Open AccessMycobacterium tuberculosis Rv1302 and Mycobacterium smegmatis MSMEG___4947 have WecA function and MSMEG__4947 is required for the growth of M. smegmatis
Author(s) -
Jin Yue,
Xin Yi,
Zhang Wenli,
Ma Yufang
Publication year - 2010
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2010.02045.x
Subject(s) - mycobacterium smegmatis , peptidoglycan , escherichia coli , transferase , biology , shuttle vector , biochemistry , cell wall , mycobacterium tuberculosis , chemistry , recombinant dna , enzyme , gene , medicine , tuberculosis , pathology , vector (molecular biology)
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‐Glc N Ac, which is catalyzed by Glc N Ac‐1‐phosphate transferase. In Gram‐negative bacteria, the wecA gene specifies the UDP‐Glc N Ac: undecaprenyl phosphate Glc N Ac‐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 .