Two cutinase‐like proteins secreted byMycobacterium tuberculosisshow very different lipolytic activities reflecting their physiological function

Abstract

Cutinases are extracellular enzymes that are able to degrade cutin, a polyester protecting plant leaves and many kinds of lipids. Although cutinases are mainly found in phytopathogenic fungi or bacteria, 7 genes related to the cutinase family have been predicted in the genome of Mycobacterium tuberculosis. These genes may encode proteins that are involved in the complex lipid metabolism of the bacterium. Here, we report on the biochemical characterization of two secreted proteins of M. tuberculosis, Rv1984c and Rv3452, belonging to the cutinase family. Although their amino acid sequence shows 50% identity with that of the well-characterized cutinase from Fusarium solani pisi, and a high level of homology has been found to exist between these two enzymes, they show distinct substrate specificities. Rv1984c preferentially hydrolyzes medium-chain carboxylic esters and monoacylglycerols, whereas Rv3452 behaves like a phospholipase A(2), and it is able to induce macrophage lysis. The tetrahydrolipstatin inhibitor, a specific lipase inhibitor, abolishes the activity of both enzymes. Site-directed mutagenesis was performed to identify the catalytic triad of Rv1984c. Structural models for Rv1984c and Rv3452 were built, based on the crystal structure of F. solani cutinase, with a view to investigating the contribution of specific residues to the substrate specificity. Our findings open new prospects for investigating the physiological roles of cutinase-like proteins in the lipid metabolism and virulence of M. tuberculosis.

Enzymatic Hydrolysis of Trehalose Dimycolate Releases Free Mycolic Acids during Mycobacterial Growth in Biofilms

“…2D). Seven cutinase-like proteins, with esterase activities in some of them, have been described in M. tuberculosis (42,51), and we suggest that one or more of these behaves similarly to Msmeg_1529, although none has yet been shown to cleave either TDM or mAGP. Rv3452, designated Culp4, is the closest relative of Msmeg_1529, has a putative signal peptide, and has been reported to be cell wallassociated (52).…”

Systematic Survey of Clonal Complexity in Tuberculosis at a Populational Level and Detailed Characterization of the Isolates Involved

“…In one case, two variants differed in an IS6110 sequence located in or absent from Rv1758. An IS6110 copy in the same sequence has also been found in strain H37Rv, and it has been reported that it might have a dual impact: on the expression of both cutinase, an enzyme involved in the lipid metabolism (27), and phospholipase plcD (16), an enzyme considered a virulence factor in certain studies (24). For the second case infected with clonal variants who was analyzed, we detected an IS6110 copy either present or absent in a region coding for the protein PPE34, which has recently been found to facilitate a shift toward the Th2 immune response, aiding the immune evasion by mycobacteria (7).…”

Discovery of a glycerol 3-phosphate phosphatase reveals glycerophospholipid polar head recycling in Mycobacterium tuberculosis

“…Such phospholipase activities have been identified in M. tuberculosis. Schué et al (28) isolated two secreted cutinases-like proteins: Rv1984c, which preferentially hydrolyzed medium-chain carboxylic esters and monoacylglycerols, and Rv3452, which behaved like a phospholipase A2. The other lipases/cutinases are most likely cell-wall-associated and surface-exposed (29).…”