Direct cloning of genes encoding novel xylanases from the human gut

2005 ◽  
Vol 51 (3) ◽  
pp. 251-259 ◽  
Author(s):  
Hidenori Hayashi ◽  
Takashi Abe ◽  
Mitsuo Sakamoto ◽  
Hiroki Ohara ◽  
Toshimichi Ikemura ◽  
...  
Keyword(s):  
Intestinal Bacteria ◽  
Fecal Microbiota ◽  
Coli Strain ◽  
Amino Acid Sequences ◽  
Self Organizing Map ◽  
Human Gut ◽  
Gene Encoding ◽  
Genes Encoding ◽  
Ph Range ◽  
Self Organizing

The aim of this study was to identify a novel 1,4-β-xylanase gene from the mixed genome DNA of human fecal bacteria without bacterial cultivation. Total DNA was isolated from a population of bacteria extracted from fecal microbiota. Using PCR, the gene fragments encoding 5 different family 10 xylanases (xyn10A, xyn10B, xyn10C, xyn10D, and xyn10E) were found. Amino acid sequences deduced from these genes were highly homologous with those of xylanases from anaerobic intestinal bacteria such as Bacteroides spp. and Prevotella spp. Self-organizing map (SOM) analysis revealed that xynA10 was classified into Bacteroidetes. To confirm that one of these genes encodes an active enzyme, a full-length xyn10A gene was obtained using nested primers specific to the internal fragments and random primers. The xyn10A gene encoding the xylanase Xyn10A consists of 1146 bp and encodes a protein of 382 amino acids and a molecular weight of 43 552. Xyn10A was a single module novel xylanase. Xyn10A was purified from a recombinant Escherichia coli strain and characterized. This enzyme was optimally active at 40 °C and stable up to 50 °C at pH 6.5 and over the pH range 4.0–11.0 at 25 °C. In addition, 2 ORFs (ORF1 and ORF2) were identified upstream of xyn10A. These results suggested that many unidentified xylanolytic bacteria exist in the human gut and may contribute to the breakdown of xylan which contains dietary fiber.Key words: xylanase, human gut, fecal microbiota, phylogenetic analysis, self-organizing map.

scholarly journals Two genes encoding ‘minor’ legumin polypeptides in pea (Pisum sativum L.). Characterization and complete sequence of the LegJ gene

1988 ◽  
Vol 250 (1) ◽  
pp. 15-24 ◽  
Author(s):  
J A Gatehouse ◽  
D Bown ◽  
J Gilroy ◽  
M Levasseur ◽  
J Castleton ◽  
...  
Keyword(s):  
Pisum Sativum ◽  
Complete Sequence ◽  
Genomic Clone ◽  
Amino Acid Sequences ◽  
Flanking Sequence ◽  
Gene Encoding ◽  
Sequence Comparisons ◽  
Control Of Gene Expression ◽  
Pisum Sativum L ◽  
Genes Encoding

A genomic clone from pea (Pisum sativum L.) contains all of one gene encoding a ‘minor’ (B-type) legumin polypeptide, and most of a second very similar gene. The two genes, designated LegJ and LegK, are arranged in tandem, separated by approx. 6 kb. A complete sequence of gene LegJ and its flanking sequences is given, with as much of the sequence of gene LegK as is present on the genomic clone. Hybridization of 3′ flanking sequence probes to seed mRNA, and sequence comparisons with cDNA species, suggested that gene LegJ, and probably gene LegK, was expressed. The partial amino acid sequences of ‘minor’ legumin α- and beta-polypeptides were used to confirm the identity of these genes. The transciption start in gene LegJ was mapped. The 5′ flanking sequence of gene LegJ contains a sequence conserved in legumin genes from pea and other species, which is likely to have functional significance in control of gene expression. Sequence comparisons with legumin genes and cDNA species from Vicia faba and soya bean show that separation of legumin genes into A- and B-type subfamilies occurred before separation of the Viciae and Glycinae tribes.

scholarly journals Multivariate Analysis in Microbiome Description: Correlation of Human Gut Protein Degraders, Metabolites, and Predicted Metabolic Functions

2021 ◽  
Vol 12 ◽  
Author(s):  
Stefano Raimondi ◽  
Rosalba Calvini ◽  
Francesco Candeliere ◽  
Alan Leonardi ◽  
Alessandro Ulrici ◽  
...  
Keyword(s):  
Intestinal Bacteria ◽  
Fecal Microbiota ◽  
Sensu Stricto ◽  
Gene Profiling ◽  
Rrna Gene ◽  
Linear Discriminant ◽  
Metabolic Functions ◽  
Genes Encoding ◽  
The Face ◽  
Microbiome Data

Protein catabolism by intestinal bacteria is infamous for releasing many harmful compounds, negatively affecting the health status, both locally and systemically. In a previous study, we enriched in protein degraders the fecal microbiota of five subjects, utilizing a medium containing protein and peptides as sole fermentable substrates and we monitored their evolution by 16S rRNA gene profiling. In the present study, we fused the microbiome data and the data obtained by the analysis of the volatile organic compounds (VOCs) in the headspace of the cultures. Then, we utilized ANOVA simultaneous component analysis (ASCA) to establish a relationship between metabolites and bacteria. In particular, ASCA allowed to separately assess the effect of subject, time, inoculum concentration, and their binary interactions on both microbiome and volatilome data. All the ASCA submodels pointed out a consistent association between indole and Escherichia–Shigella, and the relationship of butyric, 3-methyl butanoic, and benzenepropanoic acids with some bacterial taxa that were major determinants of cultures at 6 h, such as Lachnoclostridiaceae (Lachnoclostridium), Clostridiaceae (Clostridium sensu stricto), and Sutterellaceae (Sutterella and Parasutterella). The metagenome reconstruction with PICRUSt2 and its functional annotation indicated that enrichment in a protein-based medium affected the richness and diversity of functional profiles, in the face of a decrease of richness and evenness of the microbial community. Linear discriminant analysis (LDA) effect size indicated a positive differential abundance (p < 0.05) for the modules of amino acid catabolism that may be at the basis of the changes of VOC profile. In particular, predicted genes encoding functions belonging to the superpathways of ornithine, arginine, and putrescine transformation to GABA and eventually to succinyl-CoA, of methionine degradation, and various routes of breakdown of aromatic compounds yielding succinyl-CoA or acetyl-CoA became significantly more abundant in the metagenome of the bacterial community.

scholarly journals RegR Virulence Regulon of Rabbit-Specific Enteropathogenic Escherichia coli Strain E22

2013 ◽  
Vol 81 (4) ◽  
pp. 1078-1089 ◽  
Author(s):  
Yogitha N. Srikhanta ◽  
Dianna M. Hocking ◽  
Judyta Praszkier ◽  
Matthew J. Wakefield ◽  
Roy M. Robins-Browne ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Regulatory Protein ◽  
Bioinformatic Analysis ◽  
Coli Strain ◽  
Mobility Shift ◽  
Gene Encoding ◽  
Gene Target ◽  
Content Type ◽  
E Coli ◽  
Genes Encoding

ABSTRACTAraC-like regulators play a key role in the expression of virulence factors in enteric pathogens, such as enteropathogenicEscherichia coli(EPEC), enterotoxigenicE. coli, enteroaggregativeE. coli, andCitrobacter rodentium. Bioinformatic analysis of the genome of rabbit-specific EPEC (REPEC) strain E22 (O103:H2) revealed the presence of a gene encoding an AraC-like regulatory protein, RegR, which shares 71% identity to the global virulence regulator, RegA, ofC. rodentium. Microarray analysis demonstrated that RegR exerts 25- to 400-fold activation on transcription of several genes encoding putative virulence-associated factors, including a fimbrial operon (SEF14), a serine protease, and an autotransporter adhesin. These observations were confirmed by proteomic analysis of secreted and heat-extracted surface-associated proteins. The mechanism of RegR-mediated activation was investigated by using its most highly upregulated gene target,sefA. Transcriptional analyses and electrophoretic mobility shift assays showed that RegR activates the expression ofsefAby binding to a region upstream of thesefApromoter, thereby relieving gene silencing by the global regulatory protein H-NS. Moreover, RegR was found to contribute significantly to virulence in a rabbit infection experiment. Taken together, our findings indicate that RegR controls the expression of a series of accessory adhesins that significantly enhance the virulence of REPEC strain E22.

scholarly journals LT-IIc, a New Member of the Type II Heat-Labile Enterotoxin Family Encoded by an Escherichia coli Strain Obtained from a Nonmammalian Host

2010 ◽  
Vol 78 (11) ◽  
pp. 4705-4713 ◽  
Author(s):  
Hesham F. Nawar ◽  
Natalie D. King-Lyons ◽  
John C. Hu ◽  
Raymond C. Pasek ◽  
Terry D. Connell
Keyword(s):  
Escherichia Coli ◽  
Coli Strain ◽  
Amino Acid Sequences ◽  
Type I ◽  
Avian Host ◽  
Type Ii ◽  
E Coli ◽  
Heat Labile ◽  
Genes Encoding ◽  
New Type

ABSTRACT Two families of bacterial heat-labile enterotoxins (HLTs) have been described: the type I HLTs are comprised of cholera toxin (CT) of Vibrio cholerae, LT-I of Escherichia coli, and several related HLTs; the type II HLTs are comprised of LT-IIa and LT-IIb. Herein, we report LT-IIc, a new type II HLT encoded from an enterotoxigenic E. coli (ETEC) strain isolated from an avian host. Using a mouse Y1 adrenal cell bioassay, LT-IIc was shown to be less cytotoxic than CT, LT-IIa, or LT-IIb. Cytotoxicity of LT-IIc was partially neutralized by antisera recognizing LT-IIa or LT-IIb but not by anti-CT antiserum. Genes encoding putative A polypeptide and B polypeptides of LT-IIc were arranged in an operon which was flanked by potential prophage sequences. Analysis of the nucleotide and predicted amino acid sequences demonstrated that the A polypeptide of LT-IIc has moderate homology to the A polypeptides of CT and LT-I and high homology to the A polypeptides of LT-IIa and LT-IIb. The B polypeptide of LT-IIc exhibited no significant homology to the B polypeptides of CT and LT-I and only moderate homology to the B polypeptides of LT-IIa and LT-IIb. The binding pattern of LT-IIc for gangliosides was distinctive from that of either LT-IIa or LT-IIb. The data suggest that other types of the type II HLT subfamily are circulating in the environment and that host specificity of type II HLT is likely governed by changes in the B polypeptide which mediate binding to receptors.

scholarly journals Distribution of Genes Encoding the Trypsin-Dependent Lantibiotic Ruminococcin A among Bacteria Isolated from Human Fecal Microbiota

2002 ◽  
Vol 68 (7) ◽  
pp. 3424-3431 ◽  
Author(s):  
F. Marcille ◽  
A. Gomez ◽  
P. Joubert ◽  
M. Ladiré ◽  
G. Veau ◽  
...  
Keyword(s):  
Positive Response ◽  
Fecal Microbiota ◽  
Phylogenetic Group ◽  
Peptide Sequence ◽  
Amino Acid Difference ◽  
Bacterial Strains ◽  
Gene Encoding ◽  
Operational Taxonomic Units ◽  
Genes Encoding ◽  
Adults And Children

ABSTRACT Fourteen bacterial strains capable of producing a trypsin-dependent antimicrobial substance active against Clostridium perfringens were isolated from human fecal samples of various origins (from healthy adults and children, as well as from adults with chronic pouchitis). Identification of these strains showed that they belonged to Ruminococcus gnavus, Clostridium nexile, and Ruminococcus hansenii species or to new operational taxonomic units, all from the Clostridium coccoides phylogenetic group. In hybridization experiments with a probe specific for the structural gene encoding the trypsin-dependent lantibiotic ruminococcin A (RumA) produced by R. gnavus, seven strains gave a positive response. All of them harbored three highly conserved copies of rumA-like genes. The deduced peptide sequence was identical to or showed one amino acid difference from the hypothetical precursor of RumA. Our results indicate that the rumA-like genes have been disseminated among R. gnavus and phylogenetically related strains that can make up a significant part of the human fecal microbiota.

Auscultating Diagnosis for Hemodialysis Shunt Stenosis using a Self-Organizing Map and Hidden Markov Model

2012 ◽  
Vol 132 (10) ◽  
pp. 1589-1594 ◽  
Author(s):  
Hayato Waki ◽  
Yutaka Suzuki ◽  
Osamu Sakata ◽  
Mizuya Fukasawa ◽  
Hatsuhiro Kato
Keyword(s):  
Markov Model ◽  
Hidden Markov Model ◽  
Hidden Markov ◽  
Self Organizing Map ◽  
Self Organizing

An Auscultaiting Diagnosis Support System for Assessing Hemodialysis Shunt Stenosis by Using Self-organizing Map

2011 ◽  
Vol 131 (1) ◽  
pp. 160-166 ◽  
Author(s):  
Yutaka Suzuki ◽  
Mizuya Fukasawa ◽  
Osamu Sakata ◽  
Hatsuhiro Kato ◽  
Asobu Hattori ◽  
...  
Keyword(s):  
Support System ◽  
Self Organizing Map ◽  
Self Organizing

Classification of Heat Wave Events in Seoul Using Self-Organizing Map

2018 ◽  
Vol 9 (3) ◽  
pp. 209-221 ◽  
Author(s):  
Seung-Yoon Back ◽  
Sang-Wook Kim ◽  
Myung-Il Jung ◽  
Joon-Woo Roh ◽  
Seok-Woo Son
Keyword(s):  
Heat Wave ◽  
Self Organizing Map ◽  
Self Organizing
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