Abstract
The biogenesis of bacterial cell-envelope polysaccharides requires the translocation, across the plasma membrane, of sugar sub-units that are produced inside the cytoplasm. To this end, the hydrophilic sugars are anchored to a lipid phosphate carrier (undecaprenyl phosphate (C55-P)), yielding membrane intermediates which are translocated to the outer face of the membrane. Finally, the glycan moiety is transferred to a nascent acceptor polymer, releasing the carrier in the “inactive” undecaprenyl pyrophosphate (C55-PP) form. Thus, C55-P is generated through the dephosphorylation of C55-PP, itself arising from either de novo synthesis or recycling. Two types of integral membrane C55-PP phosphatases were described: BacA enzymes and a sub-group of PAP2 enzymes (type 2 phosphatidic acid phosphatases). The human pathogen Helicobacter pylori does not contain BacA homologue but has four membrane PAP2 proteins: LpxE, LpxF, HP0350 and HP0851. Here, we report the physiological role of HP0851, renamed HupA, via multiple and complementary approaches ranging from a detailed biochemical characterization to the assessment of its effect on cell envelope metabolism and microbe-host interactions. HupA displays a dual function as being the main C55-PP pyrophosphatase (UppP) and phosphatidylglycerol phosphate phosphatase (PGPase). Although not essential in vitro, HupA was essential in vivo for stomach colonization. In vitro, the remaining UppP activity was carried out by LpxE in addition to its lipid A 1-phosphate phosphatase activity. Both HupA and LpxE have crucial roles in the biosynthesis of several cell wall polysaccharides and thus constitute potential targets for new therapeutic strategies.
Highlights
The biogenesis of many bacterial cell-envelope polysaccharides (i.e., peptidoglycan (PGN), lipopolysaccharides (LPS), teichoic acids, enterobacterial common antigen) requires the translocation, across the cytoplasmic membrane, of glycan units that are produced inside the cytoplasm [1]
We show that HP0851, renamed HupA, is the major undecaprenyl pyrophosphate (C55-PP) phosphatase (UppP) and the major phosphatidylglycerol phosphate phosphatase (PGPase)
This enzyme is involved in cationic antimicrobial peptide (CAMP) resistance to which H. pylori hupA mutant shows an increased sensitivity (4 fold)
Summary
The biogenesis of many bacterial cell-envelope polysaccharides (i.e., peptidoglycan (PGN), lipopolysaccharides (LPS), teichoic acids, enterobacterial common antigen) requires the translocation, across the cytoplasmic membrane, of glycan units that are produced inside the cytoplasm [1]. In PGN biosynthesis, these intermediates are cross-linked by transglycosylase and transpeptidase activities to a nascent acceptor polymer. These polymerization reactions release the lipid carrier in an “inactive” undecaprenyl pyrophosphate form (C55-PP) which must be recycled to participate in new rounds of cell-envelope polysaccharides biosynthesis. Overexpression of BacA, PgpB or YbjG resulted in bacitracin resistance, and an increase of the UppP activity contained in membrane extracts [7]. Bacitracin is an antibiotic produced by Bacillus licheniformis [8] which strongly binds C55-PP, thereby inhibiting its dephosphorylation and leading to an arrest of PGN biosynthesis. LpxT was not able to sustain growth of the triple bacA-ybjG-pgpB mutant and its overexpression did not lead to any bacitracin resistance suggesting that LpxT displays another function. In addition to its UppP activity, PgpB is involved in phospholipids
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