scholarly journals Pyruvate Kinase of the Hyperthermophilic Crenarchaeote Thermoproteus tenax: Physiological Role and Phylogenetic Aspects

2000 ◽  
Vol 182 (7) ◽  
pp. 2001-2009 ◽  
Author(s):  
Alexander Schramm ◽  
Bettina Siebers ◽  
Britta Tjaden ◽  
Henner Brinkmann ◽  
Reinhard Hensel
Keyword(s):  
Pyruvate Kinase ◽  
Specific Activity ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Mrna Level ◽  
Energy Charge ◽  
Physiological Role ◽  
Early Gene ◽  
Bacterial Phyla ◽  
Thermoproteus Tenax

ABSTRACT Pyruvate kinase (PK; EC 2.7.1.40 ) of Thermoproteus tenax was purified to homogeneity, and its coding gene was cloned and expressed in Escherichia coli. It represents a homomeric tetramer with a molecular mass of 49 kDa per subunit. PK exhibits positive binding cooperativity with respect to phosphoenolpyruvate and metal ions such as Mg2+ and Mn2+. Heterotropic effects, as commonly found for PKs from bacterial and eucaryal sources, could not be detected. The enzyme does not depend on K+ ions. Heterotrophically grown cells exhibit specific activity of PK four times higher than autotrophically grown cells. Since the mRNA level of the PK coding gene is also accordingly higher in heterotrophic cells, we conclude that the PK activity is adjusted to growth conditions mainly on the transcript level. The enzymic properties of the PK and the regulation of its expression are discussed with respect to the physiological framework given by the T. tenax-specific variant of the Embden-Meyerhof-Parnas pathway. T. tenax PK shows moderate overall sequence similarity (25 to 40% identity) to its bacterial and eucaryal pendants. Phylogenetic analyses of the known PK sequences result in a dichotomic tree topology that divides the enzymes into two major PK clusters, probably diverged by an early gene duplication event. The phylogenetic divergence is paralleled by a striking phenotypic differentiation of PKs: PKs of cluster I, which occur in eucaryal cytoplasm, some gamma proteobacteria, and low-GC gram-positive bacteria, are only active in the presence of fructose-1,6-bisphosphate or other phosphorylated sugars, whereas PKs of cluster II, found in various bacterial phyla, plastids, and in Archaea, show activity without effectors but are commonly regulated by the energy charge of the cell.

scholarly journals Methanococci Use the Diaminopimelate Aminotransferase (DapL) Pathway for Lysine Biosynthesis

2010 ◽  
Vol 192 (13) ◽  
pp. 3304-3310 ◽  
Author(s):  
Yuchen Liu ◽  
Robert H. White ◽  
William B. Whitman
Keyword(s):  
Specific Activity ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Direct Conversion ◽  
Maximum Activity ◽  
Lysine Biosynthesis ◽  
Diaminopimelic Acid ◽  
Reading Frame ◽  
Cell Extracts ◽  
Methanothermobacter Thermautotrophicus

ABSTRACT The pathway of lysine biosynthesis in the methanococci has not been identified previously. A variant of the diaminopimelic acid (DAP) pathway uses diaminopimelate aminotransferase (DapL) to catalyze the direct conversion of tetrahydrodipicolinate (THDPA) to ll-DAP. Recently, the enzyme DapL (MTH52) was identified in Methanothermobacter thermautotrophicus and shown to belong to the DapL1 group. Although the Methanococcus maripaludis genome lacks a gene that can be unambiguously assigned a DapL function based on sequence similarity, the open reading frame MMP1527 product shares 30% amino acid sequence identity with MTH52. A Δmmp1527 deletion mutant was constructed and found to be a lysine auxotroph, suggesting that this DapL homolog in methanococci is required for lysine biosynthesis. In cell extracts of the M. maripaludis wild-type strain, the specific activity of DapL using ll-DAP and α-ketoglutarate as substrates was 24.3 ± 2.0 nmol min−1 mg of protein−1. The gene encoding the DapL homolog in Methanocaldococcus jannaschii (MJ1391) was cloned and expressed in Escherichia coli, and the protein was purified. The maximum activity of MJ1391 was observed at 70°C and pH 8.0 to 9.0. The apparent Km s of MJ1391 for ll-DAP and α-ketoglutarate were 82.8 ± 10 μM and 0.42 ± 0.02 mM, respectively. MJ1391 was not able to use succinyl-DAP or acetyl-DAP as a substrate. Phylogenetic analyses suggested that two lateral gene transfers occurred in the DapL genes, one from the archaea to the bacteria in the DapL2 group and one from the bacteria to the archaea in the DapL1 group. These results demonstrated that the DapL pathway is present in marine methanogens belonging to the Methanococcales.

Interaction of catechol O-methyltransferase with gold and silver nanoparticles

2018 ◽  
Author(s):  
◽  
Aminu Usman
Keyword(s):  
Silver Nanoparticles ◽  
Catalytic Activity ◽  
Electrostatic Interactions ◽  
Specific Activity ◽  
Sequence Similarity ◽  
Physiological Role ◽  
Spectroscopic Studies ◽  
Hydroxyl Group ◽  
Gold And Silver Nanoparticles ◽  
Silver Nps

Catechol O-methyltransferase (S-adenosyl-Z-methionine: catechol O-methyltransferase; COMT; EC 2.1.1.6) is a ubiquitous enzyme that catalyses the transfer of a methyl group from the cofactor, S-adenosyl-Z-methionine (SAM) to a hydroxyl group of endogenous and exogenous catechol-containing moieties. The physiological role of this enzyme is the methylation and thereby inactivation of the catechol-containing bio-active and bio-toxic compounds, including catechol-neurotransmitters, catechol-estrogens and catechol-containing drugs. Activity of this enzyme is implicated in the treatment of Parkinson’s disease and is associated with other diseases including breast cancer and an array neuropsychological disorders, such as schizophrenia. This thesis explores the use of gold and silver nanoparticles (NPs) (AuNPs and AgNPs) to inhibit the catalytic activity of mammalian COMT. Because of its accessibility and availability, we initially investigated bovine soluble COMT (BSCOMT) from liver tissue. Bioinformatic analyses and structural modeling revealed high (>90%) sequence similarity between BSCOMT and human soluble COMT (HSCOMT). BSCOMT was partially purified to 7.78 fold, 1.65% yield and had a specific activity of 0.052 U/mg. It had pH and temperature optima of 8.5 and 40oC, respectively. The Km, Vmax, Kcat and Kcat/Km towards esculetin methylation were respectively 1.475±0.130 pM, 0.0353±0.001 pmol/ml/min, 1.748 x 10-2±5.0x10-4 min-1 and 1.18x10-2 M-1. min-1. HSCOMT was expressed in Escherichia coli BL21(DE3) which showed optimal activity for esculetin methylation at pH and temperature of 7.0 and 30°C, respectively. It was purified to 5.62 fold, 22.6% yield with a specific activity of 3.85 U/mg. HSCOMT kinetic plots, upon incubation of the reaction mixture at 30°C for 5 min before addition of SAM was hyperbolic with Km, Vmax, Kcat and Kcat/Km values of 1.79 pM, 0.412 pmol/ml/min, 2.08 min-1 and 1.165 M-1. min-1, respectively. AuNPs and AgNPs showed a concentration dependent inhibition of HSCOMT activity upon increasing the 5 min incubation time to 1 h. Interestingly, HSCOMT kinetics, with 1 h incubation at 30°C, showed a sigmoidal curve, as well as increased activity. Incubation of the reaction mixture in the presence of 60 pM AuNPs and/or AgNPs for 1 hreversed the observed sigmoidal to a hyperbolic curve, with kinetic parameters comparable to those of 5 min incubation. SDS-PAGE analyses of HSCOMT after the kinetic experiments showed the enzyme incubated for 5 min as a monomer, while that which was incubated for 1 h migrated substantially as dimer. However, the HSCOMT incubated for 1 h in the presence of 60 pM AuNPs and/or AgNPs migrated as a monomer. This indicated that the extension of the incubation period allowed the dimerization of HSCOMT, which exhibited sigmoidal kinetics and higher activity. The presence of NPs impeded the HSCOMT dimerization which decreased the activity. Varying the concentration of SAM suggested that SAM had an allosteric modulatory effect on HSCOMT. Absorption spectroscopy indicated adsorption of HSCOMT on the gold and silver NP surfaces and the formation of NPs-HSCOMT corona. Fluorescence spectroscopy showed that the interaction of HSCOMT with both gold and silver NPs was governed by a static quenching mechanism, implying the formation of a non-fluorescent fluorophore-NP complex at the ground state. Further fluorometric analyses indicated that both gold and silver NPs had contact with Trp143; that the interactions were spontaneous and were driven by electrostatic interactions. Fourier transform infrared spectroscopic studies showed the adsorption of HSCOMT of the NPs surfaces to cause relaxation of the enzyme’s B-sheet structures. Molecular docking studies indicated involvement of largely hydrophilic amino acids, with the interacting distances of less than 3.5A. These findings signify the potential of nanotechnology in the control of COMT catalytic activity for the management of the COMT-related disorders.

scholarly journals Marisediminicola antarctica gen. nov., sp. nov., an actinobacterium isolated from the Antarctic

2010 ◽  
Vol 60 (11) ◽  
pp. 2535-2539 ◽  
Author(s):  
Hui-Rong Li ◽  
Yong Yu ◽  
Wei Luo ◽  
Yin-Xin Zeng
Keyword(s):  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Intertidal Sediment ◽  
Rrna Gene ◽  
Optimum Growth ◽  
Temperature Range ◽  
Zhongshan Station ◽  
Diamino Acid ◽  
Ph Range ◽  
The Antarctic

Strain ZS314T was isolated from a sandy intertidal sediment sample collected from the coastal area off the Chinese Antarctic Zhongshan Station, east Antarctica (6 ° 22′ 13″ S 7 ° 21′ 41″ E). The cells were Gram-positive, motile, short rods. The temperature range for growth was 0–26 °C and the pH for growth ranged from 5 to 10, with optimum growth occurring within the temperature range 18–23 °C and pH range 6.0–8.0. Growth occurred in the presence of 0–6 % (w/v) NaCl, with optimum growth occurring in the presence of 2–4 % (w/v) NaCl. Strain ZS314T had MK-10 as the major menaquinone and anteiso-C15 : 0, iso-C16 : 0 and anteiso-C17 : 0 as major fatty acids. The cell-wall peptidoglycan type was B2β with ornithine as the diagnostic diamino acid. The major polar lipids were diphosphatidylglycerol and phosphatidylglycerol. The genomic DNA G+C content was approximately 67 mol%. Phylogenetic analysis based on 16S rRNA gene sequence similarity showed that strain ZS314T represents a new lineage in the family Microbacteriaceae. On the basis of the phylogenetic analyses and phenotypic characteristics, a new genus, namely Marisediminicola gen. nov., is proposed, harbouring the novel species Marisediminicola antarctica sp. nov. with the type strain ZS314T (=DSM 22350T =CCTCC AB 209077T).

scholarly journals Nesterenkonia alkaliphila sp. nov., an alkaliphilic, halotolerant actinobacteria isolated from the western Pacific Ocean

2015 ◽  
Vol 65 (Pt_2) ◽  
pp. 516-521 ◽  
Author(s):  
Gaiyun Zhang ◽  
Yubian Zhang ◽  
Xijie Yin ◽  
Shuang Wang
Keyword(s):  
Pacific Ocean ◽  
Type Species ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Western Pacific ◽  
Rrna Gene ◽  
Western Pacific Ocean ◽  
Content Type ◽  
Link Type ◽  
The Western Pacific

A Gram-staining-positive, aerobic, motile and non-spore-forming actinobacteria, designated strain F10T, was isolated from a deep-sea sediment of the western Pacific Ocean. Phylogenetic and phenotypic properties of the organism supported that it belonged to the genus Nesterenkonia . Strain F10T shared highest 16S rRNA gene sequence similarity of 96.8 % with Nesterenkonia aethiopica DSM 17733T, followed by Nesterenkonia xinjiangensis YIM 70097T (96.7 %) and Nesterenkonia alba CAAS 252T (96.6 %). The organism grew at 4–50 °C, at pH 7.0–12.0 and in the presence of 0–12 % (w/v) NaCl, with optimal growth occurring at 40 °C, at pH 9.0 and in the presence of 1 % (w/v) NaCl. The peptidoglycan type was A4(alpha), l-Lys–Gly–l-Glu. The polar lipid profile of strain F10T consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, two unknown glycolipids and two unknown lipids. The isolate contained MK-9 (92 %) and MK-8 (5.8 %) as the major components of the menaquinone system, and anteiso-C17 : 0 (50.9 %) and anteiso-C15 : 0 (29.8 %) as the predominant fatty acids. The G+C content of the genomic DNA of strain F10T was 66.2 mol%. Based on phenotypic, genotypic and phylogenetic analyses, strain F10T represents a novel species of the genus Nesterenkonia for which the name Nesterenkonia alkaliphila sp. nov. is proposed. The type strain is F10T ( = LMG 28112T = CGMCC 1.12781T = JCM 19766T = MCCC 1A09946T).

Temperature and the Regulation of Enzyme Activity in the Hibernator. Isoenzymes of Liver Pyruvate Kinase from the Hibernating and Non-hibernating Arctic Ground Squirrel

1974 ◽  
Vol 52 (10) ◽  
pp. 894-902 ◽  
Author(s):  
Hans W. Behrisch
Keyword(s):  
Molecular Weight ◽  
Pyruvate Kinase ◽  
Wide Temperature Range ◽  
Ground Squirrel ◽  
Energy Charge ◽  
High Energy ◽  
Ground Squirrels ◽  
The Arctic ◽  
Arctic Ground Squirrel ◽  
Temperature Range

Liver of the hibernating (H) Arctic ground squirrel (Citellus undulatus) contains a single species of pyruvate kinase (PyK) that is distinct from the single isoenzyme of pyK observed in the non-hibernating (NH) ground squirrel, which has been previously described (Behrisch &Johnson (1974) Can. J. Biochem. 52, 547–559). The H-PyK has a pI value of 5.7 and a molecular weight of 241 000 – 243 000. Affinity of the H-PyK for the substrates phosphoenolpyruvate (PEP) and ADP is not affected by changing temperature. It is argued that this stability of the apparent Km's for substrate over a wide temperature range permits the hibernator to take advantage of the Q10 effect in maintaining a low rate of the PyK reaction. Similarly, affinity of H-PyK for the allosteric activator fructose-1,6-phosphate (FDP) and the inhibitor ATP is also conspicuously independent of temperature, suggesting a fine stoichiometry in the relative concentrations of the regulatory ligands in control of H-PyK over a wide temperature range. Further, affinity of H-PyK for the inhibitor ATP is about three- to fourfold lower than that of the NH-PyK, a condition that would favor the maintenance of a high energy charge in the hibernating liver cell. ATP apparently inhibits PyK by causing a dissociation of the enzyme molecule into two "halves" of about 110 000 molecular weight each. This dissociation is offset and reversed by FDP. Removal of the ATP by dialysis does not of itself result in a reassociation of the PyK "halves"; FDP and/or the substrates are required for the two subunits of PyK to reassociate. As the apparent Ki of H-PyK for ATP is higher than that of NH-PyK, substantially higher concentrations of ATP are required to effect the dissociation of H-PyK. Similarly, elevated concentrations of FDP are required to offset the ATP-caused dissociation of the H-PyK.Hibernating Arctic ground squirrels that are preparing to emerge finally from the hibernating state already possess substantial activities of the NH-PyK isoenzyme. This suggests that the animal "anticipates" its transition from one metabolic state from another. On the basis of these data a formal mechanism is proposed for the regulation of liver PyK in the Arctic ground squirrel in both the non-hibernating and hibernating states.

scholarly journals Rubritalea halochordaticola sp. nov., a carotenoid-producing verrucomicrobial species isolated from a marine chordate

2011 ◽  
Vol 61 (7) ◽  
pp. 1515-1520 ◽  
Author(s):  
Jaewoo Yoon ◽  
Satoru Matsuda ◽  
Kyoko Adachi ◽  
Hiroaki Kasai ◽  
Akira Yokota
Keyword(s):  
Dna Hybridization ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Diaminopimelic Acid ◽  
Rrna Gene ◽  
The Novel ◽  
16S Rrna Gene Sequences ◽  
Sea Squirt ◽  
Cell Biomass ◽  
Taxonomic Approach

A Gram-negative-staining, obligately aerobic, non-motile, rod-shaped and chemoheterotrophic bacterium, designated strain MN1-1006T, was isolated from an ascidian (sea squirt) sample, and was studied using a polyphasic taxonomic approach. Phylogenetic analyses based on 16S rRNA gene sequences indicated that the new isolate shared approximately 93–99% sequence similarity with recognized species of the genus Rubritalea within the phylum ‘Verrucomicrobia’. DNA–DNA hybridization values between strain MN1-1006T and Rubritalea squalenifaciens HOact23T and Rubritalea sabuli YM29-052T were 57% and 14.5%, respectively. Strain MN1-1006T produced carotenoid compounds that rendered the cell biomass a reddish pink colour. The strain also contained squalene. The cell-wall peptidoglycan of the novel strain contained muramic acid and meso-diaminopimelic acid. The DNA G+C content of strain MN1-1006T was 51.4 mol%. The major cellular fatty acids were iso-C14:0, iso-C16:0 and anteiso-C15:0. The major isoprenoid quinone was MK-9. On the basis of these data, it was concluded that strain MN1-1006T represents a novel species of the genus Rubritalea, for which the name Rubritalea halochordaticola sp. nov. is proposed. The type strain is MN1-1006T ( = KCTC 23186T = NBRC 107102T).

scholarly journals Complete WO Phage Sequences Reveal Their Dynamic Evolutionary Trajectories and Putative Functional Elements Required for Integration into the Wolbachia Genome

2009 ◽  
Vol 75 (17) ◽  
pp. 5676-5686 ◽  
Author(s):  
Kohjiro Tanaka ◽  
Seiichi Furukawa ◽  
Naruo Nikoh ◽  
Tetsuhiko Sasaki ◽  
Takema Fukatsu
Keyword(s):  
Genetic Transformation ◽  
Protein Interactions ◽  
Phylogenetic Analyses ◽  
Attachment Site ◽  
Host Bacterium ◽  
Sequence Motifs ◽  
Integrase Gene ◽  
Bacterial Phyla ◽  
Genetic Elements ◽  
Evolutionary Trajectories

ABSTRACT Wolbachia endosymbionts are ubiquitously found in diverse insects including many medical and hygienic pests, causing a variety of reproductive phenotypes, such as cytoplasmic incompatibility, and thereby efficiently spreading in host insect populations. Recently, Wolbachia-mediated approaches to pest control and management have been proposed, but the application of these approaches has been hindered by the lack of genetic transformation techniques for symbiotic bacteria. Here, we report the genome and structure of active bacteriophages from a Wolbachia endosymbiont. From the Wolbachia strain wCauB infecting the moth Ephestia kuehniella two closely related WO prophages, WOcauB2 of 43,016 bp with 47 open reading frames (ORFs) and WOcauB3 of 45,078 bp with 46 ORFs, were characterized. In each of the prophage genomes, an integrase gene and an attachment site core sequence were identified, which are putatively involved in integration and excision of the mobile genetic elements. The 3′ region of the prophages encoded genes with sequence motifs related to bacterial virulence and protein-protein interactions, which might represent effector molecules that affect cellular processes and functions of their host bacterium and/or insect. Database searches and phylogenetic analyses revealed that the prophage genes have experienced dynamic evolutionary trajectories. Genes similar to the prophage genes were found across divergent bacterial phyla, highlighting the active and mobile nature of the genetic elements. We suggest that the active WO prophage genomes and their constituent sequence elements would provide a clue to development of a genetic transformation vector for Wolbachia endosymbionts.

scholarly journals Mesonia hippocampi sp. nov., isolated from the brood pouch of a diseased Barbour's Seahorse (Hippocampus barbouri)

2015 ◽  
Vol 65 (Pt_7) ◽  
pp. 2241-2247 ◽  
Author(s):  
Judy Kolberg ◽  
Hans-Jürgen Busse ◽  
Thomas Wilke ◽  
Patrick Schubert ◽  
Peter Kämpfer ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Type Strain ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Brood Pouch ◽  
Diaminopimelic Acid ◽  
Sugar Residue ◽  
Rrna Gene ◽  
Polar Lipid Profile ◽  
Monophyletic Cluster

An orange-pigmented, Gram-staining-negative, rod-shaped bacterium, designated 96_Hippo_TS_3/13T was isolated from the brood pouch of a diseased seahorse male of the species Hippocampus barbouri from the animal facility of the University of Giessen, Germany. Phylogenetic analyses based on the nearly full-length 16S rRNA gene sequence placed strain 96_Hippo_TS_3/13T into the monophyletic cluster of the genus Mesonia within the family Flavobacteriaceae. However, the strain shared only 92.2–93.8 % sequence similarity to type strains of species of the genus Mesonia, with highest sequence similarity to the type strain of Mesonia aquimarina. Cellular fatty acid analysis showed a Mesonia-typical fatty acid profile including several branched and hydroxyl fatty acids with highest amounts of iso-C15 : 0 (40.9 %) followed by iso-C17 : 0 3-OH (14.8 %). In the polyamine pattern, sym-homospermidine was predominant. The diagnostic diamino acid of the peptidoglycan was meso-diaminopimelic acid. The quinone system contained exclusively menaquinone MK-6. The only identified compound in the polar lipid profile was phosphatidylethanolamine present in major amounts. Additionally, major amounts of an unidentified aminolipid and two unidentified lipids not containing a phosphate group, an amino group or a sugar residue were detected. The genomic G+C content of strain 96_Hippo_TS_3/13T was 30 mol%. Based on genotypic, chemotaxonomic and physiological characterizations we propose a novel species of the genus Mesonia, Mesonia hippocampi sp. nov., with strain 96_Hippo_TS_3/13T ( = CIP 110839T =  LMG 28572T = CCM 8557T) as the type strain. An emended description of the genus Mesonia is also provided.

scholarly journals Erythrobacter luteolus sp. nov., isolated from a tidal flat of the Yellow Sea in Korea

2005 ◽  
Vol 55 (3) ◽  
pp. 1167-1170 ◽  
Author(s):  
Jung-Hoon Yoon ◽  
Kook Hee Kang ◽  
Soo-Hwan Yeo ◽  
Tae-Kwang Oh
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Tidal Flat ◽  
Yellow Sea ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Major Fatty Acid ◽  
The Yellow Sea ◽  
Rrna Gene ◽  
Phenotypic Properties

A Gram-negative, non-spore-forming, yellow-pigmented, slightly halophilic bacterial strain, SW-109T, was isolated from a tidal flat of the Yellow Sea in Korea, and subjected to a polyphasic taxonomic study. This isolate did not produce bacteriochlorophyll a and contained ubiquinone-10 as the predominant respiratory lipoquinone and C18 : 1 ω7c as the major fatty acid. The DNA G+C content was 60·3 mol%. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain SW-109T is phylogenetically affiliated to the genus Erythrobacter of the family Sphingomonadaceae. Strain SW-109T exhibited levels of 16S rRNA gene sequence similarity to the type strains of Erythrobacter species of 94·0–96·3 %, making it possible to categorize strain SW-109T as a species that is separate from previously recognized Erythrobacter species. On the basis of its phenotypic properties and phylogenetic distinctiveness, SW-109T (=KCTC 12311T=JCM 12599T) was classified as the type strain of a novel Erythrobacter species, for which the name Erythrobacter luteolus sp. nov. is proposed.

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