RRC ID 33556
Author Fukushima T, Yao Y, Kitajima T, Yamamoto H, Sekiguchi J.
Title Characterization of new L,D-endopeptidase gene product CwlK (previous YcdD) that hydrolyzes peptidoglycan in Bacillus subtilis.
Journal Mol Genet Genomics
Abstract Bacillus subtilis has various cell wall hydrolases, however, the functions and hydrolase activities of some enzymes are still unknown. B. subtilis CwlK (YcdD) exhibits high sequence similarity with the peptidoglycan hydrolytic L,D-endopeptidase (PLY500) of Listeria monocytogenes phage and CwlK has the VanY motif which is a D-alanyl-D-alanine carboxypeptidase (Pfam: http://www.sanger.ac.uk/Software/Pfam/). The beta-galactosidase activity observed on cwlK-lacZ fusion indicated that the cwlK gene was expressed during the vegetative growth phase, and Western blotting suggested that CwlK seems to be localized in the membrane. Truncated CwlK fused with a histidine-tag (h-DeltaCwlK) was produced in Escherichia coli and purified on a nickel column. The h-DeltaCwlK protein hydrolyzed the peptidoglycan of B. subtilis, and the optimal pH, temperature and NaCl concentration for h-DeltaCwlK were pH 6.5, 37 degrees C, and 0 M, respectively. Interestingly, h-DeltaCwlK could hydrolyze the linkage of L-alanine-D-glutamic acid in the stem of the peptidoglycan, however, this enzyme could not hydrolyze the linkage of D-alanine-D-alanine, suggesting that CwlK is an L,D-endopeptidase not a D,D-carboxypeptidase. CwlK could not hydrolyze polyglutamate from B. natto or peptidoglycan of Staphylococcus aureus. This is the first report describing the characterization of an L,D-endopeptidase in B. subtilis and also the first report in bacteria of the characterization of a PLY500 family protein encoded in chromosomal DNA.
Volume 278(4)
Pages 371-83
Published 2007-10-1
DOI 10.1007/s00438-007-0255-8
PMID 17588176
MeSH Amino Acid Sequence Bacillus subtilis / cytology Bacillus subtilis / enzymology* Bacillus subtilis / genetics Bacterial Proteins / genetics Bacterial Proteins / metabolism Carbohydrate Sequence Endopeptidases / genetics* Endopeptidases / metabolism Hydrolysis Models, Biological Molecular Sequence Data Organisms, Genetically Modified Peptidoglycan / metabolism* Sequence Homology, Amino Acid Staphylococcus aureus / chemistry Staphylococcus aureus / metabolism Staphylococcus aureus / ultrastructure Transcription, Genetic
IF 2.879
Times Cited 34
Prokaryotes B. subtilis MBS48 MBS49 MBS50 MBS51