scholarly journals Peptidoglycan Hydrolase LytF Plays a Role in Cell Separation with CwlF during Vegetative Growth of Bacillus subtilis

1999 ◽  
Vol 181 (10) ◽  
pp. 3178-3184 ◽  
Author(s):  
Ryo Ohnishi ◽  
Shu Ishikawa ◽  
Junichi Sekiguchi
Keyword(s):  
Cell Wall ◽  
Catalytic Domain ◽  
Signal Sequence ◽  
Sequence Similarity ◽  
Terminal Region ◽  
Diaminopimelic Acid ◽  
Northern Hybridization ◽  
Peptidoglycan Hydrolase ◽  
High Sequence Similarity ◽  
Insertional Inactivation

ABSTRACT Peptidoglycan hydrolase, LytF (CwlE), was determined to be identical to YhdD (deduced cell wall binding protein) by zymography after insertional inactivation of the yhdD gene. YhdD exhibits high sequence similarity with CwlF (PapQ, LytE) and p60 ofListeria monocytogenes. The N-terminal region of YhdD has a signal sequence followed by five tandem repeated regions containing polyserine residues. The C-terminal region corresponds to the catalytic domain, because a truncated protein without the N-terminal region retained cell wall hydrolase activity. The histidine-tagged LytF protein produced in Escherichia coli cells hydrolyzed the linkage of d-γ-glutamyl-meso-diaminopimelic acid in murein peptides, indicating that it is ad,l-endopeptidase. Northern hybridization and primer extension analyses indicated that the lytF gene was transcribed by EςD RNA polymerase. Disruption oflytF led to slightly filamentous cells, and a lytF cwlF double mutant exhibited extraordinary microfiber formation, which is similar to the cell morphology of the cwlF sigDmutant.

scholarly journals A New d,l-Endopeptidase Gene Product, YojL (Renamed CwlS), Plays a Role in Cell Separation with LytE and LytF in Bacillus subtilis

2006 ◽  
Vol 188 (15) ◽  
pp. 5541-5550 ◽  
Author(s):  
Tatsuya Fukushima ◽  
Anahita Afkham ◽  
Shin-ichirou Kurosawa ◽  
Taichi Tanabe ◽  
Hiroki Yamamoto ◽  
...  
Keyword(s):  
Cell Wall ◽  
Bacillus Subtilis ◽  
Cell Separation ◽  
Sequence Similarity ◽  
Terminal Region ◽  
Diaminopimelic Acid ◽  
Amino Acid Sequence Similarity ◽  
Northern Hybridization ◽  
Triple Mutant ◽  
High Amino Acid

ABSTRACT A new peptidoglycan hydrolase, Bacillus subtilis YojL (cell wall-lytic enzyme associated with cell separation, renamed CwlS), exhibits high amino acid sequence similarity to LytE (CwlF) and LytF (CwlE), which are associated with cell separation. The N-terminal region of CwlS has four tandem repeat regions (LysM repeats) predicted to be a peptidoglycan-binding module. The C-terminal region exhibits high similarity to the cell wall hydrolase domains of LytE and LytF at their C-terminal ends. The C-terminal region of CwlS produced in Escherichia coli could hydrolyze the linkage of d-γ-glutamyl-meso-diaminopimelic acid in the cell wall of B. subtilis, suggesting that CwlS is a d,l-endopeptidase. β-Galactosidase fusion experiments and Northern hybridization analysis suggested that the cwlS gene is transcribed during the late vegetative and early stationary phases. A cwlS mutant exhibited a cell shape similar to that of the wild type; however, a lytE lytF cwlS triple mutant exhibited aggregated microfiber formation. Moreover, immunofluorescence microscopy showed that FLAG-tagged CwlS was localized at cell separation sites and cell poles during the late vegetative phase. The localization sites are similar to those of LytF and LytE, indicating that CwlS is involved in cell separation with LytF and LytE. These specific localizations may be dependent on the LysM repeats in their N-terminal domains. The roles of CwlS, LytF, and LytE in cell separation are discussed.

Acd, a peptidoglycan hydrolase of Clostridium difficile with N-acetylglucosaminidase activity

Microbiology ◽  
2005 ◽  
Vol 151 (7) ◽  
pp. 2343-2351 ◽  
Author(s):  
Anne Dhalluin ◽  
Ingrid Bourgeois ◽  
Martine Pestel-Caron ◽  
Emilie Camiade ◽  
Gregory Raux ◽  
...  
Keyword(s):  
Cell Wall ◽  
Amino Acid ◽  
Clostridium Difficile ◽  
Catalytic Domain ◽  
Sequence Similarity ◽  
Amino Acid Sequences ◽  
Cellular Growth ◽  
Peptidoglycan Hydrolase ◽  
Gene Encoding ◽  
Terminal Domain

A gene encoding a putative peptidoglycan hydrolase was identified by sequence similarity searching in the Clostridium difficile 630 genome sequence, and the corresponding protein, named Acd (autolysin of C. difficile) was expressed in Escherichia coli. The deduced amino acid sequence of Acd shows a modular structure with two main domains: an N-terminal domain exhibiting repeated sequences and a C-terminal catalytic domain. The C-terminal domain exhibits sequence similarity with the glucosaminidase domains of Staphylococcus aureus Atl and Bacillus subtilis LytD autolysins. Purified recombinant Acd produced in E. coli was confirmed to be a cell-wall hydrolase with lytic activity on the peptidoglycan of several Gram-positive bacteria, including C. difficile. The hydrolytic specificity of Acd was studied by RP-HPLC analysis and MALDI-TOF MS using B. subtilis cell-wall extracts. Muropeptides generated by Acd hydrolysis demonstrated that Acd hydrolyses peptidoglycan bonds between N-acetylglucosamine and N-acetylmuramic acid, confirming that Acd is an N-acetylglucosaminidase. The transcription of the acd gene increased during vegetative cellular growth of C. difficile 630. The sequence of the acd gene appears highly conserved in C. difficile strains. Regarding deduced amino acid sequences, the C-terminal domain with enzymic function appears to be the most conserved of the two main domains. Acd is the first known autolysin involved in peptidoglycan hydrolysis of C. difficile.

Amycolatopsis dongchuanensis sp. nov., an actinobacterium isolated from soil

2012 ◽  
Vol 62 (Pt_11) ◽  
pp. 2650-2656 ◽  
Author(s):  
Guo-Xing Nie ◽  
Hong Ming ◽  
Shuai Li ◽  
En-Min Zhou ◽  
Juan Cheng ◽  
...  
Keyword(s):  
Cell Wall ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Diaminopimelic Acid ◽  
Western China ◽  
Rrna Gene ◽  
The Novel ◽  
Biochemical Tests ◽  
Content Type ◽  
Link Type

A novel actinomycete strain, designated YIM 75904T, was isolated from a soil sample that had been collected from a dry and hot river valley in Dongchuan county, Yunnan province, south-western China. The taxonomic position of the novel strain was investigated by a polyphasic approach. In phylogenetic analyses based on 16S rRNA gene sequences, strain YIM 75904T formed a distinct clade within the genus Amycolatopsis and appeared to be closely related to Amycolatopsis sacchari K24T (99.3 % sequence similarity). Strain YIM 75904T had a type-IV cell wall, with no detectable mycolic acids, and had MK-9(H4) as its predominant menaquonine. Its cell wall contained meso-diaminopimelic acid, galactose, glucose and arabinose, and its major cellular fatty acids were iso-C16 : 0, iso-C15 : 0, anteiso-C17 : 0 and anteiso-C15 : 0. The genomic DNA G+C content of the novel strain was 68.5 mol%. Based on the results of physiological and biochemical tests and DNA–DNA hybridizations, strain YIM 75904T represents a novel species of the genus Amycolatopsis for which the name Amycolatopsis dongchuanensis sp. nov. is proposed. The type strain is YIM 75904T ( = CCTCC AA 2011016T  = JCM 18054T).

scholarly journals Coraliomargarita akajimensis gen. nov., sp. nov., a novel member of the phylum ‘Verrucomicrobia’ isolated from seawater in Japan

2007 ◽  
Vol 57 (5) ◽  
pp. 959-963 ◽  
Author(s):  
Jaewoo Yoon ◽  
Mina Yasumoto-Hirose ◽  
Atsuko Katsuta ◽  
Hiroshi Sekiguchi ◽  
Satoru Matsuda ◽  
...  
Keyword(s):  
Cell Wall ◽  
New Genus ◽  
Gene Sequence ◽  
Novel Species ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Diaminopimelic Acid ◽  
Rrna Gene ◽  
Hard Coral ◽  
The 16S Rrna Gene

An obligately aerobic, Gram-negative, non-spore-forming, non-motile, spherical bacterium, designated strain 04OKA010-24T, was isolated from seawater surrounding the hard coral Galaxea fascicularis L., collected at Majanohama, Akajima, Japan, and was subjected to a polyphasic taxonomic study. Phylogenetic analyses based on the 16S rRNA gene sequence indicated that the new strain represented a member of the phylum ‘Verrucomicrobia’ and shared 84–95 % sequence similarity with cultivated strains of ‘Verrucomicrobia’ subdivision 4. Amino acid analysis of the cell-wall hydrolysate indicated the absence of muramic acid and diaminopimelic acid, which suggested that the strain did not contain peptidoglycan in the cell wall. The G+C content of the DNA was 53.9 mol%. MK-7 was the major menaquinone and C14 : 0, C18 : 1 ω9c and C18 : 0 were the major fatty acids. On the basis of these data, it was concluded that strain 04OKA010-24T represents a novel species in a new genus in subdivision 4 of the phylum ‘Verrucomicrobia’, for which the name Coraliomargarita akajimensis gen. nov., sp. nov. is proposed. The type strain of Coraliomargarita akajimensis is 04OKA010-24T (=MBIC06463T=IAM 15411T=KCTC 12865T).

scholarly journals Pelagicoccus croceus sp. nov., a novel marine member of the family Puniceicoccaceae within the phylum ‘Verrucomicrobia’ isolated from seagrass

2007 ◽  
Vol 57 (12) ◽  
pp. 2874-2880 ◽  
Author(s):  
Jaewoo Yoon ◽  
Naoya Oku ◽  
Satoru Matsuda ◽  
Hiroaki Kasai ◽  
Akira Yokota
Keyword(s):  
Cell Wall ◽  
Novel Species ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Diaminopimelic Acid ◽  
Rrna Gene ◽  
The Novel ◽  
Antibiotic Susceptibility Test ◽  
The Family ◽  
Lactam Antibiotic

An obligately aerobic, spherical, non-motile, pale-yellow pigmented bacterium was isolated from a piece of leaf of seagrass, Enhalus acoroides (L.f.) Royle, grown in Okinawa, Japan and was subjected to a polyphasic taxonomic study. Phylogenetic analyses based on 16S rRNA gene sequences revealed that the novel isolate N5FB36-5T shared approximately 96–98 % sequence similarity with the species of the genus Pelagicoccus of the family Puniceicoccaceae within the phylum ‘Verrucomicrobia’. The DNA–DNA relatedness values of strain N5FB36-5T with Pelagicoccus mobilis 02PA-Ca-133T and Pelagicoccus albus YM14-201T were below 70 %, which is accepted as the phylogenetic definition of a novel species. β-Lactam antibiotic susceptibility test and amino acid analysis of the cell wall hydrolysates indicated the absence of muramic acid and diaminopimelic acid in the cell walls, which suggested that this strain lacks an ordinary Gram-negative type of peptidoglycan in the cell wall. The DNA G+C content of strain N5FB36-5T was 51.6 mol%; MK-7 was the major menaquinone; and the presence of C16 : 0, C16 : 1 ω7c and anteiso-C15 : 0 as the major cellular fatty acids supported the identification of the novel isolate as a member of the genus Pelagicoccus. On the basis of polyphasic taxonomic data, it was concluded that this strain should be classified as a novel species of the genus Pelagicoccus, for which the name Pelagicoccus croceus sp. nov. is proposed. The type strain is N5FB36-5T (=MBIC08282T=KCTC 12903T).

scholarly journals Distribution and Phylogenetic Analysis of Family 19 Chitinases in Actinobacteria

2004 ◽  
Vol 70 (2) ◽  
pp. 1135-1144 ◽  
Author(s):  
Tomokazu Kawase ◽  
Akihiro Saito ◽  
Toshiya Sato ◽  
Ryo Kanai ◽  
Takeshi Fujii ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Catalytic Domain ◽  
Higher Plants ◽  
Sequence Similarity ◽  
Evolutionary Relationship ◽  
Pcr Amplification ◽  
High Sequence Similarity ◽  
The Family ◽  
Family 19 Chitinase ◽  
Chitinase Genes

ABSTRACT In organisms other than higher plants, family 19 chitinase was first discovered in Streptomyces griseus HUT6037, and later, the general occurrence of this enzyme in Streptomyces species was demonstrated. In the present study, the distribution of family 19 chitinases in the class Actinobacteria and the phylogenetic relationship of Actinobacteria family 19 chitinases with family 19 chitinases of other organisms were investigated. Forty-nine strains were chosen to cover almost all the suborders of the class Actinobacteria, and chitinase production was examined. Of the 49 strains, 22 formed cleared zones on agar plates containing colloidal chitin and thus appeared to produce chitinases. These 22 chitinase-positive strains were subjected to Southern hybridization analysis by using a labeled DNA fragment corresponding to the catalytic domain of ChiC, and the presence of genes similar to chiC of S. griseus HUT6037 in at least 13 strains was suggested by the results. PCR amplification and sequencing of the DNA fragments corresponding to the major part of the catalytic domains of the family 19 chitinase genes confirmed the presence of family 19 chitinase genes in these 13 strains. The strains possessing family 19 chitinase genes belong to 6 of the 10 suborders in the order Actinomycetales, which account for the greatest part of the Actinobacteria. Phylogenetic analysis suggested that there is a close evolutionary relationship between family 19 chitinases found in Actinobacteria and plant class IV chitinases. The general occurrence of family 19 chitinase genes in Streptomycineae and the high sequence similarity among the genes found in Actinobacteria suggest that the family 19 chitinase gene was first acquired by an ancestor of the Streptomycineae and spread among the Actinobacteria through horizontal gene transfer.

scholarly journals Insights into Substrate Specificity of NlpC/P60 Cell Wall Hydrolases Containing Bacterial SH3 Domains

mBio ◽  
2015 ◽  
Vol 6 (5) ◽  
Author(s):  
Qingping Xu ◽  
Dominique Mengin-Lecreulx ◽  
Xueqian W. Liu ◽  
Delphine Patin ◽  
Carol L. Farr ◽  
...  
Keyword(s):  
Cell Wall ◽  
Amino Acid ◽  
Substrate Specificity ◽  
Catalytic Domain ◽  
Substrate Binding ◽  
Diaminopimelic Acid ◽  
Single Mutation ◽  
Molecular Architecture ◽  
Substrate Specificities ◽  
Cell Wall Hydrolases

ABSTRACTBacterial SH3 (SH3b) domains are commonly fused with papain-like Nlp/P60 cell wall hydrolase domains. To understand how the modular architecture of SH3b and NlpC/P60 affects the activity of the catalytic domain, three putative NlpC/P60 cell wall hydrolases were biochemically and structurally characterized. These enzymes all have γ-d-Glu-A2pm (A2pm is diaminopimelic acid) cysteine amidase (ordl-endopeptidase) activities but with different substrate specificities. One enzyme is a cell wall lysin that cleaves peptidoglycan (PG), while the other two are cell wall recycling enzymes that only cleave stem peptides with an N-terminall-Ala. Their crystal structures revealed a highly conserved structure consisting of two SH3b domains and a C-terminal NlpC/P60 catalytic domain, despite very low sequence identity. Interestingly, loops from the first SH3b domain dock into the ends of the active site groove of the catalytic domain, remodel the substrate binding site, and modulate substrate specificity. Two amino acid differences at the domain interface alter the substrate binding specificity in favor of stem peptides in recycling enzymes, whereas the SH3b domain may extend the peptidoglycan binding surface in the cell wall lysins. Remarkably, the cell wall lysin can be converted into a recycling enzyme with a single mutation.IMPORTANCEPeptidoglycan is a meshlike polymer that envelops the bacterial plasma membrane and bestows structural integrity. Cell wall lysins and recycling enzymes are part of a set of lytic enzymes that target covalent bonds connecting the amino acid and amino sugar building blocks of the PG network. These hydrolases are involved in processes such as cell growth and division, autolysis, invasion, and PG turnover and recycling. To avoid cleavage of unintended substrates, these enzymes have very selective substrate specificities. Our biochemical and structural analysis of three modular NlpC/P60 hydrolases, one lysin, and two recycling enzymes, show that they may have evolved from a common molecular architecture, where the substrate preference is modulated by local changes. These results also suggest that new pathways for recycling PG turnover products, such as tracheal cytotoxin, may have evolved in bacteria in the human gut microbiome that involve NlpC/P60 cell wall hydrolases.

scholarly journals Description of Domibacillus indicus sp. nov., isolated from ocean sediments and emended description of the genus Domibacillus

2014 ◽  
Vol 64 (Pt_9) ◽  
pp. 3010-3015 ◽  
Author(s):  
Avinash Sharma ◽  
Sunil Kumar Dhar ◽  
Om Prakash ◽  
Venkata Ramana Vemuluri ◽  
Vishal Thite ◽  
...  
Keyword(s):  
Cell Wall ◽  
Type Species ◽  
Sequence Similarity ◽  
Diaminopimelic Acid ◽  
Rrna Gene ◽  
Polar Lipid Profile ◽  
Content Type ◽  
Link Type ◽  
Emended Description ◽  
Marine Sediment Sample

A novel Gram-stain-positive, spore-forming, aerobic, non-motile, rod-shaped bacterium designated strain SD111T that forms red-pigmented colonies was isolated from a marine sediment sample (collected from 5 m depth) from Lakshadweep, India. Strain SD111T grew well on seawater agar at pH 6–10 (optimum pH 7.5±0.2). It showed maximum (97.6 %) 16S rRNA gene sequence similarity and formed a monophyletic clade with Domibacillus robiginosus WS 4628T ( = DSM 25058T). The genomic DNA G+C content was 37.4 mol% and the strain showed 37.7 % DNA–DNA relatedness to D. robiginosus DSM 25058T. The major fatty acids were anteiso-C15 : 0, C16 : 0, iso-C15 : 0 and iso-C16 : 0 and MK-6 was the predominant quinone. The polar lipid profile of strain SD111T consisted of unidentified phospholipids (PL1 and PL2), phosphatidylglycerol (PG) and diphosphatidylglycerol (DPG). The cell wall contained meso-diaminopimelic acid and the peptidoglycan was of A1γ type. Glucose and ribose were detected as major cell-wall sugars. Results from polyphasic studies indicated that SD111T represents a novel species of the genus Domibacillus for which the name Domibacillus indicus sp. nov. is proposed. The type strain is SD111T ( = MCC 2255T = DSM 28032T).

scholarly journals Regulation of a New Cell Wall Hydrolase Gene,cwlF, Which Affects Cell Separation in Bacillus subtilis

1998 ◽  
Vol 180 (9) ◽  
pp. 2549-2555 ◽  
Author(s):  
Shu Ishikawa ◽  
Yoshiko Hara ◽  
Ryo Ohnishi ◽  
Junichi Sekiguchi
Keyword(s):  
Cell Wall ◽  
Bacillus Subtilis ◽  
Rna Polymerase ◽  
Sequence Similarity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Exponential Growth Phase ◽  
High Sequence Similarity ◽  
Cell Wall Hydrolase ◽  
Mutant Cells

ABSTRACT Bacillus subtilis produces a 35-kDa cell wall hydrolase, CwlF, during vegetative growth. The CwlF protein was extracted from B. subtilis cwlB sigD mutant cells and separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. N-terminal amino acid sequencing revealed that its sequence is completely identical to that of the internal region of thepapQ gene product. Disruption of the papQ gene in the B. subtilis chromosome led to the complete loss of CwlF, indicating that papQ is identical tocwlF. CwlF exhibits high sequence similarity to the p60 proteins of Listeria species, NlpC proteins ofEscherichia coli and Haemophilus influenzae, and Enp2 protein of Bacillus sphaericus. The β-galactosidase activity of the cwlF-lacZ transcriptional fusion and Northern blot analysis of the cwlF gene indicated that the gene is expressed as a monocistronic operon during the exponential growth phase, and primer extension analysis suggested that the cwlF gene is transcribed mainly by EςA RNA polymerase and weakly by EςH RNA polymerase. While the cells of the cwlF-deficient mutant were about twice as long as those of the wild-type strain, the cwlF sigD double mutant cells exhibited extraordinary microfiber formation, in contrast to the filamentation of the sigD mutant. The CwlF production was not affected by the pleiotropic mutationsflaD1 and degU32(Hy), which endow cells with the ability of extensive filamentation.

scholarly journals Humicoccus flavidus gen. nov., sp. nov., isolated from soil

2007 ◽  
Vol 57 (1) ◽  
pp. 56-59 ◽  
Author(s):  
Jung-Hoon Yoon ◽  
So-Jung Kang ◽  
Seo-Youn Jung ◽  
Tae-Kwang Oh
Keyword(s):  
Fatty Acids ◽  
Cell Wall ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Diaminopimelic Acid ◽  
Rrna Gene ◽  
Polar Lipid Profile ◽  
Diamino Acid

A Gram-positive, non-motile, spherical, non-spore-forming bacterial strain, DS-52T, was isolated from soil from Dokdo, Korea, and its taxonomic position was investigated by using a polyphasic approach. It grew optimally at 25 °C and pH 6.0–7.0. Strain DS-52T had meso-diaminopimelic acid as the diagnostic diamino acid in the cell-wall peptidoglycan, and galactose, mannose, xylose and rhamnose as whole-cell sugars. It contained MK-8(H4) and MK-9(H4) as the predominant menaquinones and anteiso-C15 : 0, iso-C15 : 0 and C17 : 0 as major fatty acids. Major polar lipids were diphosphatidylglycerol, phosphatidylethanolamine and phosphatidyldimethylethanolamine. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain DS-52T is most closely related to the genus Nakamurella of the suborder Frankineae. Strain DS-52T exhibited 16S rRNA gene sequence similarity values of 96.5 % to Nakamurella multipartita JCM 9543T and 92.0–93.9 % to other members of the suborder Frankineae. The diagnostic diamino acid type and polar lipid profile of strain DS-52T were the same as those of the genus Nakamurella. However, strain DS-52T could be clearly distinguished from the genus Nakamurella by differences in predominant menaquinones, major fatty acids and cell-wall sugars. Accordingly, based on combined phenotypic, chemotaxonomic and phylogenetic data, strain DS-52T (=KCTC 19127T=CIP 108919T) is proposed as the type strain of a novel species in a new genus, Humicoccus flavidus gen. nov., sp. nov.

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