Dissecting the membrane lipid binding properties and lipase activity of Mycobacterium tuberculosis LipY domains

Abstract

The Mycobacterium tuberculosis LipY protein, a prototype of the proline‐glutamic acid (PE) family, exhibits a triacylglycerol (TAG) hydrolase activity that contributes to host cell lipid degradation and persistence of the bacilli. LipY is found either as a full‐length intracytosolic form or as a mature extracellular form lacking the N‐terminal PE domain. Even though the contribution of the extracellular form in TAG consumption has been partly elucidated, very little information is available regarding the potential interactions of either full‐length LipY with the cytoplasmic membrane, or mature form LipY with the outer membrane. Herein, several LipY variants truncated in their N‐terminal domain were produced and biochemically characterized in lipid–protein interaction assays, using the monomolecular film technique and FTIR. Comparison of the catalytic activities of these recombinant proteins showed that LipY∆149, corresponding to the extracellular form of LipY lacking the PE domain, is more active than the full‐length protein. This confirms previous studies reporting that the PE domain negatively modulates the TAG hydrolase activity of LipY. Lipid–protein interaction studies indicate that the PE domain anchors LipY onto membrane lipids. Consistent with these findings, we show that LipY∆149 is loosely associated with the mycobacterial cell wall, and that this interaction is mediated by the sole lipase domain. Overall, our results bring new information regarding the molecular mechanisms by which LipY either binds and hydrolyses host cell lipids or degrades TAG, the major source of lipids within mycobacterial intracytosolic lipid inclusions.

Lipolytic enzymes inhibitors: A new way for antibacterial drugs discovery

“…Few years later, we reported that a new substitutive OX derivative, the MmPPOX (Figure 1), was also able to inhibit M. tb growth on solid medium with moderated MIC values of 50-90 µM [28]. We further showed that MmPPOX efficiently inhibited pure recombinant M. tb enzymes belonging to the hormone-sensitive lipase (HSL) family member proteins (i.e., Lip-HSL) [28], including LipY (Rv3097c) the major M. tb Lip-HSL lipase involved in TAG acquisition from the host and in ILI breakdown [2,10,17,41]. The mechanism of action of MmPPOX involving the formation of a covalent bond with the catalytic serine residue of the enzymes (Figure 1 inset) thus resulting in a total abolition of their activities was also confirmed [28].…”

The endogenous galactofuranosidase GlfH1 hydrolyzes mycobacterial arabinogalactan

“…The importance of GlfH1 in cell wall remodeling is further emphasized by its genomic environment, as it is adjacent to Rv3097c, also known as lipY, that encodes a wellcharacterized PE protein expressing triacylglycerol (TAG) hydrolase activity. LipY participates to consumption and reprocessing of both mycobacterial lipids and host lipids, thereby contributing to persistence of the tubercle bacillus inside foamy macrophages (42)(43)(44).…”

PE_PGRS: Vital proteins in promoting mycobacterial survival and modulating host immunity and metabolism

“…LipY was the first PE/PPE protein found to have enzymatic properties, namely triacylglycerol (TAG) lipase activity (Mishra et al, 2008), allowing it to cleave TAG into a diacylglycerol. PE_PGRS35, which contains an aspartic proteinase domain, was shown to be able to process LipY, resulting in the cleavage of the PE domain of LipY, thus may serve as an activation trigger for LipY (Burggraaf et al, 2019; Daleke et al, 2011; Santucci et al, 2019). PE_PGRS10 and PE_PGRS21 are also predicted to be involved in lipid metabolism (Mazandu & Mulder, 2012, Figure 1, Table 1 ) .…”