共查询到20条相似文献,搜索用时 31 毫秒
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Seung Won Kang Bo Young Jeon Tae Sik Hwang Doo Hyun Park 《Journal of microbiology (Seoul, Korea)》2009,47(6):721-727
A bacterium growing inside yeast cytoplasm was observed by light microscope without staining. The bacterium was separately
stained from yeast cell by a fluorescent dye, 4′,6-diamidino-2-phenylindole (DAPI). The bacterium actively moved inside yeast
cytoplasm and propagated in company with the yeast growth. The bacterium was separated from the yeast cytoplasm by selective
disruption of yeast cells and the yeast without the intracellular bacterium (YWOB) was obtained by selective inactivation
of bacterial cells. The yeast and the intracellular bacterium were identified as Candida tropicalis and Microbacterium sp., respectively. The length of Microbacterium sp. and C. tropicalis measured with SEM image was smaller than 0.5 μm and was larger than 5 μm, respectively. The yeast with the intracellular
bacterium (YWIB) grew in a starch-based medium but the YWOB was not C. tropicalis has neither extracellular nor intracellular saccharification enzyme. Glucose was produced from starch by the extracellular
crude enzyme (culture fluid) of Microbacterium sp. YWIB produced significantly more ethanol from glucose than YWOB but did not from starch. Conclusively, C. tropicalis is thought to catabolize starch dependent upon Microbacterium sp. growing in its cytoplasm and furnish stable habitat for the Microbacterium sp. 相似文献
3.
Jitendra Kumar Manoj Kumar Verma Tarun Kumar Shashank Gupta Rajesh Pandey Monika Yadav Nar Singh Chauhan 《Indian journal of microbiology》2018,58(3):294-300
The human gut microbiome has a significant role in host physiology; however its role in gluten catabolism is debatable. Present study explores the role of human gut microbes in gluten catabolism and a native human gut microbe Cellulomonas sp. HM71 was identified. SSU rDNA analysis has described human gut microbiome structure and also confirmed the permanent residentship of Cellulomonas sp. HM71. Catabolic potential of Cellulomonas sp. HM71 to cleave antigenic gluten peptides indicates presence of candidate gene encoding biocatalytic machinery. Genome analysis has identified the presence of gene encoding S9A serine protease family—prolyl endopeptidase, with Ser591, Asp664 and His685 signature residues. Cellulomonas sp. HM71 prolyl endopeptidase activity was found optimal at pH 7.0 and 37 °C with a KM of 35.53 μmol and specifically cleaves at proline residue. Current study describes the gluten catabolism potential of Cellulomonas sp. HM71 depicting possible role of human gut microbes in gluten catabolism to confer resistance mechanisms for the onset of celiac diseases in populations with gluten diet. 相似文献
4.
Fatty acid amide hydrolase (FAAH) is a mammalian integral membrane enzyme responsible for the hydrolysis of a number of neuromodulatory fatty acid amides, including the endogenous cannabinoid anandamide and the sleep-inducing lipid oleamide. FAAH belongs to a large class of hydrolytic enzymes termed the "amidase signature family," whose members are defined by a conserved stretch of approximately 130 amino acids termed the "amidase signature sequence." Recently, site-directed mutagenesis studies of FAAH have targeted a limited number of conserved residues in the amidase signature sequence of the enzyme, identifying Ser-241 as the catalytic nucleophile and Lys-142 as an acid/base catalyst. The roles of several other conserved residues with potentially important and/or overlapping catalytic functions have not yet been examined. In this study, we have mutated all potentially catalytic residues in FAAH that are conserved among members of the amidase signature family, and have assessed their individual roles in catalysis through chemical labeling and kinetic methods. Several of these residues appear to serve primarily structural roles, as their mutation produced FAAH variants with considerable catalytic activity but reduced expression in prokaryotic and/or eukaryotic systems. In contrast, five mutations, K142A, S217A, S218A, S241A, and R243A, decreased the amidase activity of FAAH greater than 100-fold without detectably impacting the structural integrity of the enzyme. The pH rate profiles, amide/ester selectivities, and fluorophosphonate reactivities of these mutants revealed distinct catalytic roles for each residue. Of particular interest, one mutant, R243A, displayed uncompromised esterase activity but severely reduced amidase activity, indicating that the amidase and esterase efficiencies of FAAH can be functionally uncoupled. Collectively, these studies provide evidence that amidase signature enzymes represent a large class of serine-lysine catalytic dyad hydrolases whose evolutionary distribution rivals that of the catalytic triad superfamily. 相似文献
5.
The adipamidase of a mutant strainBrevibacterium sp. R312 involved in the degradation of adiponitrile to adipic acid was purified. Its N-terminal amino acid sequence was shown to be identical toBrevibacterium sp. R312 enantio-selective amidase andRhodococcus sp. N-774 amidase. 相似文献
6.
Kengo Yasuhira Naoki Shibata Go Mongami Yuki Uedo Yu Atsumi Yasuyuki Kawashima Atsushi Hibino Yusuke Tanaka Young-Ho Lee Dai-ichiro Kato Masahiro Takeo Yoshiki Higuchi Seiji Negoro 《The Journal of biological chemistry》2010,285(2):1239-1248
We performed x-ray crystallographic analyses of the 6-aminohexanoate cyclic dimer (Acd) hydrolase (NylA) from Arthrobacter sp., an enzyme responsible for the degradation of the nylon-6 industry byproduct. The fold adopted by the 472-amino acid polypeptide generated a compact mixed α/β fold, typically found in the amidase signature superfamily; this fold was especially similar to the fold of glutamyl-tRNAGln amidotransferase subunit A (z score, 49.4) and malonamidase E2 (z score, 44.8). Irrespective of the high degree of structural similarity to the typical amidase signature superfamily enzymes, the specific activity of NylA for glutamine, malonamide, and indoleacetamide was found to be lower than 0.5% of that for Acd. However, NylA possessed carboxylesterase activity nearly equivalent to the Acd hydrolytic activity. Structural analysis of the inactive complex between the activity-deficient S174A mutant of NylA and Acd, performed at 1.8 Å resolution, suggested the following enzyme/substrate interactions: a Ser174-cis-Ser150-Lys72 triad constitutes the catalytic center; the backbone N in Ala171 and Ala172 are involved in oxyanion stabilization; Cys316-Sγ forms a hydrogen bond with nitrogen (Acd-N7) at the uncleaved amide bond in two equivalent amide bonds of Acd. A single S174A, S150A, or K72A substitution in NylA by site-directed mutagenesis decreased the Acd hydrolytic and esterolytic activities to undetectable levels, indicating that Ser174-cis-Ser150-Lys72 is essential for catalysis. In contrast, substitutions at position 316 specifically affected Acd hydrolytic activity, suggesting that Cys316 is responsible for Acd binding. On the basis of the structure and functional analysis, we discussed the catalytic mechanisms and evolution of NylA in comparison with other Ser-reactive hydrolases. 相似文献
7.
I. Ostash B. Kolvenbach P. F.-X. Corvini V. Fedorenko B. Ostash Danuta Cichocka 《Journal of applied genetics》2018,59(1):119-121
The wide application of sulfonamide (SA) antibiotics in human and veterinary medicine contributes to the accumulation of these antibiotics in the environment and the corresponding onset of antibiotic resistance among bacteria. Microbacterium sp. BR1 is capable of mineralizing sulfamethoxazole and other SAs via a novel mechanism. The genetic basis of SA elimination by BR1 remains unknown. Development of an efficient plasmid transfer protocol for Microbacterium sp. BR1 is highly desirable, as it would open the door to genetic analysis and manipulation of its genome. Here we report that intergeneric Escherichia coli–Microbacterium spp. BR1 conjugation is an efficient way to introduce various plasmids into BR1. The generated transconjugants were stable in the presence of antibiotics and the plasmids showed no signs of rearrangements. Nevertheless, the plasmids were rapidly lost in the absence of selection. We also show that the cumate-inducible beta-glucuronidase reporter gene functions in BR1 and is strictly regulated. Our results set the working ground for further genetic manipulations of BR1, such as the overexpression of sulfonamide degradation genes or the selection of strong microbacterial promoters. 相似文献
8.
2-Chloronicotinic acid (2-CA) is an important building block for a series of agrochemicals and pharmaceuticals. Amidase-catalyzed hydrolysis of 2-chloronicotinamide is one of the most attractive approaches for 2-CA production. However, development of the bioprocess was plagued by low activity of amidase for 2-chloronicotinamide. In this work, an amidase signature (AS) family amidase from Pantoea sp. (Pa-Ami), with superior activity for nicotinamide and its chlorinated derivatives, was exploited and characterized. Kinetic analysis and molecular docking clearly indicated that chlorine substitution in the pyridine ring of nicotinamide, especially the substitution at 2-position led to a dramatic decrease of Pa-Ami activity. The productivity of the bioprocess was significantly improved using fed-batch mode at low reaction temperature and 2-CA was produced as high as 370 mM with a substrate conversion of 94.2%. These results imply that Pa-Ami is potentially promising biocatalyst for industrial production of 2-CA. 相似文献
9.
The type VI secretion systems (T6SS) have emerging roles in interspecies competition. In order to have an advantage in defense against other organisms, this system in Pseudomonas aeruginosa delivers a peptidoglycan amidase (Tse1) to the periplasmic space of a competitor. An immune protein (Tsi1) is also produced by the bacterium to protect itself from damage caused by Tse1. Tsi1 directly interacts with Tse1. We report that the crystal structure of Tse1 displays a common CHAP protein fold. Strikingly, our structures showed that the third residue in the catalytic triad may be novel as this residue type has not been observed previously. 相似文献
10.
Amidase 1 (AMI1), a specific indole-3-acetamide amidohydrolase, is an Arabidopsis thaliana amidase signature enzyme that catalyzes the synthesis of indole-3-acetic acid from indole-3-acetamide. Amidase signature family members catalyze a diverse range of enzymatic reactions and are found widespread in nature, for instance in bacteria, mammals, and plants. At the protein level, the family members share a conserved stretch of approximately 50-130 amino acids, the name-giving amidase signature. Elucidation of the crystal structures of a mammalian fatty acid amide hydrolase and the bacterial malonamidase E2 revealed an unusual Ser-cisSer-Lys catalytic triad in proteins of this family. In addition, other members, such as the amidase from Rhodococcus rhodochrous strain J1 or Sulfolobus solfataricus, seem to use an accessory Cys-cisSer-Lys center. AMI1 possesses all conserved amino-acid residues of the Ser-cisSer-Lys triad, but lacks the CX(3)C motif and therefore the Cys-cisSer-Lys catalytic site. Using a set of point-mutated variants of AMI1 and chemical modifications, we analyzed the relative importance of single amino-acid residues of AMI1 with respect to substrate conversion. These experiments revealed that a specific serine residue, Ser137, is essential for AMI1 enzymatic activity. We also report structural and functional differences of AMI1 from other amidase signature enzymes. 相似文献
11.
Fatty acid amide hydrolase (FAAH) is a mammalian amidase signature enzyme that inactivates neuromodulatory fatty acid amides, including the endogenous cannabinoid anandamide and the sleep-inducing substance oleamide. The recent determination of the three-dimensional structures of FAAH and two distantly related bacterial amidase signature enzymes indicates that these enzymes employ an unusual serine-serine-lysine triad for catalysis (Ser-241/Ser-217/Lys-142 in FAAH). Mutagenesis of each of the triad residues in FAAH has been shown to severely reduce amidase activity; however, how these residues contribute, both individually and in cooperation, to catalysis remains unclear. Here, through a combination of site-directed mutagenesis, enzyme kinetics, and chemical labeling experiments, we provide evidence that each FAAH triad residue plays a distinct role in catalysis. In particular, the mutation of Lys-142 to alanine indicates that this residue functions as both a base involved in the activation of the Ser-241 nucleophile and an acid that participates in the protonation of the substrate leaving group. This latter property appears to support the unusual ability of FAAH to hydrolyze amides and esters at equivalent rates. Interestingly, although structural evidence indicates that the impact of Lys-142 on catalysis probably occurs through the bridging Ser-217, the mutation of this latter residue to alanine impaired catalytic activity but left the amide/ester hydrolysis ratios of FAAH intact. Collectively, these findings suggest that FAAH possesses a specialized active site structure dedicated to a mechanism for competitive amide and ester hydrolysis where nucleophile attack and leaving group protonation occur in a coordinated manner dependent on Lys-142. 相似文献
12.
Cometabolic Degradation of Dibenzofuran by Biphenyl-Cultivated Ralstonia sp. Strain SBUG 290 总被引:2,自引:0,他引:2
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Drte Becher Michael Specht Elke Hammer Wittko Francke Frieder Schauer 《Applied microbiology》2000,66(10):4528-4531
Cells of the gram-negative bacterium Ralstonia sp. strain SBUG 290 grown in the presence of biphenyl are able to cooxidize dibenzofuran which has been 1,2-hydroxylated. Meta cleavage of the 1,2-dihydroxydibenzofuran between carbon atoms 1 and 9b produced 2-hydroxy-4-(3′-oxo-3′H-benzofuran-2′-yliden)but-2-enoic acid, which was degraded completely via salicylic acid. The presence of these intermediates indicates a degradation mechanism for dibenzofuran via lateral dioxygenation by Ralstonia sp. strain SBUG 290. 相似文献
13.
In the present work, we report a novel class of glutathione transferases (GSTs) originated from the pathogenic soil bacterium Agrobacterium tumefaciens C58, with structural and catalytic properties not observed previously in prokaryotic and eukaryotic GST isoenzymes. A GST-like sequence from A. tumefaciens C58 (Atu3701) with low similarity to other characterized GST family of enzymes was identified. Phylogenetic analysis showed that it belongs to a distinct GST class not previously described and restricted only in soil bacteria, called the Eta class (H). This enzyme (designated as AtuGSTH1-1) was cloned and expressed in E. coli and its structural and catalytic properties were investigated. Functional analysis showed that AtuGSTH1-1 exhibits significant transferase activity against the common substrates aryl halides, as well as very high peroxidase activity towards organic hydroperoxides. The crystal structure of AtuGSTH1-1 was determined at 1.4 Å resolution in complex with S-(p-nitrobenzyl)-glutathione (Nb-GSH). Although AtuGSTH1-1 adopts the canonical GST fold, sequence and structural characteristics distinct from previously characterized GSTs were identified. The absence of the classic catalytic essential residues (Tyr, Ser, Cys) distinguishes AtuGSTH1-1 from all other cytosolic GSTs of known structure and function. Site-directed mutagenesis showed that instead of the classic catalytic residues, an Arg residue (Arg34), an electron-sharing network, and a bridge of a network of water molecules may form the basis of the catalytic mechanism. Comparative sequence analysis, structural information, and site-directed mutagenesis in combination with kinetic analysis showed that Phe22, Ser25, and Arg187 are additional important residues for the enzyme''s catalytic efficiency and specificity. 相似文献
14.
The present paper describes the isolation, physiological and genetic characteristic of a bacterial agent which inhibits the growth of algae and causes death of laboratory cultures of Antarctic microalgal strains: prokaryotic cyanobacteria Synechocystis salina and green eukaryotic microalga Choricistis minor. The bacterial strain LB1 was isolated from algal damaged laboratory cultures of S. salina. It was established that this bacterium is obligate aerobic, Gram-positive, non-spore-forming, immotile, irregular rods with dimensions 0.3–2 μm. Our results showed that LB1 has algicidal effect to S. salina as well as to C. minor. Transmission electron microscopy observations confirmed the destruction of S. salina by the bacterium. Biochemical analysis of LB1 revealed positive reaction to d-glucose, catalase, hydrolysis of gelatin, acid production from: lactose, l-arabinose, l-ramnose, esculin and β-galactosidase. The partial sequence (1,404 bp) of the 16S rRNA gene of LB1 showed 99 % similarity with type strains of the genus Microbacterium. The results of the biochemical, antimicrobial and of 16S rRNA analysis of LB1 allowed us to identify LB1 as Microbacterium sp. Studying expression of pathogenicity of the bacteria to algal cultures will help to solve the problem of algal production for biotechnological purposes. 相似文献
15.
Z-phenylacetaldoxime degradation by a novel aldoxime dehydratase from Bacillus sp. strain OxB-1 总被引:2,自引:0,他引:2
Z-phenylacetaldoxime (Z-PAOx) degrading bacterium, identified as Bacillus sp. strain OxB-1, was isolated from soil after 2 months acclimation. The enzyme involved in the degradation of Z-PAOx was induced by the aldoxime and required FMN for its activity. The enzyme was partially purified from the cell-free extract of the strain and shown to catalyze the stoichiometric dehydration reaction of Z-PAOx to form phenylacetonitrile (PAN). Activities of nitrilase and amidase acting on PAN and phenylacetamide (PAAm), respectively, to form phenylacetate (PAA) were found in the strain grown on Z-PAOx. This is the first report of aldoxime dehydratase co-existing with nitrile degrading enzymes in bacteria. 相似文献
16.
Yun-lin Wei Tatsuo Kurihara Takeshi Suzuki Nobuyoshi Esaki 《Journal of Molecular Catalysis .B, Enzymatic》2003,23(2-6):357-365
A novel esterase that belongs to the amidase signature family was found in a psychrotrophic bacterium, Acinetobacter sp. strain no. 6, isolated from Siberian soil. The gene coding for the esterase, named EstA8, was cloned, and an open reading frame of 1488 bp corresponding to 496 amino acid residues was identified. EstA8 showed 30% sequence identity with 6-aminohexanoate-cyclic-dimer hydrolases from Pseudomonas sp. strain NK87 and Flavobacterium sp. strain K172, which degrade a by-product of the nylon-6 industry. EstA8 was overproduced in Escherichia coli JM109 under the control of the lac promoter of pUC118 and purified. Consistent with the fact that the source microorganism is cold-adapted, the enzyme was unstable at moderate temperatures. It lost 75% of its original activity by incubation at 40 °C for 30 min. Despite its structural similarity to 6-aminohexanoate-cyclic-dimer hydrolase, 6-aminohexanoate cyclic dimer did not serve as the substrate. EstA8 is a member of the amidase signature family, but its esterase activity toward p-nitrophenyl esters, such as p-nitrophenyl acetate, was much higher than its amidase activity toward p-nitroanilides, such as p-nitroacetanilide. 相似文献
17.
Wibberg D Blom J Jaenicke S Kollin F Rupp O Scharf B Schneiker-Bekel S Sczcepanowski R Goesmann A Setubal JC Schmitt R Pühler A Schlüter A 《Journal of biotechnology》2011,155(1):50-62
Agrobacterium sp. H13-3, formerly known as Rhizobium lupini H13-3, is a soil bacterium that was isolated from the rhizosphere of Lupinus luteus. The isolate has been established as a model system for studying novel features of flagellum structure, motility and chemotaxis within the family Rhizobiaceae. The complete genome sequence of Agrobacterium sp. H13-3 has been established and the genome structure and phylogenetic assignment of the organism was analysed. For de novo sequencing of the Agrobacterium sp. H13-3 genome, a combined strategy comprising 454-pyrosequencing on the Genome Sequencer FLX platform and PCR-based amplicon sequencing for gap closure was applied. The finished genome consists of three replicons and comprises 5,573,770 bases. Based on phylogenetic analyses, the isolate could be assigned to the genus Agrobacterium biovar I and represents a genomic species G1 strain within this biovariety. The highly conserved circular chromosome (2.82 Mb) of Agrobacterium sp. H13-3 mainly encodes housekeeping functions characteristic for an aerobic, heterotrophic bacterium. Agrobacterium sp. H13-3 is a motile bacterium driven by the rotation of several complex flagella. Its behaviour towards external stimuli is regulated by a large chemotaxis regulon and a total of 17 chemoreceptors. Comparable to the genome of Agrobacterium tumefaciens C58, Agrobacterium sp. H13-3 possesses a linear chromosome (2.15 Mb) that is related to its reference replicon and features chromosomal and plasmid-like properties. The accessory plasmid pAspH13-3a (0.6 Mb) is only distantly related to the plasmid pAtC58 of A. tumefaciens C58 and shows a mosaic structure. A tumor-inducing Ti-plasmid is missing in the sequenced strain H13-3 indicating that it is a non-virulent isolate. 相似文献
18.
A phenazine-1-carboxylic acid (PCA)–degrading bacterium, strain DP58, was isolated from pimiento rhizosoil. Based on morphology,
physiologic tests, 16S rDNA sequence, and phylogenetic characteristics, it was identified as Sphingomonas sp. The PCA-degradation experiments were conducted both in Luria-Bertani and inorganic salt medium at 28°C. The relationship
between bacterium growth and PCA degradation suggested that strain DP58 could use PCA as the sole source of carbon and nitrogen
and was able to completely degrade PCA in 40 hours. Newly isolated strain DP58 represents the first bacterium that can degrade
PCA. 相似文献
19.
Moslem Papizadeh Mohammad Roayaei Ardakani Gholamhossein Ebrahimipour Hossein Motamedi 《World journal of microbiology & biotechnology》2010,26(7):1195-1200
Oil-polluted soils were sampled from National Iranian South Oil Company (NISOC) for isolation and screening of C–S and not
C–C targeted Dibenzothiophene (DBT) degrading microorganisms. Microbacterium sp. NISOC-06, a C–S targeted DBT degrading bacterium, was selected and its desulfurization ability was studied in aqueous phase and water-gasoline
biphasic systems. The 16srRNA gene was amplified using universal eubacteria-specific primers, PCR product was sequenced and
the sequence of nearly 1,500 bp 16srDNA was studied. Based on Gas Chromatography results Microbacterium sp. NISOC-06 utilized 94.8% of 1 mM DBT during the 2 weeks of incubation. UV Spectrophotometry and biomass production measurements showed
that the Microbacterium sp. NISOC-06 was not able to utilize DBT as a carbon source. There was no accumulation of phenolic compounds as Gibb’s assay showed. Biomass
production in a biphasic system for which DBT-enriched gasoline was used as the sulfur source indicated the capability of
Microbacterium sp. NISOC-06 to desulfurize gasoline. 相似文献
20.
M. SantoshkumarY. Veeranagouda Kyoung LeeT.B. Karegoudar 《International biodeterioration & biodegradation》2011,65(1):153-159
A bacterial strain Paracoccus sp. SKG capable of utilizing acetonitrile as a sole source of carbon and nitrogen was isolated from the chemical waste samples. The molecular phylogram generated using the complete sequence of 16S rDNA of the strain SKG showed close links to the bacteria grouped under Brucellaceae family, that belongs to the class alphaproteobacteria. Specifically, the 16S rDNA sequence of strain SKG has shown 99% similarity to Paracoccus sp. This bacterium has also shown impressive growth on aliphatic nitriles like acetonitrile, propionitrile, acrylonitrile, valeronitrile and their corresponding amides. The nitriles degradation has led to the accumulation of ammonia and respective carboxylic acids. The acetonitrile grown cells showed the release of ammonia that contributes to the increase in pH of the medium. However, glucose grown cells failed to produce ammonia, thus indicating the inducible nature of acetonitrile degrading enzymes in Paracoccus sp. SKG. Nitrile hydratase and amidase are the two key enzymes involved in the degradation of acetonitrile. Degradation of acetonitrile in Paracoccus sp. SKG follows the bi-enzymatic pathway. Further, this strain is capable of degrading acetonitrile in the presence of other organic solvents such as methanol, ethanol and dimethylformamide. Therefore, this strain is efficiently used for the treatment of HPLC waste stream containing acetonitrile in the presence of other organic solvents. 相似文献