| Functional and biochemical analysis of the Chlamydia trachomatis ligase MurE. | |
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MedLine Citation:
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PMID: 19820100 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Chlamydiae are unusual obligately intracellular bacteria that do not synthesize detectable peptidoglycan. However, they possess genes that appear to encode products with peptidoglycan biosynthetic activity. Bioinformatic analysis predicts that chlamydial MurE possesses UDP-MurNAc-L-Ala-D-Glu:meso-diaminopimelic acid (UDP-MurNAc-L-Ala-D-Glu:meso-A(2)pm) ligase activity. Nevertheless, there are no experimental data to confirm this hypothesis. In this paper we demonstrate that the murE gene from Chlamydia trachomatis is capable of complementing a conditional Escherichia coli mutant impaired in UDP-MurNAc-L-Ala-D-Glu:meso-A(2)pm ligase activity. Recombinant MurE from C. trachomatis (MurE(Ct)) was overproduced in and purified from E. coli in order to investigate its kinetic parameters in vitro. By use of UDP-MurNAc-L-Ala-D-Glu as the nucleotide substrate, MurE(Ct) demonstrated ATP-dependent meso-A(2)pm ligase activity with pH and magnesium ion optima of 8.6 and 30 mM, respectively. Other amino acids (meso-lanthionine, the ll and dd isomers of A(2)pm, D-lysine) were also recognized by MurE(Ct.) However, the activities for these amino acid substrates were weaker than that for meso-A(2)pm. The specificity of MurE(Ct) for three possible C. trachomatis peptidoglycan nucleotide substrates was also determined in order to deduce which amino acid might be present at the first position of the UDP-MurNAc-pentapeptide. Relative k(cat)/K(m) ratios for UDP-MurNAc-L-Ala-D-Glu, UDP-MurNAc-L-Ser-D-Glu, and UDP-MurNAc-Gly-D-Glu were 100, 115, and 27, respectively. Our results are consistent with the synthesis in chlamydiae of a UDP-MurNAc-pentapeptide in which the third amino acid is meso-A(2)pm. However, due to the lack of specificity of MurE(Ct) for nucleotide substrates in vitro, it is not obvious which amino acid is present at the first position of the pentapeptide. |
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Authors:
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Delphine Patin; Julieanne Bostock; Didier Blanot; Dominique Mengin-Lecreulx; Ian Chopra |
Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't Date: 2009-10-09 |
Journal Detail:
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Title: Journal of bacteriology Volume: 191 ISSN: 1098-5530 ISO Abbreviation: J. Bacteriol. Publication Date: 2009 Dec |
Date Detail:
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Created Date: 2009-11-25 Completed Date: 2009-12-22 Revised Date: 2010-09-28 |
Medline Journal Info:
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Nlm Unique ID: 2985120R Medline TA: J Bacteriol Country: United States |
Other Details:
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Languages: eng Pagination: 7430-5 Citation Subset: IM |
Affiliation:
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Université Paris-Sud, Institut de Biochimie et Biophysique Moléculaire et Cellulaire, CNRS, Laboratoire des Enveloppes Bactériennes et Antibiotiques, UMR 8619, Orsay F-91405, France. |
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| MeSH Terms | |
Descriptor/Qualifier:
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Adenosine Triphosphate
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metabolism Bacterial Proteins / chemistry, genetics, metabolism* Chlamydia trachomatis / enzymology* Coenzymes / pharmacology Diaminopimelic Acid / metabolism Dipeptides / metabolism Enzyme Stability Escherichia coli / genetics, metabolism Genetic Complementation Test Hydrogen-Ion Concentration Kinetics Ligases / chemistry, genetics, metabolism* Magnesium / pharmacology Substrate Specificity Temperature Uridine Diphosphate N-Acetylmuramic Acid / analogs & derivatives, metabolism |
| Chemical | |
Reg. No./Substance:
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0/Bacterial Proteins; 0/Coenzymes; 0/Dipeptides; 0/Uridine Diphosphate N-Acetylmuramic Acid; 0/uridine 5'-diphosphoryl N-acetylmuramoyl-alanyl-glutamate; 56-65-5/Adenosine Triphosphate; 583-93-7/Diaminopimelic Acid; 7439-95-4/Magnesium; EC 6.-/Ligases |
| Comments/Corrections | |
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