scholarly journals New Carbenicillin-Hydrolyzing β-Lactamase (CARB-7) from Vibrio cholerae Non-O1, Non-O139 Strains Encoded by the VCR Region of the V. cholerae Genome

2002 ◽  
Vol 46 (7) ◽  
pp. 2162-2168 ◽  
Author(s):  
Roberto Melano ◽  
Alejandro Petroni ◽  
Alicia Garutti ◽  
Héctor Alex Saka ◽  
Laura Mange ◽  
...  
Keyword(s):  
Nucleotide Sequence ◽  
Vibrio Cholerae ◽  
Gene Families ◽  
Chromosome 2 ◽  
Reading Frame ◽  
West Region ◽  
Acid Protein ◽  
Lactamase Gene ◽  
Flanking Regions ◽  
Amino Acid Protein

ABSTRACT In a previous study, an analysis of 77 ampicillin-nonsusceptible (resistant plus intermediate categories) strains of Vibrio cholerae non-O1, non-O139, isolated from aquatic environment and diarrheal stool, showed that all of them produced a β-lactamase with a pI of 5.4. Hybridization or amplification by PCR with a probe for bla TEM or primers for bla CARB gene families was negative. In this work, an environmental ampicillin-resistant strain from this sample, ME11762, isolated from a waterway in the west region of Argentina, was studied. The nucleotide sequence of the structural gene of the β-lactamase was determined by bidirectional sequencing of a Sau3AI fragment belonging to this isolate. The gene encodes a new 288-amino-acid protein, designated CARB-7, that shares 88.5% homology with the CARB-6 enzyme; an overall 83.2% homology with PSE-4, PSE-1, CARB-3, and the Proteus mirabilis N29 enzymes; and 79% homology with CARB-4 enzyme. The gene for this β-lactamase could not be transferred to Escherichia coli by conjugation. The nucleotide sequence of the flanking regions of the bla CARB-7 gene showed the occurrence of three 123-bp V. cholerae repeated sequences, all of which were found outside the predicted open reading frame. The upstream fragment of the bla CARB-7 gene shared 93% identity with a locus situated inside V. cholerae's chromosome 2. These results strongly suggest the chromosomal location of the bla CARB-7 gene, making this the first communication of a β-lactamase gene located on the VCR island of the V. cholerae genome.

scholarly journals Role of rpoS in Stress Survival and Virulence of Vibrio cholerae

1998 ◽  
Vol 180 (4) ◽  
pp. 773-784 ◽  
Author(s):  
Fitnat H. Yildiz ◽  
Gary K. Schoolnik
Keyword(s):  
Vibrio Cholerae ◽  
Genomic Library ◽  
Bacterial Species ◽  
Wild Type ◽  
Reading Frame ◽  
Infant Mouse ◽  
Acid Protein ◽  
Amino Acid Protein

ABSTRACT Vibrio cholerae is known to persist in aquatic environments under nutrient-limiting conditions. To analyze the possible involvement of the alternative sigma factor encoded byrpoS, which is shown to be important for survival during nutrient deprivation in several other bacterial species, a V. cholerae rpoS homolog was cloned by functional complementation of an Escherichia coli mutant by using a wild-type genomic library. Sequence analysis of the complementing clone revealed an 1.008-bp open reading frame which is predicted to encode a 336-amino-acid protein with 71 to 63% overall identity to other reported rpoS gene products. To determine the functional role of rpoS in V. cholerae, we inactivatedrpoS by homologous recombination. V. choleraestrains lacking rpoS are impaired in the ability to survive diverse environmental stresses, including exposure to hydrogen peroxide, hyperosmolarity, and carbon starvation. These results suggest that rpoS may be required for the persistence of V. cholerae in aquatic habitats. In addition, the rpoSmutation led to reduced production or secretion of hemagglutinin/protease. However, rpoS is not critical for in vivo survival, as determined by an infant mouse intestinal competition assay.

scholarly journals Characterization and Nucleotide Sequence of CARB-6, a New Carbenicillin-Hydrolyzing β-Lactamase from Vibrio cholerae

1999 ◽  
Vol 43 (2) ◽  
pp. 297-301 ◽  
Author(s):  
Danièle Choury ◽  
Gérald Aubert ◽  
Marie-France Szajnert ◽  
Kemal Azibi ◽  
Marc Delpech ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Amino Acid ◽  
Nucleotide Sequence ◽  
Vibrio Cholerae ◽  
Plasmid Dna ◽  
Direct Sequencing ◽  
Clinical Strain ◽  
Acid Protein ◽  
K 12 ◽  
Amino Acid Protein

ABSTRACT A clinical strain of Vibrio cholerae non-O1 non-O139 isolated in France produced a new β-lactamase with a pI of 5.35. The purified enzyme, with a molecular mass of 33,000 Da, was characterized. Its kinetic constants show it to be a carbenicillin-hydrolyzing enzyme comparable to the five previously reported CARB β-lactamases and to SAR-1, another carbenicillin-hydrolyzing β-lactamase that has a pI of 4.9 and that is produced by a V. cholerae strain from Tanzania. This β-lactamase is designated CARB-6, and the gene for CARB-6 could not be transferred to Escherichia coli K-12 by conjugation. The nucleotide sequence of the structural gene was determined by direct sequencing of PCR-generated fragments from plasmid DNA with four pairs of primers covering the whole sequence of the reference CARB-3 gene. The gene encodes a 288-amino-acid protein that shares 94% homology with the CARB-1, CARB-2, and CARB-3 enzymes, 93% homology with the Proteus mirabilis N29 enzyme, and 86.5% homology with the CARB-4 enzyme. The sequence of CARB-6 differs from those of CARB-3, CARB-2, CARB-1, N29, and CARB-4 at 15, 16, 17, 19, and 37 amino acid positions, respectively. All these mutations are located in the C-terminal region of the sequence and at the surface of the molecule, according to the crystal structure of theStaphylococcus aureus PC-1 β-lactamase.

scholarly journals The bglA Gene of Aspergillus kawachii Encodes Both Extracellular and Cell Wall-Bound β-Glucosidases

1999 ◽  
Vol 65 (12) ◽  
pp. 5546-5553 ◽  
Author(s):  
Kazuhiro Iwashita ◽  
Tatsuya Nagahara ◽  
Hitoshi Kimura ◽  
Makoto Takano ◽  
Hitoshi Shimoi ◽  
...  
Keyword(s):  
Cell Wall ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Amino Acid Sequences ◽  
Enzyme Assays ◽  
Partial Amino Acid Sequence ◽  
Reading Frame ◽  
Acid Protein ◽  
Aspergillus Kawachii ◽  
Amino Acid Protein

ABSTRACT We cloned the genomic DNA and cDNA of bglA, which encodes β-glucosidase in Aspergillus kawachii, based on a partial amino acid sequence of purified cell wall-bound β-glucosidase CB-1. The nucleotide sequence of the cloned bglA gene revealed a 2,933-bp open reading frame with six introns that encodes an 860-amino-acid protein. Based on the deduced amino acid sequence, we concluded that the bglA gene encodes cell wall-bound β-glucosidase CB-1. The amino acid sequence exhibited high levels of homology with the amino acid sequences of fungal β-glucosidases classified in subfamily B. We expressed the bglA cDNA inSaccharomyces cerevisiae and detected the recombinant β-glucosidase in the periplasm fraction of the recombinant yeast.A. kawachii can produce two extracellular β-glucosidases (EX-1 and EX-2) in addition to the cell wall-bound β-glucosidase.A. kawachii in which the bglA gene was disrupted produced none of the three β-glucosidases, as determined by enzyme assays and a Western blot analysis. Thus, we concluded that thebglA gene encodes both extracellular and cell wall-bound β-glucosidases in A. kawachii.

Molecular cloning, characterization, and expression analysis of a novel sema-2a homologue in Polyrhachis vicina (Hymenoptera: Formicidae)

2010 ◽  
Vol 142 (1) ◽  
pp. 1-13 ◽  
Author(s):  
Jing Luo ◽  
Geng-Si Xi ◽  
Shu-Min Lü ◽  
Ke Li ◽  
Qing Li
Keyword(s):  
Untranslated Region ◽  
Axonal Guidance ◽  
Mrna Levels ◽  
Chain Reaction ◽  
Base Pairs ◽  
Reading Frame ◽  
Acid Protein ◽  
Polymerase Chain ◽  
Polyrhachis Vicina ◽  
Amino Acid Protein

AbstractThe semaphorin gene family plays important roles in axonal guidance in vertebrates and invertebrates. Semaphorin 2a, a member of the semaphorin family, belongs to class 2, which is found only in invertebrates. In our study, semaphorin 2a was cloned from the ant Polyrhachis vicina Roger. The full length of P. vicina semaphorin 2a (Pv-sema-2a) is 2763 base pairs (bp) and it contains a 5′-untranslated region (UTR) 92 bp long and a 3′-UTR 521 bp long. The open reading frame of Pv-sema-2a encodes a 716-amino-acid protein with a predicted molecular mass of 81.1 kilodaltons. Real-time quantitative reverse-transcription – polymerase chain reaction indicated that Pv-sema-2a mRNA is differentially expressed during P. vicina development, in the whole bodies as well as the heads of different castes. The high mRNA levels in embryos and pupae suggest that Pv-sema-2a plays an important role in ant development.

Cloning and characterization of a 2,3-oxidosqualene cyclase from Eleutherococcus trifoliatus

Holzforschung ◽  
2013 ◽  
Vol 67 (4) ◽  
pp. 463-471 ◽  
Author(s):  
Li-Ting Ma ◽  
Sheng-Yang Wang ◽  
Yen-Hsueh Tseng ◽  
Yi-Ru Lee ◽  
Fang-Hua Chu
Keyword(s):  
Physiological Role ◽  
Active Site Residues ◽  
Reading Frame ◽  
Oxidosqualene Cyclases ◽  
Oxidosqualene Cyclase ◽  
Acid Protein ◽  
Leaf Tissues ◽  
New Perspective ◽  
Amino Acid Protein

Abstract The 2,3-oxidosqualene cyclases (OSCs) are a family of enzymes that have an important role in plant triterpene biosynthesis. In this study, an OSC gene designed EtLUS from Eleutherococcus trifoliatus has been cloned. EtLUS includes a 2292-bp open reading frame and encodes a 763-amino acid protein. EtLUS has an MLCYCR motif, which is conserved in lupeol synthases. Comparison of active-site residues and gene expression in yeast showed that EtLUS synthesizes lupeol. However, EtLUS has the highest sequence identity with β-amyrin synthases from Araliaceae rather than lupeol synthases, adding new perspective to the evolution of the OSCs of Araliaceae. Furthermore, EtLUS is upregulated in leaf tissues under methyl jasmonate treatment, which can be interpreted that lupeol and its derivatives play an ecological and physiological role in plant defense against pathogens and insect herbivores.

scholarly journals Nucleotide Sequence of Rice cDNA that Encodes a Ubiquitin Protein and a 79-Amino Acid Protein

1995 ◽  
Vol 108 (2) ◽  
pp. 865-865 ◽  
Author(s):  
H. U. Kim ◽  
C. H. Yun ◽  
W. S. Cho ◽  
S. K. Kang ◽  
T. Y. Chung
Keyword(s):  
Amino Acid ◽  
Nucleotide Sequence ◽  
Acid Protein ◽  
Rice Cdna ◽  
Amino Acid Protein

scholarly journals Identification and characterization of a cDNA encoding mouse CAP: a homolog of the yeast adenylyl cyclase associated protein

1993 ◽  
Vol 105 (3) ◽  
pp. 777-785 ◽  
Author(s):  
A.B. Vojtek ◽  
J.A. Cooper
Keyword(s):  
Adenylyl Cyclase ◽  
Leading Edge ◽  
Northern Analysis ◽  
Reading Frame ◽  
Terminal Domain ◽  
Carboxy Terminal Domain ◽  
Acid Protein ◽  
Carboxy Terminal ◽  
Amino Acid Protein

CAP, an adenylyl cyclase associated protein, is present in Saccharomyces cerevisiae and Schizosaccharomyces pombe. In both organisms, CAP is bifunctional: the N-terminal domain binds to adenylyl cyclase, thereby enabling adenylyl cyclase to respond appropriately to upstream regulatory signals, such as RAS in S. cerevisiae; the C-terminal domain is required for cellular morphogenesis. Here, we describe the isolation of a cDNA encoding a CAP homolog from a higher eukaryote. The mouse CAP cDNA contains an open reading frame capable of encoding a 474 amino acid protein. The protein encoded by the mouse CAP cDNA shows extensive homology to the yeast CAP proteins, particularly in the central poly-proline rich region and in the C-terminal domain. By northern analysis, the CAP message appears to be ubiquitous, but not uniform. By indirect immunofluorescence, ectopically expressed mouse CAP protein is found in the cytoplasm of fibroblasts and, in migrating cells, at the leading edge. Expression of the mouse CAP cDNA in S. cerevisiae complements defects associated with loss of the yeast CAP carboxy-terminal domain. Hence, the function of the CAP carboxy-terminal domain has been conserved from yeast to mouse.

scholarly journals Molecular Cloning and Characterization of a cDNA Encoding Sedoheptulose-1,7-bisphosphatase from Mulberry (Morus alba var. multicaulis)

2008 ◽  
Vol 57 (1-6) ◽  
pp. 152-157 ◽  
Author(s):  
X. Ji ◽  
Y. Gai ◽  
J. Ma ◽  
C. Zheng ◽  
Z. Mu
Keyword(s):  
Morus Alba ◽  
Evolutionary Analysis ◽  
Comparison Analysis ◽  
Reading Frame ◽  
Acid Protein ◽  
Fructose 1,6 Bisphosphatase ◽  
Rapid Amplification ◽  
Molecular Evolutionary Analysis ◽  
Amino Acid Protein

Abstract A full-length cDNA encoding sedoheptulose-1,7-bisphosphatase (SBPase; EC 3.1.3.37) was cloned from mulberry (Morus alba var. multicaulis) by rapid amplification of cDNA ends (RACE). The cDNA consisted of 1,527 nucleotides with an open reading frame (ORF) of 1,179 nucleotides encoding a 393 amino acid protein of approximately 42.6 kDa. Sequence comparison analysis showed that mulberry SBPase (MSBPase) had high homology to other plant counterparts. Phylogenetic and molecular evolutionary analysis revealed that MSBPase fell into plant SBPase group. Moreover, SBPase and fructose-1,6-bisphosphatase (FBPase; EC 3.1.3.11) shared 28-32% identical residues, suggesting that the two enzymes originated from the same evolution branch. Molecular modeling indicated that each subunit of MSBPase was composed of α-helices and β-sheets joined by turns and loops, and folded into a structure of hexahedron shape which was very similar to FBPase.

scholarly journals NOP3 is an essential yeast protein which is required for pre-rRNA processing.

1992 ◽  
Vol 119 (4) ◽  
pp. 737-747 ◽  
Author(s):  
I D Russell ◽  
D Tollervey
Keyword(s):  
Northern Analysis ◽  
Rna Recognition Motif ◽  
Rna Recognition ◽  
Rrna Processing ◽  
Reading Frame ◽  
Repeat Units ◽  
Nucleolar Proteins ◽  
Acid Protein ◽  
Sds Page ◽  
Amino Acid Protein

The four nucleolar proteins NOP1, SSB1, GAR1, and NSR1 of Saccharomyces cerevisiae share a repetitive domain composed of repeat units rich in glycine and arginine (GAR domain). We have cloned and sequenced a fifth member of this family, NOP3, and shown it to be essential for cell viability. The NOP3 open reading frame encodes a 415 amino acid protein with a predicted molecular mass of 45 kD, containing a GAR domain and an RNA recognition motif. NOP3-specific antibodies recognize a 60-kD protein by SDS-PAGE and decorate the nucleolus and the surrounding nucleoplasm. A conditional lethal mutation, GAL::nop3, was constructed; growth of the mutant strain in glucose medium represses NOP3 expression. In cells depleted of NOP3, production of cytoplasmic ribosomes is impaired. Northern analysis and pulse-chase labeling indicate that pre-rRNA processing is inhibited at the late steps, in which 27SB pre-rRNA is cleaved to 25S rRNA and 20S pre-rRNA to 18S rRNA.

scholarly journals Characterization of an Endo-β-1,6-Galactanase from Streptomyces avermitilis NBRC14893

2008 ◽  
Vol 74 (8) ◽  
pp. 2379-2383 ◽  
Author(s):  
Hitomi Ichinose ◽  
Toshihisa Kotake ◽  
Yoichi Tsumuraya ◽  
Satoshi Kaneko
Keyword(s):  
Signal Sequence ◽  
Recombinant Enzyme ◽  
Streptomyces Avermitilis ◽  
Glycoside Hydrolase Family ◽  
Reading Frame ◽  
Secretion Signal ◽  
Acid Protein ◽  
Hydrolysis Of ◽  
Amino Acid Protein

ABSTRACT The putative endo-β-1,6-galactanase gene from Streptomyces avermitilis was cloned and expressed in Escherichia coli, and the enzymatic properties of the recombinant enzyme were characterized. The gene consisted of a 1,476-bp open reading frame and encoded a 491-amino-acid protein, comprising an N-terminal secretion signal sequence and glycoside hydrolase family 5 catalytic module. The recombinant enzyme, Sa1,6Gal5A, catalyzed the hydrolysis of β-1,6-linked galactosyl linkages of oligosaccharides and polysaccharides. The enzyme produced galactose and a range of β-1,6-linked galacto-oligosaccharides, predominantly β-1,6-galactobiose, from β-1,6-galactan chains. There was a synergistic effect between the enzyme and Sa1,3Gal43A in degrading tomato arabinogalactan proteins. These results suggest that Sa1,6Gal5A is the first identified endo-β-1,6-galactanase from a prokaryote.

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