Biochemical characterisation of the chlamydial MurF ligase, and possible sequence of the chlamydial peptidoglycan pentapeptide stem.
Chlamydiaceae are obligate intracellular bacteria that do not synthesise detectable peptidoglycan although they possess an almost complete arsenal of genes encoding peptidoglycan biosynthetic activities. In this paper, the murF gene from Chlamydia trachomatis was shown to be capable of complementing a conditional Escherichia coli mutant impaired in UDP-MurNAc-tripeptide:D-Ala-D-Ala ligase activity. Recombinant MurF from C. trachomatis was overproduced and purified from E. coli. It exhibited ATP-dependent UDP-MurNAc-X-γ-D-Glu-meso-A(2)pm:D-Ala-D-Ala ligase activity in vitro. No significant difference of kinetic parameters was seen when X was L-Ala, L-Ser or Gly. The L-Lys-containing UDP-MurNAc-tripeptide was a poorer substrate as compared to the meso-A(2)pm-containing one. Based on the respective substrate specificities of the chlamydial MurC, MurE, MurF and Ddl enzymes, a sequence L-Ala/L-Ser/Gly-γ-D-Glu-meso-A(2)pm-D-Ala-D-Ala is expected for the chlamydial pentapeptide stem, with Gly at position 1 being less likely.
Laboratoire des Enveloppes Bactériennes et Antibiotiques, Centre National de la Recherche Scientifique, Institut de Biochimie et Biophysique Moléculaire et Cellulaire, UMR 8619, Bâtiment 430, Université Paris-Sud, 91405 Orsay Cedex, France.
Delphine
D
Patin
Julieanne
J
Bostock
Ian
I
Chopra
Dominique
D
Mengin-Lecreulx
Didier
D
Blanot
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002692:Chlamydia trachomatis / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D004151:Dipeptides / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005816:Genetic Complementation Test; D008025:Ligases / Q000235:genetics / Q000378:metabolism*; D010457:Peptidoglycan / Q000737:chemistry; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D014169:Transformation, Bacterial
D001426:Bacterial Proteins; D004151:Dipeptides; D010457:Peptidoglycan; D011994:Recombinant Proteins; C002956:alanylalanine; D008025:Ligases