Mycobacterium tuberculosis Rv1302 and Mycobacterium smegmatis MSMEG_4947 have WecA function and MSMEG_4947 is required for the growth of M. smegmatis.
The disaccharide d-N-acetylglucosamine-l-rhamnose plays an important role in the mycobacterial cell wall as a linker connecting arabinogalactan and peptidoglycan via a phosphodiester linkage. The first step of the disaccharide linker is the formation of decaprenyl phosphate-GlcNAc, which is catalyzed by GlcNAc-1-phosphate transferase. In Gram-negative bacteria, the wecA gene specifies the UDP-GlcNAc: undecaprenyl phosphate GlcNAc-1-phosphate transferase (WecA), which catalyzes the first step in the biosynthesis of lipopolysaccharide O-antigen. Mycobacterium tuberculosis Rv1302 and Mycobacterium smegmatis MSMEG_4947 show homology to Escherichia coli WecA protein. We cloned Rv1302 and MSMEG_4947 and introduced plasmids pYJ-1 (carrying Rv1302) and pYJ-2 (carrying MSMEG_4947) into a wecA-defective strain of E. coli MV501, respectively. Lipopolysaccharide analysis demonstrated that lipopolysaccharide synthesis in MV501 (pYJ-1) and MV501 (pYJ-2) was restored upon complementation with Rv1302 and MSMEG_4947, respectively. This provides the first evidence that Rv1302 and MSMEG_4947 have the same function as E. coli WecA. We also generated an M. smegmatis MSMEG_4947 knockout mutant using a homologous recombination strategy. The disruption of MSMEG_4947 in the M. smegmatis genome resulted in the loss of viability at a nonpermissive temperature. Scanning electron microscopy and transmission electron microscopy results showed that the lack of the MSMEG_4947 protein causes drastic morphological changes in M. smegmatis.
Jin
Yue
Y
Department of Biochemistry and Molecular Biology, Dalian Medical University, 9 W. Lushun South Road, Dalian, China.
Xin
Yi
Y
Zhang
Wenli
W
Ma
Yufang
Y
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D017353:Gene Deletion; D020043:Genes, Essential; D005816:Genetic Complementation Test; D050296:Microbial Viability; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D020102:Mycobacterium smegmatis / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000648:ultrastructure; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics; D010957:Plasmids; D017386:Sequence Homology, Amino Acid; D017855:Transferases (Other Substituted Phosphate Groups) / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D017855:Transferases (Other Substituted Phosphate Groups); C083617:wecA protein, E coli