Manganese Is Essential for PlcP Metallophosphoesterase Activity Involved in Lipid Remodeling in Abundant Marine Heterotrophic Bacteria

Abstract

In vast areas of the ocean, microbes must adapt to the availability of scarce nutrients, and a key strategy for reducing the cellular phosphorus (P) quota is to remodel membranes by replacing phospholipids with non-P surrogate lipids. A metallophosphoesterase, PlcP, is essential for lipid remodeling in cosmopolitan marine bacteria of the (e.g., sp. strain MED193) and SAR11 (e.g., sp. strain HTCC7211) clades, and transcription of is known to be induced by P limitation. In order to better understand PlcP-mediated lipid remodeling, we sought to characterize PlcP for its metal ion requirement and to determine its selectivity for native bacterial phospholipids. Here, we report the occurrence of a highly conserved binuclear ion center in PlcPs from MED193 and HTCC7211 and show that manganese is the preferred metal for metallophosphoesterase activity. PlcP displayed high activity towards the major bacterial phospholipids, e.g., phosphatidylglycerol but also phosphatidic acid, a key intermediate in phospholipid biosynthesis. In contrast, phosphatidylserine and phosphatidylinositol, both of which are rare lipids in bacteria, are not preferred substrates. These data suggest that PlcP undertakes a generic lipid remodeling role during the cellular response of marine bacteria to P deficiency and that manganese availability may play a key role in regulating the lipid remodeling process. Membrane lipids form the structural basis of all cells. In the marine environment, it is well established that phosphorus availability significantly affects lipid composition in cosmopolitan marine bacteria, whereby non-phosphorus-containing lipids are used to replace phospholipids in response to phosphorus stress. Central to this lipid remodeling pathway is a newly identified phospholipase C-type metallophosphoesterase (PlcP). However, little is known about how PlcP activity is regulated. Here, we determined the role of metal ions in regulating PlcP activity and compared PlcP substrate specificities in PlcP enzymes from two model marine bacteria from the marine clade and the SAR11 clade. Our data provide new insights into the regulation of lipid remodeling in these marine bacteria.

Insights into the Mn2+ Binding Site in the Agmatinase-Like Protein (ALP): A Critical Enzyme for the Regulation of Agmatine Levels in Mammals

“…These results agree with our previous observations where wild-type-ALP increased its activity~4-fold [29]. The increased activity of ALP, produced for the heating at 65 • C with Mn 2+ , is a typical characteristic of ureohydrolases, such as the rat liver arginase [18,39] and Helicobacter pylori ARG [32]. It has been suggested that the heating of the enzyme in the presence of Mn 2+ increases the activity of the enzyme through the stabilization of its binuclear Mn 2+ center [32].…”

“…While aspartate is often found stabilizing binuclear metal centers, residues such as glutamate and asparagine can also play this role [35]. For example, it has been reported that Asn81 stabilizes the binuclear Mn 2+ center of metallophosphoesterase from a marine bacteria [39], and Asn233 plays a similar role in the binuclear Zn 2+ center of the betalactamase of Bacillus cereus [40]. Furthermore, glutamic residues (Glu235 and Glu204) have been described as stabilizing the binuclear Co 2+ center of a methionine aminopeptidase from E. coli [35] and the binuclear Mn 2+ center (Glu 56-57-58) of a pyrophosphohydrolase from E. coli [41].…”

Phosphorus stress induces the synthesis of novel glycolipids in Pseudomonas aeruginosa that confer protection against a last-resort antibiotic

“…MED193 and 46% identity to PlcP from Sinorhizobium meliloti [26][27][28]. In these bacteria, PlcP is essential in the lipid remodelling pathway for the formation of the diacylglycerol (DAG) backbone, representing the essential intermediate for the production of glycolipids [32,33]. In P. aeruginosa PAO1, PA3219 appears to form an operon with PA3218, a putative glycosyltransferase likely under the control of the PhoBR two-component system, as a highly conserved Pho box sequence was recognisable in the promoter region (Fig.…”

Characterization of a glycolipid glycosyltransferase with broad substrate specificity from the marine bacteriumCandidatusPelagibacter sp. HTCC7211

“…However, little is known about how these glycoglycerolipids are synthesized. Previous studies have shown that a manganese-dependent metallophosphoesterase, PlcP, is essential for lipid remodelling in marine heterotrophs, and that the plcP gene is organized in an operon-like structure and a putative glycosyltransferase was found down stream of plcP in numerous marine heterotrophic bacteria, such as members of the SAR11 clade (18, 21). Our previous work has shown that the GT from the marine bacterium SAR11 ( Candidatus Pelagibacter sp.…”