Characterization of the Vibrio cholerae VolA Surface-Exposed Lipoprotein Lysophospholipase

Abstract

bBacterial lipases play important roles in bacterial metabolism and environmental response. Our laboratory recently discovered that a novel lipoprotein lysophospholipase, VolA, localizes on the surface of the Gram-negative aquatic pathogen Vibrio cholerae. VolA functions to cleave exogenous lysophosphatidylcholine, freeing the fatty acid moiety for use by V. cholerae. This fatty acid is transported into the cell and can be used as a nutrient and, more importantly, as a way to alter the membrane architecture via incorporation into the phospholipid biosynthesis pathway. There are few examples of Gram-negative, surface-exposed lipoproteins, and VolA is unique, as it has a previously undercharacterized function in V. cholerae membrane remodeling. Herein, we report the biochemical characterization of VolA. We show that VolA is a canonical lipoprotein via mass spectrometry analysis and demonstrate the in vitro activity of VolA under a variety of conditions. Additionally, we show that VolA contains a conserved Gly-Xaa-Ser-Xaa-Gly motif typical of lipases. Interestingly, we report the observation of VolA homologs in other aquatic pathogens. An Aeromonas hydrophila VolA homolog complements a V. cholerae VolA mutant in growth on lysophosphatidylcholine as the sole carbon source and in enzymatic assays. These results support the idea that the lipase activity of surface-exposed VolA likely contributes to the success of V. cholerae, improving the overall adaptation and survival of the organism in different environments. R ecent work from our laboratory revealed the presence of a unique membrane-anchored lipase, VolA, localized on the surface of Vibrio cholerae cells (1). We demonstrated that VolA is required for growth when lysophosphatidylcholine (LPC) (Fig. 1) serves as the sole carbon source. We hypothesized that VolA could cleave exogenous lysophosphatidylcholine into long-chain fatty acid (LCFA) derivatives (Fig. 1), allowing them to be brought into the cell via a coexpressed fatty acid transporter, FadL. These data, taken together with the presence of a conserved lipase domain identified in V. cholerae (2) and the inability of wild-type V. cholerae to grow on phosphatidylcholine as a sole carbon source, suggest that VolA acts as a lysophospholipase in vivo.Phospholipases are members of the acylhydrolase family of enzymes, acting on ester bonds in phospholipid targets (3). The structure and function of such acylhydrolases are conserved and have been well studied. All contain a characteristic fold (4, 5), sharing a conserved lipase motif (2). In our previous work (1), we noted that VolA contains this conserved motif, which strongly implicates it as a lipase. Phospholipases vary in terms of their site of action on the phospholipid. Some target phosphate bonds, while others hydrolyze bonds between the glycerol moiety and the acyl chains. Most microbial lysophospholipases hydrolyze the ester bond between the acyl chain and the polar head group, releasing a free fatty acid (6). It is likely that VolA functions similarly to e...

Within-species variation in OMV cargo proteins: theMyxococcus xanthusOMV pan-proteome

“…VolA has been found in M. xanthus vesicles, 36,37 and is present on the surface of Vibrio cholerae cells, where the primary function is liberating fatty acids for consumption, a possible function that could also occur on the surface of vesicles. 61,62 In Pseudomonas aeruginosa, PvdQ has been linked to quorum quenching and iron homeostasis, 63 which could be useful for myxobacteria predation, as they are known to sense and respond to prey quorum signals. 64 GspD, a key member of the general secretion pathway, is typically found on the outer membrane.…”

BB0562 is a nutritional virulence determinant with lipase activity important for Borrelia burgdorferi infection and survival in fatty acid deficient environments

“…Indeed, purified, recombinant BB0562 demonstrated lipase activity, which was dependent on the serine residues in the two canonical GXSXG lipase motifs identified in the protein. In bacteria, lipases have been predominantly shown to be secreted extracellularly [ 40 , 42 ] and there are some examples of lipases that localize to the bacterial outer surface [ 63 , 64 ], both of which likely allow interaction of the lipase with target substrates in the environment. Our data indicate that BB0562 is associated with the spirochete outer membrane but is likely not surface exposed.…”

Outer membrane lipoprotein biogenesis: Lol is not the end

“…Many surface-exposed lipoproteins have been identified in a number of different organisms [36]. In those organisms, surface-exposed lipoproteins have variety of functions: they participate in iron uptake [37][38][39][40]; are enzymes such as phospholipases [41], PPIases [42] or glucanases [43]; they participate in adhesion and binding of host factors [44,45]; and others. In some organisms, like Borrelia, lipoproteins are anchored in the outer leaflet and exposed on the cell surface by default [24].…”