conditions and spore resistance. Strikingly, Geobacillus thermoglucosidans, which forms the most thermostable spores of these three species, is not able to grow in dairy intermediates as a pure culture but appears to be dependent for growth on other spoilage organisms present, probably as a result of their proteolytic activity. These results underscore the importance of abiotic and microbiotic factors in niche colonization in dairy factories, where the presence of thermophilic sporeformers can affect the quality of end products.

Zhao Y; Caspers MP; Metselaar KI; de Boer P; Roeselers G; Moezelaar R; Nierop Groot M; Montijn RC; Abee T; Kort R

2013-09-01

259

Abiotic and microbiotic factors controlling biofilm formation by thermophilic sporeformers.  

Science.gov (United States)

One of the major concerns in the production of dairy concentrates is the risk of contamination by heat-resistant spores from thermophilic bacteria. In order to acquire more insight in the composition of microbial communities occurring in the dairy concentrate industry, a bar-coded 16S amplicon sequencing analysis was carried out on milk, final products, and fouling samples taken from dairy concentrate production lines. The analysis of these samples revealed the presence of DNA from a broad range of bacterial taxa, including a majority of mesophiles and a minority of (thermophilic) spore-forming bacteria. Enrichments of fouling samples at 55°C showed the accumulation of predominantly Brevibacillus and Bacillus, whereas enrichments at 65°C led to the accumulation of Anoxybacillus and Geobacillus species. Bacterial population analysis of biofilms grown using fouling samples as an inoculum indicated that both Anoxybacillus and Geobacillus preferentially form biofilms on surfaces at air-liquid interfaces rather than on submerged surfaces. Three of the most potent biofilm-forming strains isolated from the dairy factory industrial samples, including Geobacillus thermoglucosidans, Geobacillus stearothermophilus, and Anoxybacillus flavithermus, have been characterized in detail with respect to their growth conditions and spore resistance. Strikingly, Geobacillus thermoglucosidans, which forms the most thermostable spores of these three species, is not able to grow in dairy intermediates as a pure culture but appears to be dependent for growth on other spoilage organisms present, probably as a result of their proteolytic activity. These results underscore the importance of abiotic and microbiotic factors in niche colonization in dairy factories, where the presence of thermophilic sporeformers can affect the quality of end products. PMID:23851093

Zhao, Yu; Caspers, Martien P M; Metselaar, Karin I; de Boer, Paulo; Roeselers, Guus; Moezelaar, Roy; Nierop Groot, Masja; Montijn, Roy C; Abee, Tjakko; Kort, Remco

2013-07-12

260

[Physiological and phylogenetic diversity of thermophilic spore-forming hydrocarbon-oxidizing bacteria from oil fields  

UK PubMed Central (United Kingdom)

The distribution and population density of aerobic hydrocarbon-oxidizing bacteria in the high-temperature oil fields of Western Siberia, Kazakhstan, and China were studied. Seven strains of aerobic thermophilic spore-forming bacteria were isolated from the oil fields and studied by microbiological and molecular biological methods. Based on the 16S rRNA gene sequences, phenotypic characteristics, and the results of DNA-DNA hybridization, the taxonomic affiliation of the isolates was tentatively established. The strains were assigned to the first and fifth subgroups of the genus Bacillus on the phylogenetic branch of the gram-positive bacteria. Strains B and 421 were classified as B. licheniformis. Strains X and U, located between B. stearothermophilus and B. thermocatenulatus on the phylogenetic tree, and strains K, Sam, and 34, related but not identical to B. thermodenitrificans and B. thermoleovorans, undoubtedly represent two new species. Phylogenetically and metabolically related representatives of thermophilic bacilli were found to occur in geographically distant oil fields.

Nazina TN; Turova TP; Poltaraus AB; Novikova EV; Ivanova AE; Grigor'ian AA; Lysenko AM; Beliaev SS

2000-01-01

 
 
 
 
261

[Physiological and phylogenetic diversity of thermophilic spore-forming hydrocarbon-oxidizing bacteria from oil fields].  

Science.gov (United States)

The distribution and population density of aerobic hydrocarbon-oxidizing bacteria in the high-temperature oil fields of Western Siberia, Kazakhstan, and China were studied. Seven strains of aerobic thermophilic spore-forming bacteria were isolated from the oil fields and studied by microbiological and molecular biological methods. Based on the 16S rRNA gene sequences, phenotypic characteristics, and the results of DNA-DNA hybridization, the taxonomic affiliation of the isolates was tentatively established. The strains were assigned to the first and fifth subgroups of the genus Bacillus on the phylogenetic branch of the gram-positive bacteria. Strains B and 421 were classified as B. licheniformis. Strains X and U, located between B. stearothermophilus and B. thermocatenulatus on the phylogenetic tree, and strains K, Sam, and 34, related but not identical to B. thermodenitrificans and B. thermoleovorans, undoubtedly represent two new species. Phylogenetically and metabolically related representatives of thermophilic bacilli were found to occur in geographically distant oil fields. PMID:10808498

Nazina, T N; Turova, T P; Poltaraus, A B; Novikova, E V; Ivanova, A E; Grigor'ian, A A; Lysenko, A M; Beliaev, S S

262

Bacteria isolated from amoebae/bacteria consortium  

Energy Technology Data Exchange (ETDEWEB)

New protozoan derived microbial consortia and method for their isolation are provided. Consortia and bacteria isolated therefrom are useful for treating wastes such as trichloroethylene and trinitrotoluene. Consortia, bacteria isolated therefrom, and dispersants isolated therefrom are useful for dispersing hydrocarbons such as oil, creosote, wax, and grease.

Tyndall, Richard L. (Clinton, TN)

1995-01-01

263

Cellular viability of Saccharomyces cerevisiae cultivated in association with contaminates bacteria of alcoholic fermentation  

International Nuclear Information System (INIS)

The aim of this work was to study the influence of the bacteria Bacillus and Lactobacillus, as well as their metabolic products, in reduction of cellular viability of Saccharomyces cerevisiae, when in mixed culture of yeast and active and treated bacteria. Also was to evaluated an alternative medium (MCC) for the cultivation of bacteria and yeast, constituted of sugarcane juice diluted to 5 deg Brix and supplemented with yeast extract and peptone. The bacteria Bacillus subtilis, Bacillus coagulans, Bacillus stearothermophilus, Lactobacillus fermentum and Lactobacillus plantarum were cultivated in association with yeast Saccharomyces cerevisiae (strain Y-904) for 72 h on 32 deg C, under agitation. The cellular viability, budding rate and population of S. cerevisiae, the total acidity, volatile acidity and pH of culture were determined from 0, 24, 48 e 72 h of mixed culture. Also were determined the initial and final of microorganism population across the pour plate method, in traditional culture medium (PCA for Bacillus, MRS-agar for Lactobacillus and YEPD-agar for yeast S. cerevisiae) and in medium constituted of sugarcane juice. The bacteria cultures were treated by heat sterilization (120 deg C for 20 minutes), antibacterial agent (Kamoran HJ in concentration 3,0 ppm) or irradiation (radiation gamma, with doses of 5,0 kGy for Lactobacillus and 15,0 kGy for Bacillus). The results of the present research showed that just the culture mediums more acids (with higher concentrations of total and volatile acidity, and smaller values of pH), contaminated with active bacteria L. fermentum and B. subtilis, caused reduction on yeast cellular viability. Except the bacteria B. subtilis treated with radiation, the others bacteria treated by different procedures (heat, radiation e antibacterial) did not cause reduction on yeast cellular viability and population, indicating that the isolated presence of the cellular metabolic of theses bacteria was not enough to reduce the percentage of the yeast live cells and a density population. For all microorganisms, the counts obtained with the cultivation medium constituted of sugarcane juice were similar obtained in traditional mediums, probably because the alternative medium simulate the composition of sugarcane must, that the bacteria were isolated in industrial process of ethanol yield. However, the culture medium constituted of sugarcane juice could be replacing traditional culture mediums of yeast and bacteria tested in this work. (author)

2005-01-01

264

Lactic Acid Bacteria  

Science.gov (United States)

This on-line exercise is focused on lactic acid bacteria, a group of related bacteria that produce lactic acid as a result of carbohydrate fermentation. It includes a protocol for the enrichment of lactic acid bacteria from enriched samples (like yogurt, sauerkraut, decaying plant matter, and tooth plaque). Three parameters are measured: growth, culture diversity, and pH. The exercise also includes instructions for the isolation of some of these bacteria by using the streak-plate method.

2010-03-01

265

Evaluation of peracetic acid sanitizers efficiency against spores isolated from spoiled cans in suspension and on stainless steel surfaces.  

UK PubMed Central (United Kingdom)

The aim of this study was to determine the inactivation effect of industrial formulations of peracetic acid biocides on bacterial spores adhering to stainless steel surfaces. A standardized protocol was used to validate biocide activity against spores in suspension. To validate sporicidal activity under practical conditions, we developed an additional protocol to simulate industrial sanitization of stainless steel surfaces with a foam sanitizer. Spores of three spore-forming bacteria, Clostridium sporogenes PA3679, Geobacillus stearothermophilus, and Moorella thermoacetica/thermoautotrophica, were sprayed onto stainless steel as bioaerosols. Sporicidal activity was high against the C. sporogenes spore suspension, with more than 5 log CFU ml(-1) destroyed at all liquid biocide contact times. Sporicidal activity also was high against G. stearothermophilus and M. thermoacetica/thermoautotrophica spores after 30 min of contact, but we found no population reduction at the 5-min contact time for the highest sporicide concentration tested. The foam biocide effectively inactivated C. sporogenes spores adhered to stainless steel but had a reduced decontamination effect on other species. For G. stearothermophilus spores, sanitization with the foam sporicide was more efficient on horizontal steel than on vertical steel, but foam sanitization was ineffective against M. thermoacetica/thermoautotrophica whatever the position. These results highlight that decontamination efficiency may differ depending on whether spores are suspended in an aqueous solution or adhered to a stainless steel surface. Biocide efficiency must be validated using relevant protocols and bacteria representative of the microbiological challenges and issues affecting each food industry.

André S; Hédin S; Remize F; Zuber F

2012-02-01

266

Evaluation of peracetic acid sanitizers efficiency against spores isolated from spoiled cans in suspension and on stainless steel surfaces.  

Science.gov (United States)

The aim of this study was to determine the inactivation effect of industrial formulations of peracetic acid biocides on bacterial spores adhering to stainless steel surfaces. A standardized protocol was used to validate biocide activity against spores in suspension. To validate sporicidal activity under practical conditions, we developed an additional protocol to simulate industrial sanitization of stainless steel surfaces with a foam sanitizer. Spores of three spore-forming bacteria, Clostridium sporogenes PA3679, Geobacillus stearothermophilus, and Moorella thermoacetica/thermoautotrophica, were sprayed onto stainless steel as bioaerosols. Sporicidal activity was high against the C. sporogenes spore suspension, with more than 5 log CFU ml(-1) destroyed at all liquid biocide contact times. Sporicidal activity also was high against G. stearothermophilus and M. thermoacetica/thermoautotrophica spores after 30 min of contact, but we found no population reduction at the 5-min contact time for the highest sporicide concentration tested. The foam biocide effectively inactivated C. sporogenes spores adhered to stainless steel but had a reduced decontamination effect on other species. For G. stearothermophilus spores, sanitization with the foam sporicide was more efficient on horizontal steel than on vertical steel, but foam sanitization was ineffective against M. thermoacetica/thermoautotrophica whatever the position. These results highlight that decontamination efficiency may differ depending on whether spores are suspended in an aqueous solution or adhered to a stainless steel surface. Biocide efficiency must be validated using relevant protocols and bacteria representative of the microbiological challenges and issues affecting each food industry. PMID:22289600

André, S; Hédin, S; Remize, F; Zuber, F

2012-02-01

267

Bacteria are Everywhere!  

Science.gov (United States)

Through this activity, students are introduced to the concept of engineering biological organisms and studying their growth to be able to identify periods of fast and slow growth. Students learn that bacteria are found everywhere, including on the surface of our own hands. Students study three different conditions under which bacteria are found and compare the growth of the individual bacteria from each source. In addition to monitoring the quantity of bacteria from differ conditions, students also record the growth of bacteria over time, which is an excellent tool to study binary fission and the reproduction of unicellular organisms.

Applying Mechatronics to Promote Science (AMPS) GK-12 Program,

268

Indigenous cellulolytic and hemicellulolytic bacteria enhanced rapid co-composting of lignocellulose oil palm empty fruit bunch with palm oil mill effluent anaerobic sludge.  

Science.gov (United States)

The composting of lignocellulosic oil palm empty fruit bunch (OPEFB) with continuous addition of palm oil mill (POME) anaerobic sludge which contained nutrients and indigenous microbes was studied. In comparison to the conventional OPEFB composting which took 60-90days, the rapid composting in this study can be completed in 40days with final C/N ratio of 12.4 and nitrogen (2.5%), phosphorus (1.4%), and potassium (2.8%), respectively. Twenty-seven cellulolytic bacterial strains of which 23 strains were closely related to Bacillus subtilis, Bacillus firmus, Thermobifida fusca, Thermomonospora spp., Cellulomonas sp., Ureibacillus thermosphaericus, Paenibacillus barengoltzii, Paenibacillus campinasensis, Geobacillus thermodenitrificans, Pseudoxanthomonas byssovorax which were known as lignocellulose degrading bacteria and commonly involved in lignocellulose degradation. Four isolated strains related to Exiguobacterium acetylicum and Rhizobium sp., with cellulolytic and hemicellulolytic activities. The rapid composting period achieved in this study can thus be attributed to the naturally occurring cellulolytic and hemicellulolytic strains identified. PMID:24012093

Zainudin, Mohd Huzairi Mohd; Hassan, Mohd Ali; Tokura, Mitsunori; Shirai, Yoshihito

2013-08-14

269

Indigenous cellulolytic and hemicellulolytic bacteria enhanced rapid co-composting of lignocellulose oil palm empty fruit bunch with palm oil mill effluent anaerobic sludge.  

UK PubMed Central (United Kingdom)

The composting of lignocellulosic oil palm empty fruit bunch (OPEFB) with continuous addition of palm oil mill (POME) anaerobic sludge which contained nutrients and indigenous microbes was studied. In comparison to the conventional OPEFB composting which took 60-90days, the rapid composting in this study can be completed in 40days with final C/N ratio of 12.4 and nitrogen (2.5%), phosphorus (1.4%), and potassium (2.8%), respectively. Twenty-seven cellulolytic bacterial strains of which 23 strains were closely related to Bacillus subtilis, Bacillus firmus, Thermobifida fusca, Thermomonospora spp., Cellulomonas sp., Ureibacillus thermosphaericus, Paenibacillus barengoltzii, Paenibacillus campinasensis, Geobacillus thermodenitrificans, Pseudoxanthomonas byssovorax which were known as lignocellulose degrading bacteria and commonly involved in lignocellulose degradation. Four isolated strains related to Exiguobacterium acetylicum and Rhizobium sp., with cellulolytic and hemicellulolytic activities. The rapid composting period achieved in this study can thus be attributed to the naturally occurring cellulolytic and hemicellulolytic strains identified.

Zainudin MH; Hassan MA; Tokura M; Shirai Y

2013-08-01

270

(Genetics of thermophilic bacteria): Progress report, May 1, 1987-December 20, 1987  

Energy Technology Data Exchange (ETDEWEB)

The goal of this project is the genetic analysis of Bacillus stearothermophilus NUB36. Optimal conditions were sought for chromosomal DNA transformation of protoplasts and transduction. Towards this goal a shuttle vector for cloning in B. stearothermophilus NUB36 was constructed. 2 refs.

Welker, N.E.

1987-01-01

271

Darwin y las bacterias Darwin and bacteria  

Directory of Open Access Journals (Sweden)

Full Text Available Con motivo de cumplirse 200 años del natalicio de Darwin y 150 desde la publicación de El Origen de las Especies, se revisa su obra buscando alguna mención de las bacterias, a las cuales el gran naturalista parece, o bien no haber conocido, algo muy difícil en un momento en que causaban sensación en el mundo científico, o bien haber ignorado deliberadamente, porque no encontraba para ellas lugar en su teoría de la evolución. Las bacterias, por su parte, afectaron malamente su vida familiar, falleciendo uno de sus hijos de escarlatina y su hija favorita, Arme, de una tuberculosis agravada por el mismo mal que mató a su hermano. El propio Darwin, desde el regreso del Beagle afectado por una enfermedad crónica hasta ahora no dilucidada, podría haber sufrido de la enfermedad de Chagas, cuyo agente etiológico, si bien no es una bacteria, tiene un similar nivel en la escala evolutiva.As in 2009 the scientific world celebrates two hundreds years from the birthday of Charles Darwin and one hundred and fifty from the publication of The Origin of Species, an analysis of his complete work is performed, looking for any mention of bacteria. But it seems that the great naturahst never took knowledge about its existence, something rather improbable in a time when the discovery of bacteria shook the medical world, or he deliberately ignored them, not finding a place for such microscopic beings into his theory of evolution. But the bacteria badly affected his familiar life, killing scarlet fever one of his children and worsening to death the evolution of tuberculosis of his favourite Annie. Darwin himself could suffer the sickness of Chagas, whose etiological agent has a similar level to bacteria in the scale of evolution.

Walter Ledermann D

2009-01-01

272

Darwin y las bacterias/ Darwin and bacteria  

Scientific Electronic Library Online (English)

Full Text Available Abstract in spanish Con motivo de cumplirse 200 años del natalicio de Darwin y 150 desde la publicación de El Origen de las Especies, se revisa su obra buscando alguna mención de las bacterias, a las cuales el gran naturalista parece, o bien no haber conocido, algo muy difícil en un momento en que causaban sensación en el mundo científico, o bien haber ignorado deliberadamente, porque no encontraba para ellas lugar en su teoría de la evolución. Las bacterias, por su parte, afectaron (more) malamente su vida familiar, falleciendo uno de sus hijos de escarlatina y su hija favorita, Arme, de una tuberculosis agravada por el mismo mal que mató a su hermano. El propio Darwin, desde el regreso del Beagle afectado por una enfermedad crónica hasta ahora no dilucidada, podría haber sufrido de la enfermedad de Chagas, cuyo agente etiológico, si bien no es una bacteria, tiene un similar nivel en la escala evolutiva. Abstract in english As in 2009 the scientific world celebrates two hundreds years from the birthday of Charles Darwin and one hundred and fifty from the publication of The Origin of Species, an analysis of his complete work is performed, looking for any mention of bacteria. But it seems that the great naturahst never took knowledge about its existence, something rather improbable in a time when the discovery of bacteria shook the medical world, or he deliberately ignored them, not finding a (more) place for such microscopic beings into his theory of evolution. But the bacteria badly affected his familiar life, killing scarlet fever one of his children and worsening to death the evolution of tuberculosis of his favourite Annie. Darwin himself could suffer the sickness of Chagas, whose etiological agent has a similar level to bacteria in the scale of evolution.

Ledermann D, Walter

2009-02-01

273

Viabilidade celular de Saccharomyces cerevisiae cultivada em associação com bactérias contaminantes da fermentação alcoólica/ Cellular viability of Saccharomyces cerevisiae cultivated in association with contaminant bacteria of alcoholic fermentation  

Scientific Electronic Library Online (English)

Full Text Available Abstract in portuguese O objetivo deste trabalho foi estudar a influência de bactérias dos gêneros Bacillus e Lactobacillus, bem como de seus produtos metabólicos, na redução da viabilidade celular de leveduras Saccharomyces cerevisiae. As bactérias Bacillus subtilis, Bacillus coagulans, Bacillus stearothermophilus, Lactobacillus fermentum e Lactobacillus plantarum foram cultivadas em associação com a levedura S. cerevisiae (cepa Y-904) por 72 horas a 32 °C, sob agitação. A viabi (more) lidade celular, a taxa de brotamento e a população de células de S. cerevisiae e a acidez total, a acidez volátil e o pH dos meios de cultivos foram determinados às 0, 24, 48 e 72 horas do cultivo misto. As culturas de bactérias foram tratadas através do calor, de agente antimicrobiano e de irradiação. Os resultados mostraram que apenas os meios de cultivo mais acidificados, contaminados com as bactérias ativas L. fermentum e B. subtilis, provocaram redução na viabilidade celular de S. cerevisiae. Excetuando a bactéria B. subtilis tratada com radiação gama, as demais bactérias tratadas pelos diferentes processos (calor, irradiação e antimicrobiano) não causaram diminuição da viabilidade celular e da população de S. cerevisiae, indicando que a presença isolada dos metabólitos celulares dessas bactérias não foi suficiente para reduzir a porcentagem de células vivas de S. cerevisiae. Abstract in english The aim of this project was to study the influence of the bacteria Bacillus and Lactobacillus, as well as their metabolic products to decrease the cellular viability of the yeast Saccharomyces cerevisiae. The bacteria Bacillus subtilis, Bacillus coagulans, Bacillus stearothermophilus, Lactobacillus fermentum and Lactobacillus plantarum were cultivated in association with yeast S. cerevisiae (strain Y-904) for 72 hours at 32 ºC under agitation. The cellular viability, bud (more) ding rate and population of S. Cerevisiae and the total acidity, volatile acidity and pH of culture medium were determined at 0, 24, 48 and 72 hours of incubation of the mixed culture. The bacteria cultures were treated by heat sterilization, antibacterial agent and irradiation. The results showed that only the more acidified culture medium, contaminated with active bacteria L. fermentum and B. subtilis, caused a reduction in the yeast cellular viability. Except for the bacteria B. subtilis treated for radiation, the other bacteria treated by the different procedures (heat, radiation and antibacterial) did not cause a reduction in the cellular viability of S. cerevisiae, indicating that the isolated presence of the cellular metabolic of these bacteria was not enough to reduce the percentage of the living yeast cells.

Nobre, Thais de Paula; Horii, Jorge; Alcarde, André Ricardo

2007-03-01

274

Viabilidade celular de Saccharomyces cerevisiae cultivada em associação com bactérias contaminantes da fermentação alcoólica Cellular viability of Saccharomyces cerevisiae cultivated in association with contaminant bacteria of alcoholic fermentation  

Directory of Open Access Journals (Sweden)

Full Text Available O objetivo deste trabalho foi estudar a influência de bactérias dos gêneros Bacillus e Lactobacillus, bem como de seus produtos metabólicos, na redução da viabilidade celular de leveduras Saccharomyces cerevisiae. As bactérias Bacillus subtilis, Bacillus coagulans, Bacillus stearothermophilus, Lactobacillus fermentum e Lactobacillus plantarum foram cultivadas em associação com a levedura S. cerevisiae (cepa Y-904) por 72 horas a 32 °C, sob agitação. A viabilidade celular, a taxa de brotamento e a população de células de S. cerevisiae e a acidez total, a acidez volátil e o pH dos meios de cultivos foram determinados às 0, 24, 48 e 72 horas do cultivo misto. As culturas de bactérias foram tratadas através do calor, de agente antimicrobiano e de irradiação. Os resultados mostraram que apenas os meios de cultivo mais acidificados, contaminados com as bactérias ativas L. fermentum e B. subtilis, provocaram redução na viabilidade celular de S. cerevisiae. Excetuando a bactéria B. subtilis tratada com radiação gama, as demais bactérias tratadas pelos diferentes processos (calor, irradiação e antimicrobiano) não causaram diminuição da viabilidade celular e da população de S. cerevisiae, indicando que a presença isolada dos metabólitos celulares dessas bactérias não foi suficiente para reduzir a porcentagem de células vivas de S. cerevisiae.The aim of this project was to study the influence of the bacteria Bacillus and Lactobacillus, as well as their metabolic products to decrease the cellular viability of the yeast Saccharomyces cerevisiae. The bacteria Bacillus subtilis, Bacillus coagulans, Bacillus stearothermophilus, Lactobacillus fermentum and Lactobacillus plantarum were cultivated in association with yeast S. cerevisiae (strain Y-904) for 72 hours at 32 ºC under agitation. The cellular viability, budding rate and population of S. Cerevisiae and the total acidity, volatile acidity and pH of culture medium were determined at 0, 24, 48 and 72 hours of incubation of the mixed culture. The bacteria cultures were treated by heat sterilization, antibacterial agent and irradiation. The results showed that only the more acidified culture medium, contaminated with active bacteria L. fermentum and B. subtilis, caused a reduction in the yeast cellular viability. Except for the bacteria B. subtilis treated for radiation, the other bacteria treated by the different procedures (heat, radiation and antibacterial) did not cause a reduction in the cellular viability of S. cerevisiae, indicating that the isolated presence of the cellular metabolic of these bacteria was not enough to reduce the percentage of the living yeast cells.

Thais de Paula Nobre; Jorge Horii; André Ricardo Alcarde

2007-01-01

275

Bacteria: Fossil Record  

Science.gov (United States)

This description of the fossil record of bacteria focuses on one particular group of bacteria, the cyanobacteria or blue-green algae, which have left a fossil record that extends far back into the Precambrian. The oldest cyanobacteria-like fossils known are nearly 3.5 billion years old and are among the oldest fossils currently known. Cyanobacteria are larger than most bacteria and may secrete a thick cell wall. More importantly, cyanobacteria may form large layered structures, called stromatolites (if more or less dome-shaped) or oncolites (if round). The site also refers to pseudomorphs of pyrite and siderite, and a group of bacteria known as endolithic. Two links are available for more information. One provides information on the discovery of possible remains of bacteria-like organisms on a meteorite from Mars and the other has a research report on fossilized filamentous bacteria and other microbes, found in Cretaceous amber.

276

Production of L-ribose from L-ribulose by a triple-site variant of mannose-6-phosphate isomerase from Geobacillus thermodenitrificans.  

Science.gov (United States)

A triple-site variant (W17Q N90A L129F) of mannose-6-phosphate isomerase from Geobacillus thermodenitrificans was obtained by combining variants with residue substitutions at different positions after random and site-directed mutagenesis. The specific activity and catalytic efficiency (k(cat)/K(m)) for L-ribulose isomerization of this variant were 3.1- and 7.1-fold higher, respectively, than those of the wild-type enzyme at pH 7.0 and 70°C in the presence of 1 mM Co(2+). The triple-site variant produced 213 g/liter l-ribose from 300 g/liter L-ribulose for 60 min, with a volumetric productivity of 213 g liter(-1) h(-1), which was 4.5-fold higher than that of the wild-type enzyme. The k(cat)/K(m) and productivity of the triple-site variant were approximately 2-fold higher than those of the Thermus thermophilus R142N variant of mannose-6-phosphate isomerase, which exhibited the highest values previously reported. PMID:22447612

Lim, Yu-Ri; Yeom, Soo-Jin; Oh, Deok-Kun

2012-03-23

277

Production of L-ribose from L-ribulose by a triple-site variant of mannose-6-phosphate isomerase from Geobacillus thermodenitrificans.  

UK PubMed Central (United Kingdom)

A triple-site variant (W17Q N90A L129F) of mannose-6-phosphate isomerase from Geobacillus thermodenitrificans was obtained by combining variants with residue substitutions at different positions after random and site-directed mutagenesis. The specific activity and catalytic efficiency (k(cat)/K(m)) for L-ribulose isomerization of this variant were 3.1- and 7.1-fold higher, respectively, than those of the wild-type enzyme at pH 7.0 and 70°C in the presence of 1 mM Co(2+). The triple-site variant produced 213 g/liter l-ribose from 300 g/liter L-ribulose for 60 min, with a volumetric productivity of 213 g liter(-1) h(-1), which was 4.5-fold higher than that of the wild-type enzyme. The k(cat)/K(m) and productivity of the triple-site variant were approximately 2-fold higher than those of the Thermus thermophilus R142N variant of mannose-6-phosphate isomerase, which exhibited the highest values previously reported.

Lim YR; Yeom SJ; Oh DK

2012-06-01

278

Hyperthermostable, Ca(2+)-independent, and high maltose-forming alpha-amylase production by an extreme thermophile Geobacillus thermoleovorans: whole cell immobilization.  

UK PubMed Central (United Kingdom)

The synthesis of extracellular alpha-amylase in Geobacillus thermoleovorans was constitutive. The enzyme was secreted in metabolizable carbon sources as well as non-metabolizable synthetic analogues of glucose, but the titers were higher in the former than that in the latter. G. thermoleovorans is a fast-growing facultatively anaerobic bacterium that grows under both aerobic and anaerobic conditions and produces an extracellular amylolytic enzyme alpha-amylase with the by-product of lactic acid. G. thermoleovorans is a rich source of various novel thermostable biocatalysts for different industrial applications. alpha-Amylase synthesis was subject to catabolite repression in the presence of high concentrations of glucose. The addition of cAMP to the medium containing glucose did not result in the repression of alpha-amylase synthesis. The addition of maltose (1%) to the starch arginine medium resulted in a twofold enhancement in enzyme titers. Polyurethane foam (PUF)-immobilized cells secreted alpha-amylase, which was higher than that with the free cells. PUF appeared to be a better matrix for immobilization of the thermophilic bacterium than the other commonly used matrices. The repeated use of PUF-immobilized cells was possible over 15 cycles with a sustained alpha-amylase secretion. The use of this enzyme in starch saccharification eliminates the addition of Ca(2+) in starch liquefaction and its subsequent removal by ion exchangers from the product streams.

Rao JL; Satyanarayana T

2009-11-01

279

Hyperthermostable, Ca(2+)-independent, and high maltose-forming alpha-amylase production by an extreme thermophile Geobacillus thermoleovorans: whole cell immobilization.  

Science.gov (United States)

The synthesis of extracellular alpha-amylase in Geobacillus thermoleovorans was constitutive. The enzyme was secreted in metabolizable carbon sources as well as non-metabolizable synthetic analogues of glucose, but the titers were higher in the former than that in the latter. G. thermoleovorans is a fast-growing facultatively anaerobic bacterium that grows under both aerobic and anaerobic conditions and produces an extracellular amylolytic enzyme alpha-amylase with the by-product of lactic acid. G. thermoleovorans is a rich source of various novel thermostable biocatalysts for different industrial applications. alpha-Amylase synthesis was subject to catabolite repression in the presence of high concentrations of glucose. The addition of cAMP to the medium containing glucose did not result in the repression of alpha-amylase synthesis. The addition of maltose (1%) to the starch arginine medium resulted in a twofold enhancement in enzyme titers. Polyurethane foam (PUF)-immobilized cells secreted alpha-amylase, which was higher than that with the free cells. PUF appeared to be a better matrix for immobilization of the thermophilic bacterium than the other commonly used matrices. The repeated use of PUF-immobilized cells was possible over 15 cycles with a sustained alpha-amylase secretion. The use of this enzyme in starch saccharification eliminates the addition of Ca(2+) in starch liquefaction and its subsequent removal by ion exchangers from the product streams. PMID:19280125

Rao, J L Uma Maheswar; Satyanarayana, T

2009-03-12

280

Purification, crystallization and preliminary X-ray diffraction studies of the ATP-binding subunit of an ABC transporter from Geobacillus kaustophilus.  

Science.gov (United States)

ATP-binding cassette (ABC) transporters, also known as traffic ATPases, form a large family of integral membrane proteins responsible for the translocation of a variety of chemically diverse substrates across the lipid bilayers of cellular membranes of both prokaryotes and eukaryotes by the hydrolysis of ATP. The ATP-binding subunit of an ABC transporter from Geobacillus kaustophilus, a homodimeric enzyme, was overexpressed in Escherichia coli and purified. Crystals were obtained using the microbatch-under-oil method at 291?K. X-ray diffraction data to 1.6?Å resolution were collected on SPring-8 beamline BL26B1. The crystals belonged to the orthorhombic space group I222, with unit-cell parameters a=54.94, b=78.63, c=112.96?Å. Assuming the presence of a dimer in the asymmetric unit gave a crystal volume per protein weight (VM) of 2.32?Å3?Da(-1) and a solvent content of 47%; this was consistent with the results of a dynamic light-scattering experiment, which showed a dimeric state of the protein in solution. Molecular-replacement trials using the crystal structure of HisP from the Salmonella typhimurium ATP-binding subunit of an ABC transporter as a search model did not provide a satisfactory solution, indicating that the two ATP-binding subunits of ABC transporters have substantially different structures. PMID:23143260

Manjula, Mallappa; Pampa, Kudigana J; Madan Kumar, Shankar; Kunishima, Naoki; Lokanath, Neratur K

2012-10-30

 
 
 
 
281

Purification, crystallization and preliminary X-ray diffraction studies of the ATP-binding subunit of an ABC transporter from Geobacillus kaustophilus.  

UK PubMed Central (United Kingdom)

ATP-binding cassette (ABC) transporters, also known as traffic ATPases, form a large family of integral membrane proteins responsible for the translocation of a variety of chemically diverse substrates across the lipid bilayers of cellular membranes of both prokaryotes and eukaryotes by the hydrolysis of ATP. The ATP-binding subunit of an ABC transporter from Geobacillus kaustophilus, a homodimeric enzyme, was overexpressed in Escherichia coli and purified. Crystals were obtained using the microbatch-under-oil method at 291?K. X-ray diffraction data to 1.6?Å resolution were collected on SPring-8 beamline BL26B1. The crystals belonged to the orthorhombic space group I222, with unit-cell parameters a=54.94, b=78.63, c=112.96?Å. Assuming the presence of a dimer in the asymmetric unit gave a crystal volume per protein weight (VM) of 2.32?Å3?Da(-1) and a solvent content of 47%; this was consistent with the results of a dynamic light-scattering experiment, which showed a dimeric state of the protein in solution. Molecular-replacement trials using the crystal structure of HisP from the Salmonella typhimurium ATP-binding subunit of an ABC transporter as a search model did not provide a satisfactory solution, indicating that the two ATP-binding subunits of ABC transporters have substantially different structures.

Manjula M; Pampa KJ; Madan Kumar S; Kunishima N; Lokanath NK

2012-11-01

282

The physico-chemical characterization of casein-modified surfaces and their influence on the adhesion of spores from a Geobacillus species.  

UK PubMed Central (United Kingdom)

To gain a better understanding of the factors influencing spore adhesion in dairy manufacturing plants, casein-modified glass surfaces were prepared and characterized and their effect on the adhesion kinetics of spores from a Geobacillus sp., isolated from a dairy manufacturing plant (DMP) was assessed using a flow chamber. Surfaces were produced by initially silanizing glass using (3-glycidyloxypropyl) trimethoxysilane (GPS) or (3-aminopropyl) triethoxysilane to form epoxy-functionalized (G-GPS) or amino-functionalized glass (G-NH(2)) substrata. Casein was grafted to the G-GPS directly by its primary amino groups (G-GPS-casein) or to G-NH(2) by employing glutaraldehyde as a linking agent (G-NH(2)-glutar-casein). The surfaces were characterised using streaming potential measurements, contact angle goniometry, infrared spectroscopy and scanning electron microscopy. The attachment rate of spores suspended in 0.1 M KCl at pH 6.8, was highest on the positively charged (+14 mV) G-NH(2) surface (333 spores cm(-2) s(-1)) compared to the negatively charged glass (-22 mV), G-GPS (-20 mV) or G-GPS-casein (-21 mV) surfaces (162, 17 or 6 spores cm(-2) s(-1) respectively). Whilst there was a clear decrease in attachment rate to negatively charged casein-modified surfaces compared to the positively charged amine surface, there was no clear relationship between surface hydrophobicity and spore attachment rate.

Han J; Seale RB; Silcock P; McQuillan AJ; Bremer PJ

2011-05-01

283

The physico-chemical characterization of casein-modified surfaces and their influence on the adhesion of spores from a Geobacillus species.  

Science.gov (United States)

To gain a better understanding of the factors influencing spore adhesion in dairy manufacturing plants, casein-modified glass surfaces were prepared and characterized and their effect on the adhesion kinetics of spores from a Geobacillus sp., isolated from a dairy manufacturing plant (DMP) was assessed using a flow chamber. Surfaces were produced by initially silanizing glass using (3-glycidyloxypropyl) trimethoxysilane (GPS) or (3-aminopropyl) triethoxysilane to form epoxy-functionalized (G-GPS) or amino-functionalized glass (G-NH(2)) substrata. Casein was grafted to the G-GPS directly by its primary amino groups (G-GPS-casein) or to G-NH(2) by employing glutaraldehyde as a linking agent (G-NH(2)-glutar-casein). The surfaces were characterised using streaming potential measurements, contact angle goniometry, infrared spectroscopy and scanning electron microscopy. The attachment rate of spores suspended in 0.1 M KCl at pH 6.8, was highest on the positively charged (+14 mV) G-NH(2) surface (333 spores cm(-2) s(-1)) compared to the negatively charged glass (-22 mV), G-GPS (-20 mV) or G-GPS-casein (-21 mV) surfaces (162, 17 or 6 spores cm(-2) s(-1) respectively). Whilst there was a clear decrease in attachment rate to negatively charged casein-modified surfaces compared to the positively charged amine surface, there was no clear relationship between surface hydrophobicity and spore attachment rate. PMID:21598124

Han, Jingtian; Seale, R Brent; Silcock, Patrick; McQuillan, A James; Bremer, Phil J

2011-05-01

284

Characterization of recombinant amylopullulanase (gt-apu) and truncated amylopullulanase (gt-apuT) of the extreme thermophile Geobacillus thermoleovorans NP33 and their action in starch saccharification.  

UK PubMed Central (United Kingdom)

A gene encoding amylopullulanase (gt-apu) of the extremely thermophilic Geobacillus thermoleovorans NP33 was cloned and expressed in Escherichia coli. The gene has an open reading frame of 4,965 bp that encodes a protein of 1,655 amino acids with molecular mass of 182 kDa. The six conserved regions, characteristic of GH13 family, have been detected in gt-apu. The recombinant enzyme has only one active site for ?-amylase and pullulanase activities based on the enzyme kinetic analyses in a system that contains starch as well as pullulan as competing substrates and response to inhibitors. The end-product analysis confirmed that this is an endoacting enzyme. The specific enzyme activities for ?-amylase and pullulanase of the truncated amylopullulanase (gt-apuT) are higher than gt-apu. Both enzymes exhibited similar temperature (60 °C) and pH (7.0) optima, although gt-apuT possessed a higher thermostability than gt-apu. The overall catalytic efficiency (K(cat)/K(m)) of gt-apuT is greater than that of gt-apu, with almost similar substrate specificities. The C-terminal region of gt-apu appeared to be non-essential, and furthermore, it negatively affects the substrate binding and stability of the enzyme.

Nisha M; Satyanarayana T

2013-07-01

285

Characterization of recombinant amylopullulanase (gt-apu) and truncated amylopullulanase (gt-apuT) of the extreme thermophile Geobacillus thermoleovorans NP33 and their action in starch saccharification.  

Science.gov (United States)

A gene encoding amylopullulanase (gt-apu) of the extremely thermophilic Geobacillus thermoleovorans NP33 was cloned and expressed in Escherichia coli. The gene has an open reading frame of 4,965 bp that encodes a protein of 1,655 amino acids with molecular mass of 182 kDa. The six conserved regions, characteristic of GH13 family, have been detected in gt-apu. The recombinant enzyme has only one active site for ?-amylase and pullulanase activities based on the enzyme kinetic analyses in a system that contains starch as well as pullulan as competing substrates and response to inhibitors. The end-product analysis confirmed that this is an endoacting enzyme. The specific enzyme activities for ?-amylase and pullulanase of the truncated amylopullulanase (gt-apuT) are higher than gt-apu. Both enzymes exhibited similar temperature (60 °C) and pH (7.0) optima, although gt-apuT possessed a higher thermostability than gt-apu. The overall catalytic efficiency (K(cat)/K(m)) of gt-apuT is greater than that of gt-apu, with almost similar substrate specificities. The C-terminal region of gt-apu appeared to be non-essential, and furthermore, it negatively affects the substrate binding and stability of the enzyme. PMID:23132347

Nisha, M; Satyanarayana, T

2012-11-07

286

Characteristics of thermostable endoxylanase and ?-xylosidase of the extremely thermophilic bacterium Geobacillus thermodenitrificans TSAA1 and its applicability in generating xylooligosaccharides and xylose from agro-residues.  

UK PubMed Central (United Kingdom)

An extremely thermophilic bacterial isolate that produces a high titer of thermostable endoxylanase and ?-xylosidase extracellularly in an inducible manner was identified as Geobacillus thermodenitrificans TSAA1. The distinctive features of this strain are alkalitolerance and halotolerance. The endoxylanase is active over a broad range of pH (5.0-10.0) and temperatures (30-100 °C) with optima at pH 7.5 and 70 °C, while ?-xylosidase is optimally active at pH 7.0 and 60 °C. The T 1/2 values of the endoxylanase and ?-xylosidase are 30 min at 80 °C, and 180 min at 70 °C, respectively. The endoxylanase activity is stimulated by dithiothreitol, but inhibited strongly by EDAC and Woodward's reagent K. N-BS and DEPC strongly inhibited ?-xylosidase. MALDI-ToF (MS/MS) analysis of tryptic digest of ?-xylosidase revealed similarity with that of G. thermodenitrificans NG 80-2, and suggested that this belongs to the GH 52 glycosyl hydrolase super family. The action of endoxylanase on birch wood xylan and agro-residues such as wheat bran and wheat straw liberated xylooligosaccharides similar to endoxylanases of the family 10 glycoside hydrolases, while the enzyme preparation having both endoxylanase and ?-xylosidase liberated xylose as main hydrolysis product.

Anand A; Kumar V; Satyanarayana T

2013-05-01

287

Genomics of Probiotic Bacteria  

Science.gov (United States)

Probiotic bacteria from the Lactobacillus and Bifidobacterium species belong to the Firmicutes and the Actinobacteria phylum, respectively. Lactobacilli are members of the lactic acid bacteria (LAB) group, a broadly defined family of microorganisms that ferment various hexoses into primarily lactic acid. Lactobacilli are typically low G + C gram-positive species which are phylogenetically diverse, with over 100 species documented to date. Bifidobacteria are heterofermentative, high G + C content bacteria with about 30 species of bifidobacteria described to date.

O'Flaherty, Sarah; Goh, Yong Jun; Klaenhammer, Todd R.

288

Bacteria-Antagonists  

International Science & Technology Center (ISTC)

Development of Biological Control Agents Through Use of Recombinant Antagonistic Bacteria Possessing Variable Mechanisms of Antagonisms, High Colonizing Capacity and Marker Traits for their Monitoring in Nature

289

Introduction to Bacteria  

Science.gov (United States)

This science site has students research how bacteria move, where they live, and how they reproduce; learn how bacteria can be helpful or harmful; and create a design illustrating what they have learned about bacteria. Included in the lesson plan are the objectives, needed materials and Web sites, procedures, discussion questions, evaluation, extensions, suggested reading, and vocabulary. Teachers can link to Teaching Tools to create custom worksheets, puzzles, and quizzes. A printable version of the lesson plan can be downloaded. The video Bacteria, Viruses and Allergies can be purchased and comprehension questions and answers can be downloaded.

Discoveryschool.com; Fenichel, Marilyn

2007-12-12

290

Rapid identification of dairy mesophilic and thermophilic sporeforming bacteria using DNA high resolution melt analysis of variable 16S rDNA regions.  

Science.gov (United States)

Due to their ubiquity in the environment and ability to survive heating processes, sporeforming bacteria are commonly found in foods. This can lead to product spoilage if spores are present in sufficient numbers and where storage conditions favour spore germination and growth. A rapid method to identify the major aerobic sporeforming groups in dairy products, including Bacillus licheniformis group, Bacillus subtilis group, Bacillus pumilus group, Bacillus megaterium, Bacillus cereus group, Geobacillus species and Anoxybacillus flavithermus was devised. This method involves real-time PCR and high resolution melt analysis (HRMA) of V3 (~70 bp) and V6 (~100 bp) variable regions in the 16S rDNA. Comparisons of HRMA curves from 194 isolates of the above listed sporeforming bacteria obtained from dairy products which were identified using partial 16S rDNA sequencing, allowed the establishment of criteria for differentiating them from each other and several non-sporeforming bacteria found in samples. A blinded validation trial on 28 bacterial isolates demonstrated complete accuracy in unambiguous identification of the 7 different aerobic sporeformers. The reliability of HRMA method was also verified using boiled extractions of crude DNA, thereby shortening the time needed for identification. The HRMA method described in this study provides a new and rapid approach to identify the dominant mesophilic and thermophilic aerobic sporeforming bacteria found in a wide variety of dairy products. PMID:23743474

Chauhan, Kanika; Dhakal, Rajat; Seale, R Brent; Deeth, Hilton C; Pillidge, Christopher J; Powell, Ian B; Craven, Heather; Turner, Mark S

2013-05-17

291

Rapid identification of dairy mesophilic and thermophilic sporeforming bacteria using DNA high resolution melt analysis of variable 16S rDNA regions.  

UK PubMed Central (United Kingdom)

Due to their ubiquity in the environment and ability to survive heating processes, sporeforming bacteria are commonly found in foods. This can lead to product spoilage if spores are present in sufficient numbers and where storage conditions favour spore germination and growth. A rapid method to identify the major aerobic sporeforming groups in dairy products, including Bacillus licheniformis group, Bacillus subtilis group, Bacillus pumilus group, Bacillus megaterium, Bacillus cereus group, Geobacillus species and Anoxybacillus flavithermus was devised. This method involves real-time PCR and high resolution melt analysis (HRMA) of V3 (~70 bp) and V6 (~100 bp) variable regions in the 16S rDNA. Comparisons of HRMA curves from 194 isolates of the above listed sporeforming bacteria obtained from dairy products which were identified using partial 16S rDNA sequencing, allowed the establishment of criteria for differentiating them from each other and several non-sporeforming bacteria found in samples. A blinded validation trial on 28 bacterial isolates demonstrated complete accuracy in unambiguous identification of the 7 different aerobic sporeformers. The reliability of HRMA method was also verified using boiled extractions of crude DNA, thereby shortening the time needed for identification. The HRMA method described in this study provides a new and rapid approach to identify the dominant mesophilic and thermophilic aerobic sporeforming bacteria found in a wide variety of dairy products.

Chauhan K; Dhakal R; Seale RB; Deeth HC; Pillidge CJ; Powell IB; Craven H; Turner MS

2013-07-01

292

Structural homology between elongation factors EF-Tu from Bacillus stearothermophilus and Escherichia coli in the binding site for aminoacyl-tRNA.  

Science.gov (United States)

Elongation factor EF-Tu (Mr approximately equal to 50 000) and elongation factor EF-G (Mr approximately equal to 78 000) were isolated from Bacillus stearothermophilus in a homogeneous form. The ability of EF-Tu to participate in protein synthesis is rapidly inactivated by N-tosyl-L-phenyl-alanylchloromethane (Tos-PheCH2Cl). EF-Tu X GTP is more susceptible to the inhibition by Tos-PheCH2Cl than is EF-Tu X GDP. Tos-PheCH2Cl forms a covalent equimolar complex with the factor by reacting with a cysteine residue in its molecule. The labelling of EF-Tu by the reagent irreversibly destroys its ability to bind aminoacyl-tRNA, which in turn protects the protein from this inactivation. This indicates that the modification of EF-Tu by Tos-PheCH2Cl occurs at the aminoacyl-tRNA binding site of the protein. To identify and characterize the site of aminoacyl-tRNA binding in EF-Tu, the factor was labelled with [14C]Tos-PheCH2Cl, digested with trypsin, the resulting peptides were separated by high-performance liquid chromatography and the sequence of the radioactive peptide was determined. The peptide has identical structure with an Escherichia coli EF-Tu tryptic peptide comprising the residues 75-89 and the Tos-PheCH2Cl-reactive cysteine at position 81 [Jonák, J., Petersen, T. E., Clark, B. F. C. and Rychlík, I. (1982) FEBS Lett. 150, 485-488]. Experiments on photo-oxidation of EF-Tu by visible light in the presence of rose bengal dye showed that there are apparently two histidine residues in elongation factor Tu from B. stearothermophilus which are essential for the interaction with aminoacyl-tRNA. This is clearly reminiscent of a similar situation in E. coli EF-Tu [Jonák, J., Petersen, T. E., Meloun, B. and Rychlík, I. (1984) Eur. J. Biochem. 144, 295-303]. Our results provide further evidence for the conserved nature of the site of aminoacyl-tRNA binding in elongation factor EF-Tu and show that Tos-PheCH2Cl reagent might be a favourable tool for the identification of the site in the structure of prokaryotic EF-Tus. PMID:3510872

Jonák, J; Pokorná, K; Meloun, B; Karas, K

1986-01-15

293

Structural homology between elongation factors EF-Tu from Bacillus stearothermophilus and Escherichia coli in the binding site for aminoacyl-tRNA.  

UK PubMed Central (United Kingdom)

Elongation factor EF-Tu (Mr approximately equal to 50 000) and elongation factor EF-G (Mr approximately equal to 78 000) were isolated from Bacillus stearothermophilus in a homogeneous form. The ability of EF-Tu to participate in protein synthesis is rapidly inactivated by N-tosyl-L-phenyl-alanylchloromethane (Tos-PheCH2Cl). EF-Tu X GTP is more susceptible to the inhibition by Tos-PheCH2Cl than is EF-Tu X GDP. Tos-PheCH2Cl forms a covalent equimolar complex with the factor by reacting with a cysteine residue in its molecule. The labelling of EF-Tu by the reagent irreversibly destroys its ability to bind aminoacyl-tRNA, which in turn protects the protein from this inactivation. This indicates that the modification of EF-Tu by Tos-PheCH2Cl occurs at the aminoacyl-tRNA binding site of the protein. To identify and characterize the site of aminoacyl-tRNA binding in EF-Tu, the factor was labelled with [14C]Tos-PheCH2Cl, digested with trypsin, the resulting peptides were separated by high-performance liquid chromatography and the sequence of the radioactive peptide was determined. The peptide has identical structure with an Escherichia coli EF-Tu tryptic peptide comprising the residues 75-89 and the Tos-PheCH2Cl-reactive cysteine at position 81 [Jonák, J., Petersen, T. E., Clark, B. F. C. and Rychlík, I. (1982) FEBS Lett. 150, 485-488]. Experiments on photo-oxidation of EF-Tu by visible light in the presence of rose bengal dye showed that there are apparently two histidine residues in elongation factor Tu from B. stearothermophilus which are essential for the interaction with aminoacyl-tRNA. This is clearly reminiscent of a similar situation in E. coli EF-Tu [Jonák, J., Petersen, T. E., Meloun, B. and Rychlík, I. (1984) Eur. J. Biochem. 144, 295-303]. Our results provide further evidence for the conserved nature of the site of aminoacyl-tRNA binding in elongation factor EF-Tu and show that Tos-PheCH2Cl reagent might be a favourable tool for the identification of the site in the structure of prokaryotic EF-Tus.

Jonák J; Pokorná K; Meloun B; Karas K

1986-01-01

294

How bacteria choose phosphate.  

UK PubMed Central (United Kingdom)

Discriminating against arsenate: Determination of the structure of periplasmic phosphate binding proteins at very high resolution provides the basis for understanding the high discrimination of bacteria against arsenate. The results complete our insight into earlier erroneous conclusions on the ability of certain bacteria to use arsenate instead of phosphate.

Goody RS

2013-02-01

295

Cellular viability of Saccharomyces cerevisiae cultivated in association with contaminates bacteria of alcoholic fermentation;Viabilidade celular de Saccharomyces cerevisiae cultivada em associacao com bacterias contaminantes da fermentacao alcoolica  

Energy Technology Data Exchange (ETDEWEB)

The aim of this work was to study the influence of the bacteria Bacillus and Lactobacillus, as well as their metabolic products, in reduction of cellular viability of Saccharomyces cerevisiae, when in mixed culture of yeast and active and treated bacteria. Also was to evaluated an alternative medium (MCC) for the cultivation of bacteria and yeast, constituted of sugarcane juice diluted to 5 deg Brix and supplemented with yeast extract and peptone. The bacteria Bacillus subtilis, Bacillus coagulans, Bacillus stearothermophilus, Lactobacillus fermentum and Lactobacillus plantarum were cultivated in association with yeast Saccharomyces cerevisiae (strain Y-904) for 72 h on 32 deg C, under agitation. The cellular viability, budding rate and population of S. cerevisiae, the total acidity, volatile acidity and pH of culture were determined from 0, 24, 48 e 72 h of mixed culture. Also were determined the initial and final of microorganism population across the pour plate method, in traditional culture medium (PCA for Bacillus, MRS-agar for Lactobacillus and YEPD-agar for yeast S. cerevisiae) and in medium constituted of sugarcane juice. The bacteria cultures were treated by heat sterilization (120 deg C for 20 minutes), antibacterial agent (Kamoran HJ in concentration 3,0 ppm) or irradiation (radiation gamma, with doses of 5,0 kGy for Lactobacillus and 15,0 kGy for Bacillus). The results of the present research showed that just the culture mediums more acids (with higher concentrations of total and volatile acidity, and smaller values of pH), contaminated with active bacteria L. fermentum and B. subtilis, caused reduction on yeast cellular viability. Except the bacteria B. subtilis treated with radiation, the others bacteria treated by different procedures (heat, radiation e antibacterial) did not cause reduction on yeast cellular viability and population, indicating that the isolated presence of the cellular metabolic of theses bacteria was not enough to reduce the percentage of the yeast live cells and a density population. For all microorganisms, the counts obtained with the cultivation medium constituted of sugarcane juice were similar obtained in traditional mediums, probably because the alternative medium simulate the composition of sugarcane must, that the bacteria were isolated in industrial process of ethanol yield. However, the culture medium constituted of sugarcane juice could be replacing traditional culture mediums of yeast and bacteria tested in this work. (author)

Nobre, Thais de Paula

2005-07-01

296

Simultaneous hydrocarbon biodegradation and biosurfactant production by oilfield-selected bacteria.  

UK PubMed Central (United Kingdom)

AIMS: To study the bacterial diversity associated with hydrocarbon biodegradation potentiality and biosurfactant production of Tunisian oilfields bacteria. METHODS AND RESULTS: Eight Tunisian hydrocarbonoclastic oilfields bacteria have been isolated and selected for further characterization studies. Phylogenetic analysis revealed that three thermophilic strains belonged to the genera Geobacillus, Bacillus and Brevibacillus, and that five mesophilic strains belonged to the genera Pseudomonas, Lysinibacillus, Achromobacter and Halomonas. The bacterial strains were cultivated on crude oil as sole carbon and energy sources, in the presence of different NaCl concentrations (1, 5 and 10%, w/v), and at 37 or 55°C. The hydrocarbon biodegradation potential of each strain was quantified by GC-MS. Strain C450R, phylogenetically related to the species Pseudomonas aeruginosa, showed the maximum crude oil degradation potentiality. During the growth of strain C450R on crude oil (2%, v/v), the emulsifying activity (E24) and glycoside content increased and reached values of 77 and 1.33 g l(-1), respectively. In addition, the surface tension (ST) decreased from 68 to 35.1 mN m(-1), suggesting the production of a rhamnolipid biosurfactant. Crude biosurfactant had been partially purified and characterized. It showed interest stability against temperature and salinity increasing and important emulsifying activity against oils and hydrocarbons. CONCLUSIONS: The results of this study showed the presence of diverse aerobic bacteria in Tunisian oilfields including mesophilic, thermophilic and halotolerant strains with interesting aliphatic hydrocarbon degradation potentiality, mainly for the most biosurfactant produced strains. SIGNIFICANCE AND IMPACT OF THE STUDY: It may be suggested that the bacterial isolates are suitable candidates for practical field application for effective in situ bioremediation of hydrocarbon-contaminated sites.

Mnif S; Chamkha M; Labat M; Sayadi S

2011-09-01

297

Synthesis of cinnamyl alcohol from cinnamaldehyde with Bacillus stearothermophilus alcohol dehydrogenase as the isolated enzyme and in recombinant E. coli cells.  

UK PubMed Central (United Kingdom)

The synthesis of the aroma chemical cinnamyl alcohol (CMO) by means of enzymatic reduction of cinnamaldehyde (CMA) was investigated using NADH-dependent alcohol dehydrogenase from Bacillus stearothermophilus both as an isolated enzyme, and in recombinant Escherichia coli whole cells. The influence of parameters such as reaction time and cofactor, substrate, co-substrate 2-propanol and biocatalyst concentrations on the bioreduction reaction was investigated and an efficient and sustainable one-phase system developed. The reduction of CMA (0.5 g/L, 3.8 mmol/L) by the isolated enzyme occurred in 3 h at 50 °C with 97% conversion, and yielded high purity CMO (?98%) with a yield of 88% and a productivity of 50 g/genzyme. The reduction of 12.5 g/L (94 mmol/L) CMA by whole cells in 6 h, at 37 °C and no requirement of external cofactor occurred with 97% conversion, 82% yield of 98% pure alcohol and a productivity of 34 mg/gwet cell weight. The results demonstrate the microbial system as a practical and efficient method for larger-scale synthesis of CMO.

Pennacchio A; Rossi M; Raia CA

2013-07-01

298

Synthesis of cinnamyl alcohol from cinnamaldehyde with Bacillus stearothermophilus alcohol dehydrogenase as the isolated enzyme and in recombinant E. coli cells.  

Science.gov (United States)

The synthesis of the aroma chemical cinnamyl alcohol (CMO) by means of enzymatic reduction of cinnamaldehyde (CMA) was investigated using NADH-dependent alcohol dehydrogenase from Bacillus stearothermophilus both as an isolated enzyme, and in recombinant Escherichia coli whole cells. The influence of parameters such as reaction time and cofactor, substrate, co-substrate 2-propanol and biocatalyst concentrations on the bioreduction reaction was investigated and an efficient and sustainable one-phase system developed. The reduction of CMA (0.5 g/L, 3.8 mmol/L) by the isolated enzyme occurred in 3 h at 50 °C with 97% conversion, and yielded high purity CMO (?98%) with a yield of 88% and a productivity of 50 g/genzyme. The reduction of 12.5 g/L (94 mmol/L) CMA by whole cells in 6 h, at 37 °C and no requirement of external cofactor occurred with 97% conversion, 82% yield of 98% pure alcohol and a productivity of 34 mg/gwet cell weight. The results demonstrate the microbial system as a practical and efficient method for larger-scale synthesis of CMO. PMID:23686507

Pennacchio, Angela; Rossi, Mosè; Raia, Carlo A

2013-05-19

299

Application of artificial neural networks to describe the combined effect of pH and NaCl on the heat resistance of Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

A model for prediction of bacterial spore inactivation was developed. The influence of temperature, pH and NaCl on the heat resistance of Bacillus stearothermophilus spores was described using low-complexity, black box models based on artificial neural networks. Literature data were used to build and train the neural network, and new experimental data were used to evaluate it. The neural network models gave better predictions than the classical quadratic response surface model in all the experiments tried. When the neural networks were evaluated using new experimental data, also good predictions were obtained, providing fail-safe predictions of D values in all cases. The weights and biases values of neurons of the neural network that gave the best results are presented, so the reader can use the model for their own purposes. The use of this non-linear modelling technique makes it possible to describe more accurately interacting effects of environmental factors when compared with classical predictive microbial models.

Esnoz A; Periago PM; Conesa R; Palop A

2006-02-01

300

Application of artificial neural networks to describe the combined effect of pH and NaCl on the heat resistance of Bacillus stearothermophilus.  

Science.gov (United States)

A model for prediction of bacterial spore inactivation was developed. The influence of temperature, pH and NaCl on the heat resistance of Bacillus stearothermophilus spores was described using low-complexity, black box models based on artificial neural networks. Literature data were used to build and train the neural network, and new experimental data were used to evaluate it. The neural network models gave better predictions than the classical quadratic response surface model in all the experiments tried. When the neural networks were evaluated using new experimental data, also good predictions were obtained, providing fail-safe predictions of D values in all cases. The weights and biases values of neurons of the neural network that gave the best results are presented, so the reader can use the model for their own purposes. The use of this non-linear modelling technique makes it possible to describe more accurately interacting effects of environmental factors when compared with classical predictive microbial models. PMID:16216369

Esnoz, A; Periago, P M; Conesa, R; Palop, A

2005-10-10

 
 
 
 
301

Presence and potential role of thermophilic bacteria in temperate terrestrial environments  

Science.gov (United States)

Organic sulfur and nitrogen are major reservoirs of these elements in terrestrial systems, although their cycling remains to be fully understood. Both sulfur and nitrogen mineralization are directly related to microbial metabolism. Mesophiles and thermophiles were isolated from temperate environments. Thermophilic isolates were classified within the Firmicutes, belonging to the Geobacillus, Brevibacillus, and Ureibacillus genera, and showed optimum growth temperatures between 50°C and 60°C. Sulfate and ammonium produced were higher during growth of thermophiles both for isolated strains and natural bacterial assemblages. They were positively related to organic nutrient load. Temperature also affected the release of sulfate and ammonium by thermophiles. Quantitative, real-time reverse-transcription polymerase chain reaction on environmental samples indicated that the examined thermophilic Firmicutes represented up to 3.4% of the total bacterial community RNA. Temperature measurements during summer days showed values above 40°C for more than 10 h a day in soils from southern Spain. These results support a potential role of thermophilic bacteria in temperate terrestrial environments by mineralizing organic sulfur and nitrogen ruled by the existence and length of warm periods.

Portillo, M. C.; Santana, M.; Gonzalez, J. M.

2012-01-01

302

Presence and potential role of thermophilic bacteria in temperate terrestrial environments.  

UK PubMed Central (United Kingdom)

Organic sulfur and nitrogen are major reservoirs of these elements in terrestrial systems, although their cycling remains to be fully understood. Both sulfur and nitrogen mineralization are directly related to microbial metabolism. Mesophiles and thermophiles were isolated from temperate environments. Thermophilic isolates were classified within the Firmicutes, belonging to the Geobacillus, Brevibacillus, and Ureibacillus genera, and showed optimum growth temperatures between 50°C and 60°C. Sulfate and ammonium produced were higher during growth of thermophiles both for isolated strains and natural bacterial assemblages. They were positively related to organic nutrient load. Temperature also affected the release of sulfate and ammonium by thermophiles. Quantitative, real-time reverse-transcription polymerase chain reaction on environmental samples indicated that the examined thermophilic Firmicutes represented up to 3.4% of the total bacterial community RNA. Temperature measurements during summer days showed values above 40°C for more than 10 h a day in soils from southern Spain. These results support a potential role of thermophilic bacteria in temperate terrestrial environments by mineralizing organic sulfur and nitrogen ruled by the existence and length of warm periods.

Portillo MC; Santana M; Gonzalez JM

2012-01-01

303

Purification studies on a thermo-active amidase of Geobacillus pallidus BTP-5x MTCC 9225 isolated from thermal springs of Tatapani (Himachal Pradesh).  

Science.gov (United States)

An intracellular aliphatic amide degrading inducible thermo-active amidase produced by Geobacillus pallidus BTP-5x MTCC 9225 was purified to apparent homogeneity using anion exchange and gel filtration chromatography, giving a yield of 6.7 % and a specific activity of 30.49 units mg(-1). The purified protein migrated as a single band of estimated molecular mass of 158 kDa (homo-tetramer) in 8 % polyacrylamide gel electrophoresis and ?38.5 kDa in 12 % sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Optima of pH and temperature varied widely in broad pH range (pH 6-9) and temperature range (45-70 °C). The purified amidase was stable up to 6 h at 50 °C, with a t (1/2) of 7 h at 55 °C. The multimeric nature of the holozyme (tetramer) contributed to protection of the enzyme against thermal denaturation. The enzyme showed resistance to metal chelating agents (EDTA, 8-hydroxyquinoline, and sodium azide), explaining its non-metallic nature, and is strongly inhibited by thiol reagents that means cysteine is involved in catalysis. The amidase of G. pallidus BTP-5x preferentially hydrolyzed only small aliphatic amides and has a narrow substrate spectrum. The K (M) value for acrylamide is 10.54 mM, V (max) 45.19 ?mol(-1)?min(-1)?mg(-1) protein, and k (cat) 4.29 min(-1). The sequence of amino acids of the purified enzyme MRHGDISSSHDTVGI appears similar to thermophilic amidases. Sequence analysis of the amidase gene showed that the enzyme is 347 amino-acid-long with a molecular weight of 38.4 kDa (as observed in SDS-PAGE), theoretical pI 5.38, and show strong similarity to thermostable amidases, possessing unique restriction sites. PMID:23096998

Sharma, Monica; Sharma, Nitya Nand; Bhalla, Tek Chand

2012-10-25

304

Improvement of Thermal Stability via Outer-Loop Ion Pair Interaction of Mutated T1 Lipase from Geobacillus zalihae Strain T1  

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Full Text Available Mutant D311E and K344R were constructed using site-directed mutagenesis to introduce an additional ion pair at the inter-loop and the intra-loop, respectively, to determine the effect of ion pairs on the stability of T1 lipase isolated from Geobacillus zalihae. A series of purification steps was applied, and the pure lipases of T1, D311E and K344R were obtained. The wild-type and mutant lipases were analyzed using circular dichroism. The Tm for T1 lipase, D311E lipase and K344R lipase were approximately 68.52 °C, 70.59 °C and 68.54 °C, respectively. Mutation at D311 increases the stability of T1 lipase and exhibited higher Tm as compared to the wild-type and K344R. Based on the above, D311E lipase was chosen for further study. D311E lipase was successfully crystallized using the sitting drop vapor diffusion method. The crystal was diffracted at 2.1 Å using an in-house X-ray beam and belonged to the monoclinic space group C2 with the unit cell parameters a = 117.32 Å, b = 81.16 Å and c = 100.14 Å. Structural analysis showed the existence of an additional ion pair around E311 in the structure of D311E. The additional ion pair in D311E may regulate the stability of this mutant lipase at high temperatures as predicted in silico and spectroscopically.

Rudzanna Ruslan; Raja Noor Zaliha Raja Abd. Rahman; Thean Chor Leow; Mohd Shukuri Mohamad Ali; Mahiran Basri; Abu Bakar Salleh

2012-01-01

305

Purification studies on a thermo-active amidase of Geobacillus pallidus BTP-5x MTCC 9225 isolated from thermal springs of Tatapani (Himachal Pradesh).  

UK PubMed Central (United Kingdom)

An intracellular aliphatic amide degrading inducible thermo-active amidase produced by Geobacillus pallidus BTP-5x MTCC 9225 was purified to apparent homogeneity using anion exchange and gel filtration chromatography, giving a yield of 6.7 % and a specific activity of 30.49 units mg(-1). The purified protein migrated as a single band of estimated molecular mass of 158 kDa (homo-tetramer) in 8 % polyacrylamide gel electrophoresis and ?38.5 kDa in 12 % sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Optima of pH and temperature varied widely in broad pH range (pH 6-9) and temperature range (45-70 °C). The purified amidase was stable up to 6 h at 50 °C, with a t (1/2) of 7 h at 55 °C. The multimeric nature of the holozyme (tetramer) contributed to protection of the enzyme against thermal denaturation. The enzyme showed resistance to metal chelating agents (EDTA, 8-hydroxyquinoline, and sodium azide), explaining its non-metallic nature, and is strongly inhibited by thiol reagents that means cysteine is involved in catalysis. The amidase of G. pallidus BTP-5x preferentially hydrolyzed only small aliphatic amides and has a narrow substrate spectrum. The K (M) value for acrylamide is 10.54 mM, V (max) 45.19 ?mol(-1)?min(-1)?mg(-1) protein, and k (cat) 4.29 min(-1). The sequence of amino acids of the purified enzyme MRHGDISSSHDTVGI appears similar to thermophilic amidases. Sequence analysis of the amidase gene showed that the enzyme is 347 amino-acid-long with a molecular weight of 38.4 kDa (as observed in SDS-PAGE), theoretical pI 5.38, and show strong similarity to thermostable amidases, possessing unique restriction sites.

Sharma M; Sharma NN; Bhalla TC

2013-01-01

306

Bacteria-virus coevolution.  

UK PubMed Central (United Kingdom)

Phages, viruses of bacteria, are ubiquitous. Many phages require host cell death to successfully complete their life cycle, resulting in reciprocal evolution of bacterial resistance and phage infectivity (antagonistic coevolution). Such coevolution can have profound consequences at all levels of biological organisation. Here, we review genetic and ecological factors that contribute to determining coevolutionary dynamics between bacteria and phages. We also consider some of the consequences of bacteria-phage coevolution, such as determining rates of molecular evolution and structuring communities, and how these in turn feedback into driving coevolutionary dynamics.

Buckling A; Brockhurst M

2012-01-01

307

[Respiration of oligonitrophilic bacteria  

UK PubMed Central (United Kingdom)

The respiration rate of 25 studied oligonitrophilous strains belonging to various genera and species of bacteria varied within wide limits depending on their taxonomy, conditions of cultivation, substrates, and other factors. No strict correlation was established between the respiration rate and the activity of nitrogen fixation. One of the typical properties of oligonitrophilous bacteria is a high rate of endogeneous respiration --50% and more of the respiration rate during assimilation of exogeneous substrates. The level of oxidative metabolism of oligonitrophilous bacteria was found to be higher on media containing bound nitrogen than on media without nitrogen.

Mal'tseva NN; Ivanitskaia LM; Skliar IuV

1975-11-01

308

Magnetite biomineralization in bacteria.  

UK PubMed Central (United Kingdom)

Magnetotactic bacteria are able to biomineralize magnetic crystals in intracellular organelles, so-called "magnetosomes." These particles exhibit species- and strain-specific size and morphology. They are of great interest for biomimetic nanotechnological and biotechnological research due to their fine-tuned magnetic properties and because they challenge our understanding of the classical principles of crystallization. Magnetotactic bacteria use these highly optimized particles, which form chains within the bacterial cells, as a magnetic field actuator, enabling them to navigate. In this chapter, we discuss the current biological and chemical knowledge of magnetite biomineralization in these bacteria. We highlight the extraordinary properties of magnetosomes and some resulting potential applications.

Baumgartner J; Faivre D

2011-01-01

309

[Darwin and bacteria].  

UK PubMed Central (United Kingdom)

As in 2009 the scientific world celebrates two hundreds years from the birthday of Charles Darwin and one hundred and fifty from the publication of The Origin of Species, an analysis of his complete work is performed, looking for any mention of bacteria. But it seems that the great naturahst never took knowledge about its existence, something rather improbable in a time when the discovery of bacteria shook the medical world, or he deliberately ignored them, not finding a place for such microscopic beings into his theory of evolution. But the bacteria badly affected his familiar life, killing scarlet fever one of his children and worsening to death the evolution of tuberculosis of his favourite Annie. Darwin himself could suffer the sickness of Chagas, whose etiological agent has a similar level to bacteria in the scale of evolution.

Ledermann D W

2009-02-01

310

Bacteria and Foodborne Illness  

Science.gov (United States)

... foodborne illnesses may lead to chronic disorders, including reactive arthritis, a type of joint inflammation that usually affects ... by certain bacteria, including C. jejuni and Salmonella . Reactive arthritis usually lasts fewer than 6 months, but this ...

311

Quorum Sensingin Bacteria  

UK PubMed Central (United Kingdom)

Bacteria can co ordinate group activities by cell cell communicate with each other. The interbacterial communication system known as quorum sensing(QS) utilizes hormones, such as acyl homoserine lactone(AHL), oligopeptide and furanosyl borate diester, to regulate bacterial gene expression. This review describes bacterial QS regulatory cascades of gram negative and postive bacteria. Finally we discuss strategy of anti QS system for plant controlling bacterial disease.

Zhou Yi; Liu Xiaojin; Zhu Chenguang; Sun Ming; Yu Ziniu

2004-01-01

312

Immobilization of BSA, enzymes and cells of Bacillus stearothermophilus onto cellulose polygalacturonic acid and starch based graft copolymers containing maleic arhydride  

Energy Technology Data Exchange (ETDEWEB)

Poly(maleic anhydride styrene) graft copolymers of cellulose, pectin polygalacturonic acid salt, calcium polygalacturonate, and starch were prepared and used to immobilize proteins. The cellulose grafts coupled quite appreciable quantities of acid phosphatase, glucose oxidase, and trypsin. However, the general retention of activity was somewhat disappointing. Further investigation with acid phosphatase showed that the amount of enzyme immobilized increased as the amount of anhydride in the graft copolymer increased but no such relationship existed for the enzymic activity. The cellulose graft copolymers were hydrolyzed and it appeared that the carboxyl group aided adsorption of the enzyme. Attempts to couple acid phosphatase using CMC through the free carboxyl groups, created by hydrolysis, gave only a small increase in the extent of protein coupling. However, the unhydrolyzed system gave a useful degree of immobilization of cells of Bacillus stearothermophilus, as did a poly(maleic anhydride/styrene)-cocellulose system. Attempts to improve the activity by using grafts based on other polysaccharide supports met with mixed success. Pectin products were soluble. Polygalacturonic acid products were partially soluble and extremely high levels of enzymic activity were obtained. This was probably due in part to the hydrophilic nature of the system, which also encouraged absorption of the enzyme. Attempts were made to reduce the solubility by using the calcium pectinate salt. Immobilization of acid phosphatase and trypsin resulted in increased protein coupling but relatively poor activities were attained. Calcium polygalacturonate was used to prepare an insoluble graft copolymeric system containing acrylonitrile-comaleic anhydride. The resulting gels gave excellent coupling with acid phosphatase which had a very good retention of activity.

Beddows, C.G.; Gil, M.H.; Guthrie, J.T.

1986-01-01

313

Ice-Nucleating Bacteria  

Science.gov (United States)

Since the discovery of ice-nucleating bacteria in 1974 by Maki et al., a large number of studies on the biological characteristics, ice-nucleating substance, ice nucleation gene and frost damage etc. of the bacteria have been carried out. Ice-nucleating bacteria can cause the freezing of water at relatively warm temperature (-2.3°C). Tween 20 was good substrates for ice-nucleating activity of Pseudomonas fluorescens KUIN-1. Major fatty acids of Isolate (Pseudomonas fluorescens) W-11 grown at 30°C were palmitic, cis-9-hexadecenoic and cis-11-octadecenoic which amounted to 90% of the total fatty acids. Sequence analysis shows that an ice nucleation gene from Pseudomonas fluorescens is related to the gene of Pseudomonas syringae.

Obata, Hitoshi

314

Cytokinesis in Bacteria  

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Work on two diverse rod-shaped bacteria, Escherichia coli and Bacillus subtilis, has defined a set of about 10 conserved proteins that are important for cell division in a wide range of eubacteria. These proteins are directed to the division site by the combination of two negative regulatory systems...

Errington, Jeffery; Daniel, Richard A.; Scheffers, Dirk-Jan

315

Immunity to intracellular bacteria  

Directory of Open Access Journals (Sweden)

Full Text Available Immunity to intracellular bacteria including Mycobacterium tuberculosis. Mycobacterium leprae, and Listeria monocytogenes depends on specific T cells. Evidence to be described suggests that CD4 (alpha/beta)T cells which interact with each other and with macrophages contribute to acquired resistence against as well as pathogenesis of intracellular bacterial infections.

Stefan H. E. Kaufmann; George A. Follows; Martin E. Munik

1992-01-01

316

Bacteria, Phages and Septicemia  

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The use of phages is an attractive option to battle antibiotic resistant bacteria in certain bacterial infections, but the role of phage ecology in bacterial infections is obscure. Here we surveyed the phage ecology in septicemia, the most severe type of bacterial infection. We observed that the maj...

Gaidelyt?, Aušra; Vaara, Martti; Bamford, Dennis H.

317

Bacteria-surface interactions.  

UK PubMed Central (United Kingdom)

The interaction of bacteria with surfaces has important implications in a range of areas, including bioenergy, biofouling, biofilm formation, and the infection of plants and animals. Many of the interactions of bacteria with surfaces produce changes in the expression of genes that influence cell morphology and behavior, including genes essential for motility and surface attachment. Despite the attention that these phenotypes have garnered, the bacterial systems used for sensing and responding to surfaces are still not well understood. An understanding of these mechanisms will guide the development of new classes of materials that inhibit and promote cell growth, and complement studies of the physiology of bacteria in contact with surfaces. Recent studies from a range of fields in science and engineering are poised to guide future investigations in this area. This review summarizes recent studies on bacteria-surface interactions, discusses mechanisms of surface sensing and consequences of cell attachment, provides an overview of surfaces that have been used in bacterial studies, and highlights unanswered questions in this field.

Tuson HH; Weibel DB

2013-05-01

318

Receptor-transporter interactions of canonical ATP-binding cassette import systems in prokaryotes.  

Science.gov (United States)

ATP-binding cassette (ABC) transport systems mediate the translocation of solutes across biological membranes at the expense of ATP. They share a common modular architecture comprising two pore-forming transmembrane domains and two nucleotide binding domains. In prokaryotes, ABC transporters are involved in the uptake of a large variety of chemicals, including nutrients, osmoprotectants and signal molecules. In pathogenic bacteria, some ABC importers are virulence factors. Canonical ABC import systems require an additional component, a substrate-specific receptor or binding protein for function. Interaction of the liganded receptor with extracytoplasmic loop regions of the transmembrane domains initiate the transport cycle. In this review we summarize the current knowledge on receptor-transporter interplay provided by crystal structures as well as by biochemical and biophysical means. In particular, we focus on the maltose/maltodextrin transporter of enterobacteria and the transporters for positively charged amino acids from the thermophile Geobacillus stearothermophilus and Salmonella enterica serovar Typhimurium. PMID:21561685

Schneider, Erwin; Eckey, Viola; Weidlich, Daniela; Wiesemann, Nicole; Vahedi-Faridi, Ardeshir; Thaben, Paul; Saenger, Wolfram

2011-05-10

319

Receptor-transporter interactions of canonical ATP-binding cassette import systems in prokaryotes.  

UK PubMed Central (United Kingdom)

ATP-binding cassette (ABC) transport systems mediate the translocation of solutes across biological membranes at the expense of ATP. They share a common modular architecture comprising two pore-forming transmembrane domains and two nucleotide binding domains. In prokaryotes, ABC transporters are involved in the uptake of a large variety of chemicals, including nutrients, osmoprotectants and signal molecules. In pathogenic bacteria, some ABC importers are virulence factors. Canonical ABC import systems require an additional component, a substrate-specific receptor or binding protein for function. Interaction of the liganded receptor with extracytoplasmic loop regions of the transmembrane domains initiate the transport cycle. In this review we summarize the current knowledge on receptor-transporter interplay provided by crystal structures as well as by biochemical and biophysical means. In particular, we focus on the maltose/maltodextrin transporter of enterobacteria and the transporters for positively charged amino acids from the thermophile Geobacillus stearothermophilus and Salmonella enterica serovar Typhimurium.

Schneider E; Eckey V; Weidlich D; Wiesemann N; Vahedi-Faridi A; Thaben P; Saenger W

2012-04-01

320

Impact of an oil-based lubricant on the effectiveness of the sterilization processes .  

UK PubMed Central (United Kingdom)

Surgical instruments, including hinged instruments, were inoculated with test microorganisms (ie, methicillin-resistant Staphylococcus aureus, approximately 2 x 10(6) colony-forming units [cfu]; Pseudomonas aeruginosa, approximately 3 x 10(6) cfu; Escherichia coli, approximately 2 x 10(5) cfu; vancomycin-resistant enterococci, 1 x 10(5) cfu; Geobacillus stearothermophilus spores, 2 x 10(5) cfu or more; or Bacillus atrophaeus spores, 9 x 10(4) cfu or more), coated with an oil-based lubricant (hydraulic fluid), subjected to a sterilization process, and then samples from the instruments were cultured. We found that the oil-based lubricant did not alter the effectiveness of the sterilization process because high numbers of clinically relevant bacteria and standard test spores (which are relatively resistant to the sterilization process) were inactivated.

Rutala WA; Gergen MF; Weber DJ

2008-01-01

 
 
 
 
321

Impact of an oil-based lubricant on the effectiveness of the sterilization processes .  

Science.gov (United States)

Surgical instruments, including hinged instruments, were inoculated with test microorganisms (ie, methicillin-resistant Staphylococcus aureus, approximately 2 x 10(6) colony-forming units [cfu]; Pseudomonas aeruginosa, approximately 3 x 10(6) cfu; Escherichia coli, approximately 2 x 10(5) cfu; vancomycin-resistant enterococci, 1 x 10(5) cfu; Geobacillus stearothermophilus spores, 2 x 10(5) cfu or more; or Bacillus atrophaeus spores, 9 x 10(4) cfu or more), coated with an oil-based lubricant (hydraulic fluid), subjected to a sterilization process, and then samples from the instruments were cultured. We found that the oil-based lubricant did not alter the effectiveness of the sterilization process because high numbers of clinically relevant bacteria and standard test spores (which are relatively resistant to the sterilization process) were inactivated. PMID:18171191

Rutala, William A; Gergen, Maria F; Weber, David J

2008-01-01

322

Lose a Million (Bacteria) Game  

Science.gov (United States)

... Lose a Million (Bacteria) Game. "Lose a Million (Bacteria)" is a fun game based on the popular TV game show, "Who wants to be a Millionaire.". ... More results from www.fda.gov/food/foodscienceresearch/toolsmaterials

323

Backbone resonance assignments of the homotetrameric (48 kD) copper sensor CsoR from Geobacillus thermodenitrificans in the apo- and Cu(I)-bound states: insights into copper-mediated allostery.  

Science.gov (United States)

Prokaryotes are highly susceptible to exogenous copper and employ metalloregulatory proteins to control the intracellular concentration. CsoR (copper-sensitive operon repressor) is one such protein that represses transcription of a Cu(I)-effluxing ATPase in its apo form. Cu(I)-binding leads to transcriptional derepression and cellular copper resistance. Herein, we present substantially complete backbone (H(N), N, C', C?, C?) resonance assignments of tetrameric (48 kD) Geobacillus thermodenitrificans (Gt) CsoR in its apo- and Cu(I)-saturated states. These data provide the first spectroscopic evidence that Cu(I)-binding induces an interruption in the long ?2 helix of CsoR. PMID:23001947

Coyne, H Jerome; Giedroc, David P

2012-09-22

324

Thermophilic bacteria in cool temperate soils: are they metabolically active or continually added by global atmospheric transport?  

UK PubMed Central (United Kingdom)

Thermophilic soil geobacilli isolated from cool temperate geographical zone environments have been shown to be metabolically inactive under aerobic conditions at ambient temperatures (-5 to 25 degrees C). It is now confirmed that a similar situation exists for their anaerobic denitrification activity. It is necessary therefore to determine the mechanisms that sustain the observed significant viable populations in these soils. Population analysis of thermophiles in rainwater and air samples has shown different species compositions which support the view that long distance global transport and deposition in rainwater is a possible source of replenishment of the soil thermophile populations. Survival experiments using a representative Geobacillus isolate have indicated that while cells lose viability rapidly at most temperatures, populations can increase only when the temperature allows growth to take place at a rate which exceeds death rate. Long term (9-month) experiments at 4 degrees C show population increases which can be accounted for by very slow growth rates complemented by negligible death rates. These results are interpreted in the context of current hypotheses on the biogeography patterns of bacteria.

Marchant R; Franzetti A; Pavlostathis SG; Tas DO; Erdbrugger I; Unyayar A; Mazmanci MA; Banat IM

2008-04-01

325

Lipoprotein sorting in bacteria.  

UK PubMed Central (United Kingdom)

Bacterial lipoproteins are synthesized as precursors in the cytoplasm and processed into mature forms on the cytoplasmic membrane. A lipid moiety attached to the N terminus anchors these proteins to the membrane surface. Many bacteria are predicted to express more than 100 lipoproteins, which play diverse functions on the cell surface. The Lol system, composed of five proteins, catalyzes the localization of Escherichia coli lipoproteins to the outer membrane. Some lipoproteins play vital roles in the sorting of other lipoproteins, lipopolysaccharides, and ?-barrel proteins to the outer membrane. On the basis of results from biochemical, genetic, and structural studies, we discuss the biogenesis of lipoproteins in bacteria, their importance in cellular functions, and the molecular mechanisms underlying efficient sorting of hydrophobic lipoproteins to the outer membrane through the hydrophilic periplasm.

Okuda S; Tokuda H

2011-01-01

326

Lipoprotein sorting in bacteria.  

Science.gov (United States)

Bacterial lipoproteins are synthesized as precursors in the cytoplasm and processed into mature forms on the cytoplasmic membrane. A lipid moiety attached to the N terminus anchors these proteins to the membrane surface. Many bacteria are predicted to express more than 100 lipoproteins, which play diverse functions on the cell surface. The Lol system, composed of five proteins, catalyzes the localization of Escherichia coli lipoproteins to the outer membrane. Some lipoproteins play vital roles in the sorting of other lipoproteins, lipopolysaccharides, and ?-barrel proteins to the outer membrane. On the basis of results from biochemical, genetic, and structural studies, we discuss the biogenesis of lipoproteins in bacteria, their importance in cellular functions, and the molecular mechanisms underlying efficient sorting of hydrophobic lipoproteins to the outer membrane through the hydrophilic periplasm. PMID:21663440

Okuda, Suguru; Tokuda, Hajime

2011-01-01

327

Reanimation of Ancient Bacteria  

Energy Technology Data Exchange (ETDEWEB)

Recent highly publicized experiments conducted on salt crystals taken from the Permian Salado Formation in Southeastern New Mexico have shown that some ancient crystals contain viable microorganisms trapped within fluid inclusions. Stringent geological and microbiological selection criteria were used to select crystals and conduct all sampling. This talk will focus on how each of these lines of data support the conclusion that such isolated bacteria are as old as the rock in which they are trapped. In this case, the isolated microbes are salt tolerant bacilli that grow best in media containing 8% NaCl, and respond to concentrated brines by forming spores. One of the organisms is phylogenetically related to several bacilli, but does have several unique characteristics. This talk will trace the interdisciplinary data and procedures supporting these discoveries, and describe the various isolated bacteria.

Vreeland, Russell H. (West Chester University)

2002-01-09

328

Manufacture of Probiotic Bacteria  

Science.gov (United States)

Lactic acid bacteria (LAB) have been used for many years as natural biopreservatives in fermented foods. A small group of LAB are also believed to have beneficial health effects on the host, so called probiotic bacteria. Probiotics have emerged from the niche industry from Asia into European and American markets. Functional foods are one of the fastest growing markets today, with estimated growth to 20 billion dollars worldwide by 2010 (GIA, 2008). The increasing demand for probiotics and the new food markets where probiotics are introduced, challenges the industry to produce high quantities of probiotic cultures in a viable and stable form. Dried concentrated probiotic cultures are the most convenient form for incorporation into functional foods, given the ease of storage, handling and transport, especially for shelf-stable functional products. This chapter will discuss various aspects of the challenges associated with the manufacturing of probiotic cultures.

Muller, J. A.; Ross, R. P.; Fitzgerald, G. F.; Stanton, C.

329

Glacial lake hides bacteria  

Science.gov (United States)

This article highlights the published work of a geomicrobiology research team led by Eric Gaidos from the University of Hawaii and Brian Lanoil, from the University of California, Riverside. This group reports the identification of bacteria from an Icelandic sub-glacial lake, and how the collection and description of these microorganisms immured within glacial ice and sub-surface water serve as a model in the search for extra-terrestrial life.

Peplow, Mark; Online, Bioed

330

Antibiotic Resistant Bacteria  

Science.gov (United States)

This week's Topic In Depth is about antibiotic resistant bacteria.The first site is a recent news report from BBC news (1) that describes some recent research on resistant strains of two "of the world's most dangerous bacteria. Next is a Centers for Disease Control (CDC) page (2) with a brief background on antibiotic resistance and how to prevent it. A much more in-depth report is provided by the Select Committee on Science and Technology of the British House of Lords (3). There has been some public concern over the use of antibiotic resistant bacteria strains as markers in genetically modified food crops. The next two resources present information specific to this topic. The first is from the European Federation of Biotechnology (4), and the second is a shorter report from the Council for Biotechnology Information (5). The Alliance for the Prudent Use of Antibiotics (6) has a consumer and patient information section that explains what individuals can do to help prevent the problem from increasing. Readers who need a brief primer on antibiotics may appreciate this Web site from the University of Edinburgh (7). The last site is a "bugs in the news" feature from the University of Kansas (8), which is an easy-to-read explanation of "what the heck" antibiotic resistance is.

Lee, Amy.

2002-01-01

331

Biocide tolerance in bacteria.  

UK PubMed Central (United Kingdom)

Biocides have been employed for centuries, so today a wide range of compounds showing different levels of antimicrobial activity have become available. At the present time, understanding the mechanisms of action of biocides has also become an important issue with the emergence of bacterial tolerance to biocides and the suggestion that biocide and antibiotic resistance in bacteria might be linked. While most of the mechanisms providing antibiotic resistance are agent specific, providing resistance to a single antimicrobial or class of antimicrobial, there are currently numerous examples of efflux systems that accommodate and, thus, provide tolerance to a broad range of structurally unrelated antimicrobials, both antibiotics and biocides. If biocide tolerance becomes increasingly common and it is linked to antibiotic resistance, not only resistant (even multi-resistant) bacteria could be passed along the food chain, but also there are resistance determinants that can spread and lead to the emergence of new resistant microorganisms, which can only be detected and monitored when the building blocks of resistance traits are understood on the molecular level. This review summarizes the main advances reached in understanding the mechanism of action of biocides, the mechanisms of bacterial resistance to both biocides and antibiotics, and the incidence of biocide tolerance in bacteria of concern to human health and the food industry.

Ortega Morente E; Fernández-Fuentes MA; Grande Burgos MJ; Abriouel H; Pérez Pulido R; Gálvez A

2013-03-01

332

Comparative study on disinfection potency of spore forming bacteria by electron-beam irradiation and gamma-ray irradiation  

International Nuclear Information System (INIS)

[en] Along with gamma-ray irradiation, electron-beam irradiation (EB) is a method to disinfect microorganisms which cause food decomposition and food-poisoning. The present study was undertaken to compare sterilization efficacy of EB and gamma-ray irradiation on bacterial spores and vegetative cells under various conditions. Spores of Bacillus pumilus, a marker strain for irradiation study, and Bacillus stearothermophilus known as a thermophilic bacteria were irradiated by electron-beam and gamma-ray separately at irradiation dose of 0 to 10 kGy on combination of wet/dry and aerobic/anaerobic conditions. Sterilization effect of irradiation on spores was evaluated by colony counting on agar plates. Results showed that both EB and gamma-ray irradiation gave sufficient sterilization effect on spores, and the sterilization effect increased exponentially with irradiation dose. The sterilization effect of gamma-ray irradiation was higher than that of EB in all cases. Higher disinfection effect was observed under aerobic condition. The present study suggests that oxygen supply in EB is more important than gamma-ray irradiation. No results suggesting that chlorine ion at 0.1 ppm (as available chlorine concentration) enhanced the sterilization efficacy of either EB or gamma-ray irradiation was obtained under any conditions examined. (author)

1990-01-01

333

Comparative study on disinfection potency of spore forming bacteria by electron-beam irradiation and gamma-ray irradiation  

Energy Technology Data Exchange (ETDEWEB)

Along with gamma-ray irradiation, electron-beam irradiation (EB) is a method to disinfect microorganisms which cause food decomposition and food-poisoning. The present study was undertaken to compare sterilization efficacy of EB and gamma-ray irradiation on bacterial spores and vegetative cells under various conditions. Spores of Bacillus pumilus, a marker strain for irradiation study, and Bacillus stearothermophilus known as a thermophilic bacteria were irradiated by electron-beam and gamma-ray separately at irradiation dose of 0 to 10 kGy on combination of wet/dry and aerobic/anaerobic conditions. Sterilization effect of irradiation on spores was evaluated by colony counting on agar plates. Results showed that both EB and gamma-ray irradiation gave sufficient sterilization effect on spores, and the sterilization effect increased exponentially with irradiation dose. The sterilization effect of gamma-ray irradiation was higher than that of EB in all cases. Higher disinfection effect was observed under aerobic condition. The present study suggests that oxygen supply in EB is more important than gamma-ray irradiation. No results suggesting that chlorine ion at 0.1 ppm (as available chlorine concentration) enhanced the sterilization efficacy of either EB or gamma-ray irradiation was obtained under any conditions examined. (author).

Takizawa, Hironobu; Suzuki, Satoru; Suzuki, Tetsuya; Takama, Kozo (Hokkaido Univ., Hakodate (Japan). Faculty of Fisheries); Hayashi, Toru; Yasumoto, Kyoden

1990-10-01

334

Performance of bacteria filters.  

UK PubMed Central (United Kingdom)

Several kinds and brands of bacteria filters are commercially available for use in anesthesia and respiratory therapy applications. Clinical experience of high airflow resistance, ruptured media, failure to retain visible dust particles, and lack of consistent performance statements or warranties by manufacturers about their bacteria filters prompted a study of the performance of 13 different filters. The filters were challenged by mineral oil droplets, Serratia marcescens and Excherichia coli bacteriophages T4 and T7, tobacco smoke, nebulized india ink, dioctylphthalate smoke (DOP), and water. Results showed that viable bacterial passed through some filters, many filters were unable to retain visible ink or tobacco smoke particles, and resistance to airflow was increased two-fold or more in many filters when the filters were laden with 10 ml of water. Conflicting data resulted from two different types of DOP testing machines. There was a wide variation in performance among the different brands of filters; variable results also were seen within a given brand. Five brands of filters met the federal DOP standards for HEPA filters, but the wide variation in DOP testing results with two different kinds of DOP machines indicates a need for better standards. The DOP 0.3-micron bubble test is the most readily available nontoxic test to rate filtration efficiency; however, the DOP efficiency rating cannot be used to equate equivalent performance against infectious organisms.

Dryden GE; Dryden SR; Brown DG; Schatzle KC; Godzeski C

1980-11-01

335

Performance of bacteria filters.  

Science.gov (United States)

Several kinds and brands of bacteria filters are commercially available for use in anesthesia and respiratory therapy applications. Clinical experience of high airflow resistance, ruptured media, failure to retain visible dust particles, and lack of consistent performance statements or warranties by manufacturers about their bacteria filters prompted a study of the performance of 13 different filters. The filters were challenged by mineral oil droplets, Serratia marcescens and Excherichia coli bacteriophages T4 and T7, tobacco smoke, nebulized india ink, dioctylphthalate smoke (DOP), and water. Results showed that viable bacterial passed through some filters, many filters were unable to retain visible ink or tobacco smoke particles, and resistance to airflow was increased two-fold or more in many filters when the filters were laden with 10 ml of water. Conflicting data resulted from two different types of DOP testing machines. There was a wide variation in performance among the different brands of filters; variable results also were seen within a given brand. Five brands of filters met the federal DOP standards for HEPA filters, but the wide variation in DOP testing results with two different kinds of DOP machines indicates a need for better standards. The DOP 0.3-micron bubble test is the most readily available nontoxic test to rate filtration efficiency; however, the DOP efficiency rating cannot be used to equate equivalent performance against infectious organisms. PMID:10315105

Dryden, G E; Dryden, S R; Brown, D G; Schatzle, K C; Godzeski, C

1980-11-01

336

Platelet Interaction with Bacteria: II. Fate of the Bacteria  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Several common strains of bacteria have been studied to determine their influence on human and rabbit platelets in vitro. Bacteria at a nominal ratio of 1:1 were added to platelets in their native plasma or to platelets in a balanced salt solution. The platelet-bacterial interaction was examined by ...

Clawson, C. C.; White, James G.

337

Bacteriophages of methanotrophic bacteria  

Energy Technology Data Exchange (ETDEWEB)

Bacteriophages of methanotrophic bacteria have been found in 16 out of 88 studied samples (underground waters, pond water, soil, gas and oil installation waters, fermentor cultural fluids, bacterial paste, and rumen of cattle) taken in different geographic zones of the Soviet Union. Altogether, 23 phage strains were isolated. By fine structure, the phages were divided into two types (with very short or long noncontractile tails); by host range and serological properties, they fell into three types. All phages had guanine- and cytosine-rich double-stranded deoxyribonucleic acid consisting of common nitrogen bases. By all of the above-mentioned properties, all phages within each of the groups were completely identical to one another, but differed from phages of other groups.

Tyutikow, F.M. (All-Union Research Inst. for Genetics and Selection of Industrial Microorganisms, Moscow, USSR); Bespalova, I.A.; Rebentish, B.A.; Aleksandrushkina, N.N.; Krivisky, A.S.

1980-10-01

338

Arsenic-tolerant, arsenite-oxidising bacterial strains in the contaminated soils of West Bengal, India.  

Science.gov (United States)

As biological agents represent an affordable alternative to costly metal decontamination technologies, we isolated arsenic (As) oxidising bacteria from the As-contaminated soils of West Bengal, India. These strains were closely related to various species of Bacillus and Geobacillus based on their 16S rRNA gene sequences. They were found to be hyper-resistant to both As(V) (167-400mM) and As(III) (16-47mM). Elevated rates of As(III) oxidation (278-1250?Mh(-1)) and arsenite oxidase activity (2.1-12.5nMmin(-1)mg(-1) protein) were observed in these isolates. Screening identified four strains as superior As-oxidisers. Among them, AMO-10 completely (100%) oxidised 30mM of As(III) within 24h. The presence of the aoxB gene was confirmed in the screened isolates. Phylogenetic tree construction based on the aoxB sequence revealed that two strains, AGO-S5 and AGH-02, clustered with Achromobacter and Variovorax, whereas the other two (AMO-10 and ADP-25) remained unclustered. The increased rate of As(III) oxidation by these native strains might be exploited for the remediation of As in contaminated environments. Notably, this study presents the first correlation regarding the presence of the aoxB gene and As(III) oxidation ability in Geobacillus stearothermophilus. PMID:23876545

Majumder, Aparajita; Bhattacharyya, K; Bhattacharyya, S; Kole, S C

2013-07-20

339

[Isolation of viscous-oil degrading microorganism and biodegradation to resin].  

UK PubMed Central (United Kingdom)

OBJECTIVE: The aim of this study was to isolate bacterial strains with high-efficiency to degrade resins. METHODS: We used resin-plate to isolate resin-degrading bacteria from the formation water of Nanbao35-2 oil field, China National Offshore Oil Corporation. The morphological properties and the sequence homology of 16S rRNA were used to identify the strains. The changes of four fractions contents and the infrared spectrometry of the heavy oil were used to analyze the degradation properties. RESULTS: Four strains, Q4, QB9, QB26 and QB36, were isolated using resin as the sole carbon source. Based on the high sequence similarities (more than 99%) of 16S rDNA sequences analysis. These strains were identified as member of the Petrobacter sp., Geobacillus stearothermophilus, Bacillus licheniformis, Geobacillus pallidus, respectively. QB26, a Bacillus licheniformis, was the most efficient strain, it can grow well under anaerobic conditions, emulsify heavy oil well, and degrade resin and asphaltene in heavy oil. The relative content of saturated hydrocarbons in heavy oil increase after degradation, and the relative content of resin and asphaltene in heavy oil decreased 5.1% and 2.7%, respectively. CONCLUSION: The strains isolated from Nanbao 35-2 oil field formation water could degrade resin and heavy oil. They have potential values in microbial enhanced oil recovery and oil pollution treatment.

Wang D; Zhang J; Qi Y; Ma T

2012-03-01

340

Arsenic-tolerant, arsenite-oxidising bacterial strains in the contaminated soils of West Bengal, India.  

UK PubMed Central (United Kingdom)

As biological agents represent an affordable alternative to costly metal decontamination technologies, we isolated arsenic (As) oxidising bacteria from the As-contaminated soils of West Bengal, India. These strains were closely related to various species of Bacillus and Geobacillus based on their 16S rRNA gene sequences. They were found to be hyper-resistant to both As(V) (167-400mM) and As(III) (16-47mM). Elevated rates of As(III) oxidation (278-1250?Mh(-1)) and arsenite oxidase activity (2.1-12.5nMmin(-1)mg(-1) protein) were observed in these isolates. Screening identified four strains as superior As-oxidisers. Among them, AMO-10 completely (100%) oxidised 30mM of As(III) within 24h. The presence of the aoxB gene was confirmed in the screened isolates. Phylogenetic tree construction based on the aoxB sequence revealed that two strains, AGO-S5 and AGH-02, clustered with Achromobacter and Variovorax, whereas the other two (AMO-10 and ADP-25) remained unclustered. The increased rate of As(III) oxidation by these native strains might be exploited for the remediation of As in contaminated environments. Notably, this study presents the first correlation regarding the presence of the aoxB gene and As(III) oxidation ability in Geobacillus stearothermophilus.

Majumder A; Bhattacharyya K; Bhattacharyya S; Kole SC

2013-10-01

 
 
 
 
341

Testing antimicrobials against biofilm bacteria.  

UK PubMed Central (United Kingdom)

Standard laboratory methods are needed to assess the efficacy of antimicrobial agents that are applied to biofilm bacteria. Existing standard suspension tests and dried surface tests show much greater efficacy than antimicrobial agents applied to biofilms. The greater resistance of biofilm bacteria to antimicrobial agents can be attributed to a number of interacting factors, including reaction and diffusion processes that limit an agent's accessibility to bacteria, phenotypic changes in biofilm bacteria caused by stress, and adaptation of the bacteria. Because biofilm systems are so diverse, a variety of new biofilm tests with features that differ in important ways from existing tests will ultimately be required. For example, the biofilm test apparatus may include a pump and a continuous-flow stirred tank reactor. This report provides an overview of biofilm testing and suggests a strategy for creating standard test methods.

Hamilton MA

2002-03-01

342

Sterilization of single-use helical stone baskets: an experimental study  

Directory of Open Access Journals (Sweden)

Full Text Available Objectives: To experimentally evaluate the efficacy of a standard sterilization protocol employed during reuse of disposable helical stone baskets. Methods: Study performed on 20 helical stone baskets: 10 were used in the initial validation process, contaminated with Escherichia coli ATCC 25922 and imprinted on Müeller-Hinton media; 10 catheters were contaminated with Geobacillus stearothermophilus ATCC 7953, processed, inoculated in TSB and incubated in a water bath at a temperature of 55ºC. Bacterial growth was evaluated after 1, 3, 5 and 7 days. After sterilization, stone baskets were also opened and closed 40 times to check for functional problems. All plastic and basket parts were carefully checked for damages. Results: After the 72-hour incubation period, there was growth of E. coli ATCC 25922 in 100% of imprints. After the sterilization process and up to 7 days incubation period on a blood agar plate, there was no growth of G. stearothermophilus ATCC 7953 or any other bacteria. There were no functional problems or damage to baskets after the sterilization process. Conclusion: The ethylene oxide system is efficacious and safe for sterilization of disposable helical stone baskets. However, further clinical studies are required and should provide more safety information.

Fernando Korkes; Alex Menezes; Cely Barreto da Silva; Roni de Carvalho Fernandes; Marjo Deninson Cardenuto Perez

2011-01-01

343

Clean-Test self-contained biological indicator performance used to steam sterilization process validation.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Clean-Test self-contained biological indicators are used to monitor steam sterilization cycles. Each biological indicator has a minimum population of 105 or 106 UFC bacterial spores of Geobacillus stearothermophilus ATCC 7953. The pu...

Thiago José Pelissari; Heron Oliveira dos Santos Lima; Mirela Vanin dos Santos Lima

344

Bacteria Growth Inquiry: Bodily Bacteria and Healthy Hygiene Habits  

Science.gov (United States)

In this inquiry activity, students generate investigable questions to explore the link between hygiene/cleanliness and bacteria growth/population. The students will present their conclusions, and video clips containing additional information will be discussed.

345

L-Ribose Production from L-Arabinose by Immobilized Recombinant Escherichia coli Co-expressing the L-Arabinose Isomerase and Mannose-6-Phosphate Isomerase Genes from Geobacillus thermodenitrificans.  

UK PubMed Central (United Kingdom)

L-Ribose is an important precursor for antiviral agents, and thus its high-level production is urgently demanded. For this aim, immobilized recombinant Escherichia coli cells expressing the L-arabinose isomerase and variant mannose-6-phosphate isomerase genes from Geobacillus thermodenitrificans were developed. The immobilized cells produced 99 g/l L-ribose from 300 g/l L-arabinose in 3 h at pH 7.5 and 60 °C in the presence of 1 mM Co(2+), with a conversion yield of 33 % (w/w) and a productivity of 33 g/l/h. The immobilized cells in the packed-bed bioreactor at a dilution rate of 0.2 h(-1) produced an average of 100 g/l L-ribose with a conversion yield of 33 % and a productivity of 5.0 g/l/h for the first 12 days, and the operational half-life in the bioreactor was 28 days. Our study is first verification for L-ribose production by long-term operation and feasible for cost-effective commercialization. The immobilized cells in the present study also showed the highest conversion yield among processes from L-arabinose as the substrate.

Kim KR; Seo ES; Oh DK

2013-09-01

346

Cytokinesis in bacteria.  

UK PubMed Central (United Kingdom)

Work on two diverse rod-shaped bacteria, Escherichia coli and Bacillus subtilis, has defined a set of about 10 conserved proteins that are important for cell division in a wide range of eubacteria. These proteins are directed to the division site by the combination of two negative regulatory systems. Nucleoid occlusion is a poorly understood mechanism whereby the nucleoid prevents division in the cylindrical part of the cell, until chromosome segregation has occurred near midcell. The Min proteins prevent division in the nucleoid-free spaces near the cell poles in a manner that is beginning to be understood in cytological and biochemical terms. The hierarchy whereby the essential division proteins assemble at the midcell division site has been worked out for both E. coli and B. subtilis. They can be divided into essentially three classes depending on their position in the hierarchy and, to a certain extent, their subcellular localization. FtsZ is a cytosolic tubulin-like protein that polymerizes into an oligomeric structure that forms the initial ring at midcell. FtsA is another cytosolic protein that is related to actin, but its precise function is unclear. The cytoplasmic proteins are linked to the membrane by putative membrane anchor proteins, such as ZipA of E. coli and possibly EzrA of B. subtilis, which have a single membrane span but a cytoplasmic C-terminal domain. The remaining proteins are either integral membrane proteins or transmembrane proteins with their major domains outside the cell. The functions of most of these proteins are unclear with the exception of at least one penicillin-binding protein, which catalyzes a key step in cell wall synthesis in the division septum.

Errington J; Daniel RA; Scheffers DJ

2003-03-01

347

Cytokinesis in bacteria.  

Science.gov (United States)

Work on two diverse rod-shaped bacteria, Escherichia coli and Bacillus subtilis, has defined a set of about 10 conserved proteins that are important for cell division in a wide range of eubacteria. These proteins are directed to the division site by the combination of two negative regulatory systems. Nucleoid occlusion is a poorly understood mechanism whereby the nucleoid prevents division in the cylindrical part of the cell, until chromosome segregation has occurred near midcell. The Min proteins prevent division in the nucleoid-free spaces near the cell poles in a manner that is beginning to be understood in cytological and biochemical terms. The hierarchy whereby the essential division proteins assemble at the midcell division site has been worked out for both E. coli and B. subtilis. They can be divided into essentially three classes depending on their position in the hierarchy and, to a certain extent, their subcellular localization. FtsZ is a cytosolic tubulin-like protein that polymerizes into an oligomeric structure that forms the initial ring at midcell. FtsA is another cytosolic protein that is related to actin, but its precise function is unclear. The cytoplasmic proteins are linked to the membrane by putative membrane anchor proteins, such as ZipA of E. coli and possibly EzrA of B. subtilis, which have a single membrane span but a cytoplasmic C-terminal domain. The remaining proteins are either integral membrane proteins or transmembrane proteins with their major domains outside the cell. The functions of most of these proteins are unclear with the exception of at least one penicillin-binding protein, which catalyzes a key step in cell wall synthesis in the division septum. PMID:12626683

Errington, Jeffery; Daniel, Richard A; Scheffers, Dirk-Jan

2003-03-01

348

Bacteriophages of methanotrophic bacteria.  

UK PubMed Central (United Kingdom)

Bacteriophages of methanotrophic bacteria have been found in 16 out of 88 studied samples (underground waters, pond water, soil, gas and oil installation waters, fermentor cultural fluids, bacterial paste, and rumen of cattle) taken in different geographic zones of the Soviet Union. Altogether, 23 phage strains were isolated: 10 strains that specifically lysed only Methylosinus sporium strains, 2 strains that each lysed 1 of 5 Methylosinus trichosporium strains studied, and 11 strains that lysed Flavobacterium gasotypicum and, at the same time, 1 M. sporium strain. By fine structure, the phages were divided into two types (with very short or long noncontractile tails); by host range and serological properties, they fell into three types. One-step growth characteristics of the phages differed only slightly; the latent period varied from 6 to 8 h, the rise period varied from 4 to 6 h, and the average burst size was 100. All phages had guanine- and cytosine-rich double-stranded deoxyribonucleic acid consisting of common nitrogen bases. The molecular mass of the deoxyribonucleic acid as determined by restriction endonuclease analysis was 29.4 X 10(6) for M. sporium phages and 44 X 10(6) for F. gasotypicum phages. By all of the above-mentioned properties, all phages within each of the groups were completely identical to one another, but differed from phages of other groups. Bacteriophages lysing M. sporium and M. trichosporium GB2 were identical to phages M1 and M4, respectively, which were isolated earlier in the German Democratic Republic on the same methanotrophic species.

Tyutikov FM; Bespalova IA; Rebentish BA; Aleksandrushkina NN; Krivisky AS

1980-10-01

349

Where Bacteria and Languages Concur  

Science.gov (United States)

Access to the article is free, however registration and sign-in are required. Genetic data from human gastric bacteria provide independent support for a linguistic analysis of Pacific population dispersals.

Colin Renfrew (University of Cambridge;McDonald Institute for Archaeological Research)

2009-01-23

350

Geobiology of Marine Magnetotactic Bacteria.  

Science.gov (United States)

Magnetotactic bacteria (MTB) biomineralize intracellular membrane- bound crystals of magnetite (Fe304) or greigite (Fe3S4), and are abundant in the suboxic to anoxic zones of stratified marine environments worldwide. Their population densities (up to 10(5...

S. L. Simmons

2006-01-01

351

Thymidine kinase diversity in bacteria  

DEFF Research Database (Denmark)

Thymidine kinases (TKs) appear to be almost ubiquitous and are found in nearly all prokaryotes, eukaryotes, and several viruses. They are the key enzymes in thymidine salvage and activation of several anti-cancer and antiviral drugs. We show that bacterial TKs can be subdivided into 2 groups. The TKs from Gram-positive bacteria are more closely related to the eukaryotic TK1 enzymes than are TKs from Gram-negative bacteria.

Sandrini, Michael; Clausen, A.R.

2006-01-01

352

Bioreporter bacteria for landmine detection  

Energy Technology Data Exchange (ETDEWEB)

Landmines (and other UXO) gradually leak explosive chemicals into the soil at significant concentrations. Bacteria, which have adapted to scavenge low concentrations of nutrients, can detect these explosive chemicals. Uptake of these chemicals results in the triggering of specific bacterial genes. The authors have created genetically recombinant bioreporter bacteria that detect small concentrations of energetic chemicals. These bacteria are genetically engineered to produce a bioluminescent signal when they contact specific explosives. A gene for a brightly fluorescent compound can be substituted for increased sensitivity. By finding the fluorescent bacteria, you find the landmine. Detection might be accomplished using stand-off illumination of the minefield and GPS technology, which would result in greatly reduced risk to the deminers. Bioreporter technology has been proven at the laboratory scale, and will be tested under field conditions in the near future. They have created a bacterial strain that detects sub-micromolar concentrations of o- and p-nitrotoluene. Related bacterial strains were produced using standard laboratory protocols, and bioreporters of dinitrotoluene and trinitrotoluene were produced, screening for activity with the explosive compounds. Response time is dependent on the growth rate of the bacteria. Although frill signal production may require several hours, the bacteria can be applied over vast areas and scanned quickly, producing an equivalent detection speed that is very fast. This technology may be applicable to other needs, such as locating buried explosives at military and ordnance/explosive manufacturing facilities.

Burlage, R.S. [Oak Ridge National Lab., TN (United States); Youngblood, T. [Frisby Technologies, Aiken, SC (United States); Lamothe, D. [American Technologies, Inc., Huntsville, AL (United States). Ordnance/Explosives Environmental Services Div.

1998-04-01

353

Quorum Sensing- Communication Between Bacteria  

Directory of Open Access Journals (Sweden)

Full Text Available The latest discoveries in the field of microbiology have proved that bacteriacommunicate between each other. It is common knowledge that bacterial diseasessuch as cholera, anthrax, meningitis, and many others are among the deadliestin the world. It may be the case, however, that bacteria cannot cause an illnessin small quantities. Only when there are a sufficient number of them can theyact. Some, Vibrio Fischeri or Vibrio Harveyi, can glow in the dark. Others,like Pseudomonas Aeruginosa, form biofilms on the surface of human organs, andattack virulently those organs multiplying with tremendous speed, making itpractically impossible for antibiotics to interfere.What is quorum sensing? Bacteria produce and release chemical signals - autoinducers- in search of similar cells in their close surroundings. This is also called"cell-cell communication." Other bacteria release the same autoinducers in response.One-cell organisms in effect become multi-cellular organisms and can act together.People or animals may have bacteria that cause some serious infectious diseasesand yet not be infected unless the number of bacteria is large enough; thatis to say, unless the quorum has been reached. Even though this phenomenon hasbeen known to scientists since the 1960s, only now they are able to study itin detail. This new branch of microbiology, quorum sensing, discovered by BonnyBassler, professor of microbiology from Princeton, is dedicated to studyingthis phenomenon.

Mrs. Lakshmi Sivasubramaniam; Madhumathi Seshadri

2005-01-01

354

Periodontitis, periodontopathic bacteria and lactoferrin.  

UK PubMed Central (United Kingdom)

Lactoferrin (LF) is a component of saliva and is suspected to be a defense factor against oral pathogens including Streptococcus mutans and Candida albicans. Periodontitis is a very common oral disease caused by periodontopathic bacteria. Antimicrobial activities and other biological effects of LF against representative periodontopathic bacteria, Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, and Prevotella intermedia, have been widely studied. Association of polymorphisms in LF with incidence of aggressive periodontitis and the role of LF in the gingival crevicular fluid as a marker of periodontitis severity have also been reported. Periodontopathic bacteria reside as a biofilm in supragingival and subgingival plaque. Our recent study indicated that LF exhibits antibacterial activity against planktonic forms of P. gingivalis and P. intermedia at higher concentrations, and furthermore, LF effectively inhibits biofilm formation and reduces the established biofilm of these bacteria at physiological concentrations. A small-scale clinical study indicated that oral administration of bovine LF reduces P. gingivalis and P. intermedia in the subgingival plaque of chronic periodontitis patients. LF seems to be a biofilm inhibitor of periodontopathic bacteria in vitro and in vivo.

Wakabayashi H; Kondo I; Kobayashi T; Yamauchi K; Toida T; Iwatsuki K; Yoshie H

2010-06-01

355

Genetic transfer in acidophilic bacteria  

Energy Technology Data Exchange (ETDEWEB)

There is increasing interest in the use of microorganisms to recover metals from ores, as well as to remove sulfur from coal. These so-called bioleaching processes are mediated by a number of bacteria. The best-studied of these organisms are acidophiles including Thiobacillus and Acidiphilium species. Our laboratory has focused on developing genetic strategies to allow the manipulation of acidophilic bacteria to improve and augment their utility in large scale operations. We have recently been successful in employing conjugation for interbacterial transfer of genetic information, as well as in directly transforming Acidiphilium by use of electroporation. We are now testing the properties of IncPl, IncW and IncQ plasmid vectors in Acidiphilium to determine their relative usefulness in routine manipulation of acidophiles and transfer between organisms. This study also allows us to determine the natural ability of these bacteria to transfer genetic material amongst themselves in their particular environment. 21 refs., 3 figs., 2 tabs.

Roberto, F.F.; Glenn, A.W.; Bulmer, D.; Ward, T.E.

1990-01-01

356

Bacteria as bioavailability enhancing agents  

Energy Technology Data Exchange (ETDEWEB)

Although limited bioavailability appears to be primarily a physical, i.e. mass transfer-controlled process, different recent observations indicate that organism-specific bioavailability-enhancing strategies may exist and that generalizations about the bioavailability of sorbed, solid or dissolved substrates are inappropriate. Several reports, using polycyclic aromatic hydrocarbons (PAH), indicate that sorption-limited bioavailability plays an important role in the selection of PAH-degrading bacteria and that different PAH-degrading bacteria inhabiting the same soil may be adapted to different degrees of PAH-bioavailability. In these studies, different bacterial strains or microbial assemblages were selected depending on how the compound was provided to the bacteria. This indicates that the physiology of a bacterium or its particular life style might play an important role in its adaptation to degrade HOC by providing a strategy for enhancing the compound's bioavailability. (orig.)

Wick, L.Y. [UFZ Centre for Environmental Research Leipzig-Halle, Leipzig (Germany). Dept. of Environmental Microbiology; Harms, H.

2004-07-01

357

Microrheology of swimming bacteria suspension.  

Science.gov (United States)

We study rheology of suspension of swimming bacteria Bacillus Subtilis at high concentrations. Experiments were performed in a free standing fluid film contained in a transparent chamber with adjustable Oxygen/Nitrogen ratio. The swimming velocity of bacteria is controlled by the concentration of dissolved Oxygen: it reduces to zero when Oxygen is completely replaced by Nitrogen. Macroscopic flow in a film is produced by oscillations and rotations of magnetic particles by rotating external magnetic field. To extract the effective viscosity, we measured macroscopic velocity field generated by the particles using PIV of fluorescent markers seeded to the film. We discovered that viscosity of bacterial suspension is increasing with decreasing swimming speed of bacteria due lack of Oxygen.

Sokolov, Andrey; Aronson, Igor

2009-03-01

358

Assessing chronological aging in bacteria.  

UK PubMed Central (United Kingdom)

Bacteria, which are often considered as avid reproductive organisms under constant selective pressure to utilize available nutrients to proliferate, might seem an inappropriate model to study aging. However, environmental conditions are rarely supporting the exponential growth that is most often studied in laboratories. In the wild, Escherichia coli inhabits environments of relative nutritional paucity. Not surprisingly, under such circumstances, members of an E. coli population age and progressively lose the ability to reproduce, even when environmental conditions provide such an opportunity. Here, we review the methods to study chronological aging in bacteria and some of the mechanisms that may contribute to their age-dependent loss of viability.

Gonidakis S; Longo VD

2013-01-01

359

Using bacteria to treat diseases.  

UK PubMed Central (United Kingdom)

INTRODUCTION: Mosquito-borne diseases such as malaria and dengue fever result in significant morbidity and mortality in developing countries. Vector control is often the most effective strategy to prevent disease transmission and novel methods are required to complement existing insecticide-based strategies. Biological control uses natural predators or pathogens to kill mosquitoes or reduce their capacity to transmit disease. Bacteria such as Wolbachia have been proposed to have the potential to provide effective biological control of mosquitoes. AREAS COVERED: A review of the potential role of bacteria in the control of mosquito-borne diseases highlighting the advantages and disadvantages of each strategy. In particular, a comprehensive summary of the progress made using the bacterial endosymbiont Wolbachia for dengue control. EXPERT OPINION: Pathogenic bacteria such as Bti can be used to kill mosquito larvae and several endosymbiotic bacteria such as Asaia could be genetically transformed to alter the mosquito's ability to transmit pathogens. The endosymbiotic bacterium Wolbachia has been successfully introduced into the principal vector of dengue, Aedes aegypti, and induces a variety of phenotypic effects that are predicted to reduce dengue transmission. The release of Wolbachia-infected mosquitoes has been undertaken as part of preliminary trials to determine the applied use of this bacterium for mosquito-borne disease control.

Caragata EP; Walker T

2012-06-01

360

Bacteria, it's what's for dinner.  

UK PubMed Central (United Kingdom)

While intestinal epithelial cells are known for secreting antimicrobial molecules, cell intrinsic defense mechanisms are less characterized. In this issue, Benjamin et al. (2013) demonstrate that MyD88 and autophagy within the intestinal epithelium detect invasive bacteria and prevent dissemination.

Cadwell K

2013-06-01

 
 
 
 
361

Functional genomics of intracellular bacteria.  

UK PubMed Central (United Kingdom)

During the genomic era, a large amount of whole-genome sequences accumulated, which identified many hypothetical proteins of unknown function. Rapidly, functional genomics, which is the research domain that assign a function to a given gene product, has thus been developed. Functional genomics of intracellular pathogenic bacteria exhibit specific peculiarities due to the fastidious growth of most of these intracellular micro-organisms, due to the close interaction with the host cell, due to the risk of contamination of experiments with host cell proteins and, for some strict intracellular bacteria such as Chlamydia, due to the absence of simple genetic system to manipulate the bacterial genome. To identify virulence factors of intracellular pathogenic bacteria, functional genomics often rely on bioinformatic analyses compared with model organisms such as Escherichia coli and Bacillus subtilis. The use of heterologous expression is another common approach. Given the intracellular lifestyle and the many effectors that are used by the intracellular bacteria to corrupt host cell functions, functional genomics is also often targeting the identification of new effectors such as those of the T4SS of Brucella and Legionella.

de Barsy M; Greub G

2013-07-01

362

Folate Production by Probiotic Bacteria  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Probiotic bacteria, mostly belonging to the genera Lactobacillus and Bifidobacterium, confer a number of health benefits to the host, including vitamin production. With the aim to produce folate-enriched fermented products and/or develop probiotic supplements that accomplish folate biosynthesis in v...

Maddalena Rossi; Alberto Amaretti; Stefano Raimondi

363

Photosynthetic reaction centers in bacteria  

Energy Technology Data Exchange (ETDEWEB)

The photochemistry of photosynthesis begins in complexes called reaction centers. These have become model systems to study the fundamental process by which plants and bacteria convert and store solar energy as chemical free energy. In green plants, photosynthesis occurs in two systems, each of which contains a different reaction center, working in series. In one, known as photosystem 1, oxidized nicotinamide adenine dinucleotide phosphate (NADP[sup +]) is reduced to NADPH for use in a series of dark reactions called the Calvin cycle, named for Nobel Laureate Melvin Calvin, by which carbon dioxide is converted into useful fuels such as carbohydrates and sugars. In the other half of the photosynthetic machinery of green plants, called photosystem 2, water is oxidized to produce molecular oxygen. A different form of photosynthesis occurs in photosynthetic bacteria, which typically live at the bottom of ponds and feed on organic debris. Two main types of photosynthetic bacteria exist: purple and green. Neither type liberates oxygen from water. Instead, the bacteria feed on organic media or inorganic materials, such as sulfides, which are easier to reduce or oxidize than carbon dioxide or water. Perhaps in consequence, their photosynthetic machinery is simpler than that of green, oxygen-evolving plants and their primary photochemistry is better understood.

Norris, J.R. (Argonne National Lab., IL (United States) Univ. of Chicago, IL (United States)); Schiffer, M. (Argonne National Lab., IL (United States))

1990-07-30

364

Fuzzy species among recombinogenic bacteria  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background It is a matter of ongoing debate whether a universal species concept is possible for bacteria. Indeed, it is not clear whether closely related isolates of bacteria typically form discrete genotypic clusters that can be assigned as species. The most challenging test of whether species can be clearly delineated is provided by analysis of large populations of closely-related, highly recombinogenic, bacteria that colonise the same body site. We have used concatenated sequences of seven house-keeping loci from 770 strains of 11 named Neisseria species, and phylogenetic trees, to investigate whether genotypic clusters can be resolved among these recombinogenic bacteria and, if so, the extent to which they correspond to named species. Results Alleles at individual loci were widely distributed among the named species but this distorting effect of recombination was largely buffered by using concatenated sequences, which resolved clusters corresponding to the three species most numerous in the sample, N. meningitidis, N. lactamica and N. gonorrhoeae. A few isolates arose from the branch that separated N. meningitidis from N. lactamica leading us to describe these species as 'fuzzy'. Conclusion A multilocus approach using large samples of closely related isolates delineates species even in the highly recombinogenic human Neisseria where individual loci are inadequate for the task. This approach should be applied by taxonomists to large samples of other groups of closely-related bacteria, and especially to those where species delineation has historically been difficult, to determine whether genotypic clusters can be delineated, and to guide the definition of species.

Hanage William P; Fraser Christophe; Spratt Brian G

2005-01-01

365

Killer Pigments in Bacteria: An Ecological Nightmare.  

Science.gov (United States)

Describes an alternative to teaching ecology by using bacteria to test competitor survival. Students observe a time-dependent selective killing of other unrelated bacteria by Pseudomonas aeruginosa. (SAH)

Benathen, Isaiah A.; Saccardi, Marion

2000-01-01

366

Barbecue Bliss: Keeping Bacteria at Bay  

Science.gov (United States)

... Tobacco Products Vaccines, Blood & Biologics Barbecue Bliss: Keeping Bacteria at Bay Search the Consumer Updates Section Get ... 664 KB) Summer brings out barbecue grills—and bacteria, which multiply in food faster in warm weather ...

367

Identification of a novel gene cluster participating in menaquinone (vitamin K2) biosynthesis. Cloning and sequence determination of the 2-heptaprenyl-1,4-naphthoquinone methyltransferase gene of Bacillus stearothermophilus.  

UK PubMed Central (United Kingdom)

We recently described the isolation and sequence analysis of a DNA region containing the genes of Bacillus stearothermophilus heptaprenyl diphosphate synthase, which catalyzes the synthesis of the prenyl side chain of menaquinone-7 of this bacterium. Sequence analyses revealed the presence of three open reading frames (ORFs), designated as ORF-1, ORF-2, and ORF-3, and the structural genes of the heptaprenyl diphosphate synthase were proved to consist of ORF-1 (heps-1) and ORF-3 (heps-2) (Koike-Takeshita, A., Koyama, T., Obata, S., and Ogura, K. (1995) J. Biol. Chem. 270, 18396-18400). The predicted amino acid sequence of ORF-2 (234 amino acids) contains a methyltransferase consensus sequence and shows a 22% identity with UbiG of Escherichia coli, which catalyzes S-adenosyl-L-methionine-dependent methylation of 2-octaprenyl-3-methyl-5-hydroxy-6-methoxy-1,4-benzoquinone. These pieces of information led us to identify the ORF-2 gene product. The cell-free homogenate of the transformant of E. coli with an expression vector of ORF-2 catalyzed the incorporation of S-adenosyl-L-methionine into menaquinone-8, indicating that ORF-2 encodes 2-heptaprenyl-1,4-naphthoquinone methyltransferase, which participates in the terminal step of the menaquinone biosynthesis. Thus it is concluded that the ORF-1, ORF-2, and ORF-3 genes, designated heps-1, menG, and heps-2, respectively, form another cluster involved in menaquinone biosynthesis in addition to the cluster of menB, menC, menD, and menE already identified in the Bacillus subtilis and E. coli chromosomes.

Koike-Takeshita A; Koyama T; Ogura K

1997-05-01

368

Selection-driven genome reduction in bacteria  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Gene loss by deletion is a common evolutionary process in bacteria, as exemplified by bacteria with small genomes that have evolved from bacteria with larger genomes by reductive processes. The driving force(s) for genome reduction remains unclear, and here we examined the hypothesis that gene loss ...

Koskiniemi, Sanna; Sun, Song; Berg, Otto; Andersson, Dan I.

369

Laser-Based Identification of Pathogenic Bacteria  

Science.gov (United States)

Bacteria are ubiquitous in our world. From our homes, to our work environment, to our own bodies, bacteria are the omnipresent although often unobserved companions to human life. Physicists are typically untroubled professionally by the presence of these bacteria, as their study usually falls safely outside the realm of our typical domain. In the…

Rehse, Steven J.

2009-01-01

370

Ecology of mycophagous collimonas bacteria in soil  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Bacteria belonging to the genus Collimonas consist of soil bacteria that can grow at expense of living fungal hyphae i.e. they are mycophagous. This PhD studies deals with the ecology of mycophagous bacteria in soil using collimonads as model organisms. Collimonads were found to be widely distribut...

Höppener-Ogawa, Sachie

371

Selection-Driven Gene Loss in Bacteria  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Gene loss by deletion is a common evolutionary process in bacteria, as exemplified by bacteria with small genomes that have evolved from bacteria with larger genomes by reductive processes. The driving force(s) for genome reduction remains unclear, and here we examined the hypothesis that gene loss ...

Koskiniemi, Sanna; Sun, Song; Berg, Otto G.; Andersson, Dan I.

372

Effect of Nitric Oxide on Anammox Bacteria?  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The effects of nitrogen oxides on anammox bacteria are not well known. Therefore, anammox bacteria were exposed to 3,500 ppm nitric oxide (NO) in the gas phase. The anammox bacteria were not inhibited by the high NO concentration but rather used it to oxidize additional ammonium to dinitrogen gas un...

Kartal, Boran; Tan, Nico C. G.; Van de Biezen, Erwin; Kampschreur, Marlies J.; Van Loosdrecht, Mark C. M.; Jetten, Mike S. M.

373

Magnetotactic bacteria at the geomagnetic equator  

International Nuclear Information System (INIS)

Magnetotatic bacteria are observed in freshwater and marine sediments of Fortaleza, Brazil, situated close to the geomagnetic equator. Both South-seeking and North-seeking bacteria are present in roughly equal numbers in the same samples. This observation is consistent with the hypothesis that the vertical component of the geomagnetic field selects the predominant polarity type among magnetotactic bacteria in natural environments. (Author)

1981-01-01

374

Chondroitinase-Producing Bacteria in Natural Habitats  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A search was undertaken for bacteria which degrade chondroitin sulfate in nature and to find bacteria with a usefully high rate of chondroitinase (ChSase) productivity. First, 253 ChSase-producing bacteria were obtained from aquatic and land environments in Japan by aerobic and anaerobic screening m...

Kitamikado, Manabu; Lee, Young-Zoon

375

Antibiotic-resistant bacteria in drinking water.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

We analyzed drinking water from seven communities for multiply antibiotic-resistant (MAR) bacteria (bacteria resistant to two or more antibiotics) and screened the MAR bacterial isolates obtained against five antibiotics by replica plating. Overall, 33.9% of 2,653 standard plate count bacteria from ...

Armstrong, J L; Shigeno, D S; Calomiris, J J; Seidler, R J

376

Genetics of Lactic Acid Bacteria  

Science.gov (United States)

Many meat (or fish) products, obtained by the fermentation of meat originating from various animals by the flora that naturally contaminates it, are part of the human diet since millenaries. Historically, the use of bacteria as starters for the fermentation of meat, to produce dry sausages, was thus performed empirically through the endogenous micro-biota, then, by a volunteer addition of starters, often performed by back-slopping, without knowing precisely the microbial species involved. It is only since about 50 years that well defined bacterial cultures have been used as starters for the fermentation of dry sausages. Nowadays, the indigenous micro-biota of fermented meat products is well identified, and the literature is rich of reports on the identification of lactic acid bacteria (LAB) present in many traditional fermented products from various geographical origin, obtained without the addition of commercial starters (See Talon, Leroy, & Lebert, 2007, and references therein).

Zagorec, Monique; Anba-Mondoloni, Jamila; Coq, Anne-Marie Crutz-Le; Champomier-Vergès, Marie-Christine

377

Aggregation Patterns in Stressed Bacteria  

CERN Multimedia

We study the formation of spot patterns seen in a variety of bacterial species when the bacteria are subjected to oxidative stress due to hazardous byproducts of respiration. Our approach consists of coupling the cell density field to a chemoattractant concentration as well as to nutrient and waste fields. The latter serves as a triggering field for emission of chemoattractant. Important elements in the proposed model include the propagation of a front of motile bacteria radially outward form an initial site, a Turing instability of the uniformly dense state and a reduction of motility for cells sufficiently far behind the front. The wide variety of patterns seen in the experiments is explained as being due the variation of the details of the initiation of the chemoattractant emission as well as the transition to a non-motile phase.

Tsimring, L S; Aranson, I S; Ben-Jacob, E; Cohen, I; Shochet, O; Tsimring, Lev; Levine, Herbert; Aranson, Igor; Ben-Jacob, Eshel; Cohen, Inon; Shochet, Ofer

1995-01-01

378

Swimming bacteria power microscopic gears  

Science.gov (United States)

While the laws of thermodynamics prohibit extraction of useful work from the Brownian motion of particles in systems at equilibrium, under non-equlibrium conditions their motions can be ``rectified'', for example, in the presence of asymmetric geometrical obstacles. We describe a class of systems in which aerobic bacteria Bacillus subtilis moving randomly in a fluid film power submillimeter gears and primitive systems of gears decorated with asymmetric teeth. The directional rotation is observed only in the regime of collective bacterial swimming and the gears' angular velocities depend on and can be controlled by the amount of oxygen available to the bacteria. The ability to harness and control the power of collective motions appears an important requirement for further development of mechanical systems driven by microorganism.

Sokolov, Andrey; Aronson, Igor; Apodaca, Mario; Grzybowski, Bartosz

2010-03-01

379

Swimming bacteria power microscopic gears  

Energy Technology Data Exchange (ETDEWEB)

Whereas the laws of thermodynamics prohibit extraction of useful work from the Brownian motion of particles in equilibrium, these motions can be “rectified” under nonequilibrium conditions, for example, in the presence of asymmetric geometrical obstacles. Here, we describe a class of systems in which aerobic bacteria Bacillus subtilis moving randomly in a fluid film power submillimeter gears and primitive systems of gears decorated with asymmetric teeth. The directional rotation is observed only in the regime of collective bacterial swimming and the gears’ angular velocities depend on and can be controlled by the amount of oxygen available to the bacteria. The ability to harness and control the power of collective motions appears an important requirement for further development of mechanical systems driven by microorganisms.

Sokolov, Andrey; Apodaca, Mario M.; Grzybowski, Bartosz A.; Aranson, Igor S.

2010-01-01

380

Swimming bacteria power microscopic gears.  

Energy Technology Data Exchange (ETDEWEB)

Whereas the laws of thermodynamics prohibit extraction of useful work from the Brownian motion of particles in equilibrium, these motions can be 'rectified' under nonequilibrium conditions, for example, in the presence of asymmetric geometrical obstacles. Here, we describe a class of systems in which aerobic bacteria Bacillus subtilis moving randomly in a fluid film power submillimeter gears and primitive systems of gears decorated with asymmetric teeth. The directional rotation is observed only in the regime of collective bacterial swimming and the gears angular velocities depend on and can be controlled by the amount of oxygen available to the bacteria. The ability to harness and control the power of collective motions appears an important requirement for further development of mechanical systems driven by microorganisms.

Sokolov, A.; Apodaca, M. M.; Grzybowski, B. A.; Aranson, I. S.; Materials Science Division; Princeton Univ.; Northwestern Univ.

2010-01-19

 
 
 
 
381

Synergetic effect in bacteria radiobiology  

International Nuclear Information System (INIS)

Synergetic effect in bacteria radiobiology is considered on the example of combined thermoradiation and magneto-radiation effect. When considering leading mechanisms of synergetic effects the main role is played by the violation of DNA repair processes. The formation of complex non-repair genome damages proceeds under conditions of inhibition of fermentative complexes providing stability of genetic cell information. In the case of combined radiation effect and other bactericide factors additional damages of membranes and energy supply systems of cells are very important in the formation of the final radiobiologic effect. An important role in the synergetic intensification of bacteria death is played by violations in the balance of DNA and protein synthesis reducing the effectiveness of processes of cell restoration.

1984-01-01

382

Virulence properties of cariogenic bacteria  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract The importance of Streptococcus mutans in the etiology of dental caries has been well documented. However, there is growing recognition that the cariogenic potential of dental plaque may be determined by the composite interactions of the total plaque bacteria rather than solely the virulence properties of a single organism. This study will examine how the interactions of S. mutans with other biofilm constituents may influence the cariogenicity of plaque samples. In order to begin to investigate the effects of nonmutans streptococci on the cariogenic potential of S. mutans, we have examined the effects of Streptococcus gordonii on the virulence properties of the former organisms. These studies have indicated that S.gordonii can attenuate several potential virulence properties of S. mutans including bacteriocin production, genetic transformation, and biofilm formation. Therefore, modulation of the interactions between plaque bacteria might be a novel approach for attenuating dental caries initiation.

Kuramitsu Howard K; Wang Bing-Yan

2006-01-01

383

Lima Bean Bacteria DNA Extraction  

Science.gov (United States)

This laboratory exercise is designed to show learners how DNA can easily be extracted from lima bean bacteria. This experiment requires the use of a centrifuge (not included in cost of materials). Use this experiment to supplement any unit on genetics and to demonstrate how scientists study DNA. Adult supervision is recommended. This resource guide includes tips and suggestions for instructors as well as other DNA extraction experiments and a chart for learners to answer questions.

Hays, Lana

2009-01-01

384

--- but bacteria fuel the debate  

Energy Technology Data Exchange (ETDEWEB)

Bacteria that convert carbon dioxide in the air into energy could be used as an alternative source of fuel with no conflict with food production. Chlorella, a green alga has a fast growth rate and it is estimated that each hectare of growth area could produce the equivalent of 50,000 barrels of crude oil a year. Arthrobactir AK19 can accumulate over 85% of its body weight as fat which can then be refined and burnt.

1981-01-15

385

Geobiology of Marine Magnetotactic Bacteria  

Canada Institute for Scientific and Technical Information (Canada)

Magnetotactic bacteria (MTB) biomineralize intracellular membrane-bound crystals of magnetite (Fe304) or greigite (Fe3S4), and are abundant in the suboxic to anoxic zones of stratified marine environments worldwide. Their population densities (up to 10(5) cells ml(-1)) and high intracellular iron content suggest a potentially significant role in iron cycling, but very little is known about their population dynamics and regulation by environmental geochemistry.

2006-01-01

386

Memory in bacteria and phage.  

Science.gov (United States)

Whenever the state of a biological system is not determined solely by present conditions but depends on its past history, we can say that the system has memory. Bacteria and bacteriophage use a variety of memory mechanisms, some of which seem to convey adaptive value. A genetic type of heritable memory is the programmed inversion of specific DNA sequences, which causes switching between alternative patterns of gene expression. Heritable memory can also be based on epigenetic circuits, in which a system with two possible steady states is locked in one or the other state by a positive feedback loop. Epigenetic states have been observed in a variety of cellular processes, and are maintained by diverse mechanisms. Some of these involve alternative DNA methylation patterns that are stably transmitted to daughter molecules and can affect DNA-protein interactions (e.g., gene transcription). Other mechanisms exploit autocatalytic loops whereby proteins establish the proper conditions for their continued synthesis. Template polymers other than nucleic acids (e.g., components of the cell wall) may also propagate epigenetic states. Non-heritable memory is exemplified by parasitic organisms that bear a signature of their previous host, such as host-controlled modification of phage DNA or porin hitchhiking in predatory bacteria. The heterogeneous nature of the examples known may be indicative of widespread occurrence of memory mechanisms in bacteria and phage. However, the actual extent, variety and potential selective value of prokaryotic memory devices remain open questions, still to be addressed experimentally. PMID:12111734

Casadesús, Josep; D'Ari, Richard

2002-06-01

387

Memory in bacteria and phage.  

UK PubMed Central (United Kingdom)

Whenever the state of a biological system is not determined solely by present conditions but depends on its past history, we can say that the system has memory. Bacteria and bacteriophage use a variety of memory mechanisms, some of which seem to convey adaptive value. A genetic type of heritable memory is the programmed inversion of specific DNA sequences, which causes switching between alternative patterns of gene expression. Heritable memory can also be based on epigenetic circuits, in which a system with two possible steady states is locked in one or the other state by a positive feedback loop. Epigenetic states have been observed in a variety of cellular processes, and are maintained by diverse mechanisms. Some of these involve alternative DNA methylation patterns that are stably transmitted to daughter molecules and can affect DNA-protein interactions (e.g., gene transcription). Other mechanisms exploit autocatalytic loops whereby proteins establish the proper conditions for their continued synthesis. Template polymers other than nucleic acids (e.g., components of the cell wall) may also propagate epigenetic states. Non-heritable memory is exemplified by parasitic organisms that bear a signature of their previous host, such as host-controlled modification of phage DNA or porin hitchhiking in predatory bacteria. The heterogeneous nature of the examples known may be indicative of widespread occurrence of memory mechanisms in bacteria and phage. However, the actual extent, variety and potential selective value of prokaryotic memory devices remain open questions, still to be addressed experimentally.

Casadesús J; D'Ari R

2002-06-01

388

Bacteria under simulated Martian conditions.  

UK PubMed Central (United Kingdom)

The behavior of organisms in simulated Martian conditions is of great importance to exobiology for two reasons: (1) Because of the extreme environment of Mars, the likelihood of contamination of the planet by earth organisms is considered slight by some scientists. To date, there has been little evidence to contradict this supposition. Such evidence is presented. (2) The selection and adaptation of earth bacteria to Martian conditions is potentially significant in understanding Martian life, if it exists, and may be helpful in designing life-detection techniques and devices. Of course, simulation attempts, based on current knowledge of the Mars environment, may be far from the actual conditions, and extrapolations made from such situations of no real significance. However, generalizations can be made and cautious interpretation of the results of those experiments seems well worth reporting. A new technique for simulation of known parameters of the Martian environment is discussed along with possible biological implications. The response of bacteria to such simulation is demonstrated in terms of survival and growth, showing that certain bacteria will not only survive, but grow during simulated Martian freeze-thaw cycling if water is present. Ways are demonstrated in which water can be present on Mars although not detectable with current technology. Plans for future experimentation are discussed.

Young RS; Deal PH; Bell J; Allen JL

1964-01-01

389

The sulphate-reducing bacteria  

Energy Technology Data Exchange (ETDEWEB)

This monograph surveys knowledge about an unusual and little-studied group of microbes, bringing together information that has hitherto been widely scattered throughout the scientific literature. The sulphate-reducing bacteria cannot grow in air; they respire s