Discovery of a bifunctional cardiolipin/phosphatidylethanolamine synthase in bacteria

Abstract

SummaryPhosphatidylethanolamine (PE) and cardiolipin (CL) are major components of bacterial and eukaryotic membranes. In bacteria, synthesis of PE usually occurs via decarboxylation of phosphatidylserine (PS) by PS decarboxylases (Psd). CL is produced by various CL synthases (Cls). Membranes of the plant pathogen Xanthomonas campestris predominantly contain PE, phosphatidylglycerol (PG) and CL. The X. campestris genome encodes one Psd and six putative CLs. Deletion of psd resulted in loss of PE and accumulation of PS. The mutant was severely affected in growth and cell size. PE synthesis, growth and cell division were partially restored when cells were supplied with ethanolamine (EA) suggesting a previously unknown PE synthase activity. Via mutagenesis, we identified a Cls enzyme (Xc_0186) responsible for EA-dependent PE biosynthesis. Xanthomonas lacking xc_0186 not only lost its ability to utilize EA for PE synthesis but also produced less CL suggesting a bifunctional enzyme. Recombinant Xc_0186 in E. coli and in cell-free extracts uses cytidine diphosphate diacylglycerol (CDP-DAG) and PG for CL synthesis. It is also able to use CDP-DAG and EA for PE synthesis. Owing to its dual function in CL and PE production, we consider Xc_0186 the founding member of a new class of enzymes called CL/PE synthase (CL/PEs).

Tat transport in Escherichia coli requires zwitterionic phosphatidylethanolamine but no specific negatively charged phospholipid

“…However, as pssA and pgsA cannot be deleted in the same strain, it is not easy to address this question by molecular biology tools. Our data are also in agreement with the recently discovered contribution of ClsB to PG synthesis , as no PG accumulates in the pgsA / clsA / clsB / clsC quadruple mutant relative to the pgsA single mutant strain (Fig. E).…”

Unconventional membrane lipid biosynthesis in Xanthomonas campestris

“…The relative content of CL in bacteria depends on the growth phase and culture conditions and can vary from a few to 30%. As in most bacteria, CL is a predominant lipid in early stationary phase in X. campestris (Moser et al ., 2014a,b). Many bacteria including E. coli , Staphylococcus aureus and Bacillus subtilis contain multiple CL synthases (Lopez et al ., ; Tan et al ., ; Kuhn et al ., ).…”

“…The most important factors contributing to virulence are the Type3 secretion system and effector proteins delivered by this system such as the transcription activator‐like effectors (TALEs) unique to Xanthomonads (Boch et al ., ; Boch and Bonas, ; Büttner and Bonas, ). Recent research on membrane lipid biosynthesis in X. campestris suggests that Xanthomonads are not only unique in pathogenicity concepts but also have established non‐conventional pathways for membrane lipid synthesis (Moser et al ., 2014a,b).…”

“…Xanthonomas campestris membranes contain the typical bacterial phospholipids PE (∼50% of the total PL composition), CL (∼33%) and PG (∼13%) and small amounts of MMPE (∼4%). When grown in complex medium, Xanthomonas is able to produce small amounts of PC (∼6%) (Moser et al ., 2014a,b). Xanthonomas campestris contains the complete gene repertoire required for conventional biosynthesis of the common phospholipid precursor CDP‐DAG and the major PLs PE, PG and CL (Fig.…”

“…). Apparently, the remaining putative Cls enzymes cannot compensate these defects (Moser et al ., ). Heterologous expression of the Xanthomonas CL/PEs gene in E. coli resulted in CL accumulation and EA‐dependent PE production providing conclusive evidence for the bifunctional activity.…”