Abstract
ABSTRACTMycobacterium tuberculosis, the etiological agent of tuberculosis (TB), has a unique cell envelope which accounts for its unusual low permeability and contributes to resistance against common antibiotics. The main structural elements of the cell wall consist of a cross-linked network of peptidoglycan (PG) in which some of the muramic acid residues are covalently attached to a complex polysaccharide, arabinogalactan (AG), via a unique α-l-rhamnopyranose–(1→3)-α-d-GlcNAc-(1→P) linker unit. While the molecular genetics associated with PG and AG biosynthetic pathways have been largely delineated, the mechanism by which these two major pathways converge has remained elusive. In Gram-positive organisms, the LytR-CpsA-Psr (LCP) family of proteins are responsible for ligating cell wall teichoic acids to peptidoglycan, through a linker unit that bears a striking resemblance to that found in mycobacterial arabinogalactan. In this study, we have identified Rv3267 as a mycobacterial LCP homolog gene that encodes a phosphotransferase which we have named Lcp1. We demonstrate that lcp1 is an essential gene required for cell viability and show that recombinant Lcp1 is capable of ligating AG to PG in a cell-free radiolabeling assay.
Highlights
Mycobacterium tuberculosis, the etiological agent of tuberculosis (TB), has a unique cell envelope which accounts for its unusual low permeability and contributes to resistance against common antibiotics
Unlike archetypal PG commonly found in bacteria such as Escherichia coli, the muramic acid residues in Mycobacterium tuberculosis contains a mixture of N-acetyl and N-glycolyl derivatives, whereby the N-acetyl function has been oxidized to an N-glycolyl function to form MurNGly [4,5,6]
We identified three putative LCP homologs in M. tuberculosis, with Rv3267 sharing 20% amino acid sequence identity with Cps2A from S. pneumoniae, which is in accordance with a previous phylogenetic study of bacterial LCP proteins [16]
Summary
Mycobacterium tuberculosis, the etiological agent of tuberculosis (TB), has a unique cell envelope which accounts for its unusual low permeability and contributes to resistance against common antibiotics. The main structural elements of the cell wall consist of a cross-linked network of peptidoglycan (PG) in which some of the muramic acid residues are covalently attached to a complex polysaccharide, arabinogalactan (AG), via a unique ␣-L-rhamnopyranose–(1¡3)-␣-D-GlcNAc-(1¡P) linker unit. In Gram-positive organisms, the LytR-CpsA-Psr (LCP) family of proteins are responsible for ligating cell wall teichoic acids to peptidoglycan, through a linker unit that bears a striking resemblance to that found in mycobacterial arabinogalactan. PG and AG form a huge macropolymer positioned between the cytoplasmic membrane and the outer mycolic acid layer of the TB bacillus Our understanding of both the structure and biosynthesis of mycobacterial AG has developed steadily over the last 2 decades, with much emphasis being placed on investigating the molecular genetics of how this complex structure is assembled (extensively reviewed in reference 9). We demonstrate that the open reading frame contained in Rv3267, one of three LCP ortholog genes in M. tuberculosis, encodes a peptidoglycan– arabinogalactan ligase, which we have termed Lcp
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