Deciphering the metabolism of undecaprenyl-phosphate: the bacterial cell-wall unit carrier at the membrane frontier

Abstract

During the biogenesis of bacterial cell-wall polysaccharides, such as peptidoglycan, cytoplasmic synthesized precursors should be trafficked across the plasma membrane. This essential process requires a dedicated lipid, undecaprenyl-phosphate that is used as a glycan lipid carrier. The sugar is linked to the lipid carrier at the inner face of the membrane and is translocated toward the periplasm, where the glycan moiety is transferred to the growing polymer. Undecaprenyl-phosphate originates from the dephosphorylation of its precursor undecaprenyl-diphosphate, with itself generated by de novo synthesis or by recycling after the final glycan transfer. Undecaprenyl-diphosphate is de novo synthesized by the cytosolic cis-prenyltransferase undecaprenyl-diphosphate synthase, which has been structurally and mechanistically characterized in great detail highlighting the condensation process. In contrast, the next step toward the formation of the lipid carrier, the dephosphorylation step, which has been overlooked for many years, has only started revealing surprising features. In contrast to the previous step, two unrelated families of integral membrane proteins exhibit undecaprenyl-diphosphate phosphatase activity: BacA and members of the phosphatidic acid phosphatase type 2 super-family, raising the question of the significance of this multiplicity. Moreover, these enzymes establish an unexpected link between the synthesis of bacterial cell-wall polymers and other biological processes. In the present review, the current knowledge in the field of the bacterial lipid carrier, its mechanism of action, biogenesis, recycling, regulation, and future perspective works are presented.

FIG. 1.

Structures of different polyprenyl-phosphate carrier lipids (A) and lipid intermediates from various glycoconjugate biosynthetic pathways (B).

FIG. 2.

C₅₅-P metabolism and membrane steps of peptidoglycan biosynthesis. C₅₅-PP is de novo synthesized by undecaprenyl-diphosphate synthase (UppS) enzyme in the cytoplasm, from which it partitions in the inner side of the plasma membrane. Subsequently, C₅₅-PP should be dephosphorylated to function as a lipid carrier in various polysaccharide biosynthetic pathways, such as the peptidoglycan synthesis. The enzymes MraY and MurG catalyze the successive transfers of the phospho-MurNAc-pentapeptide and GlcNAc motifs from the nucleotide precursors to the lipid carrier C₅₅-P, generating the lipid I and lipid II intermediates, respectively. Lipid II is then translocated by the flippase FtsW to the periplasmic side of the membrane, where polymerization reactions of peptidoglycan that are catalyzed by the penicillin-binding proteins (PBPs) occur. The lipid carrier is released in its pyrophosphate form (C₅₅-PP), which should be dephosphorylated and shuttled back to the inner face of the membrane to be reused. C₅₅-P metabolism is the target of bacitracin that sequestrates C₅₅-PP, thereby inhibiting its dephosphorylation, and of colicin M which cleaves lipid II in dead-end products, C₅₅-OH and 1-pyrophospho-MurNAc(-peptide)-GlcNAc.

FIG. 3.

Mechanisms of C₅-PP condensation on an allylic substrate by cis-prenyltransferases (A) and trans-prenyltransferases (B).

FIG. 4.

Structures of UppS enzyme from Escherichia coli. Overall structure of UppS (1UEH atomic coordinate): one dimer shown with purple α-helices and blue β-strands and the other monomer shown with red α-helices and green β-strands (A); structure of UppS in complex with C₁₅-PP homoallylic substrate (represented with spheres) (1V7U atomic coordinate), highlighting conformational changes occurring in the α3-helix on substrate binding (B). Details of the ternary complex of UppS with Mg²⁺, C₁₅-sPP, and C₅-PP of wild-type enzyme (left) and a D26A mutant (right) (1X06, 1X07, 1X08, and 1X09 atomic coordinates) emphasizing the condensation mechanism and the role played by the aspartate 26 residue in this process (C).

FIG. 5.

Phosphorylation of lipid A by LpxT in E. coli, Salmonella enterica, and Salmonella typhimurium. LpxT transfers the distal phosphate group from C₅₅-PP to the 1-phosphate group of nascent lipid A at the periplasmic side of the plasma membrane, releasing lipid A-1-diphosphate and C₅₅-P. The 6′ position of lipid A is linked to two Kdo (3-deoxy-D-manno-octulosonic acid) residues from the core region. Subsequently, the core-lipid A (followed or not by the outermost O-antigen depending on the bacterial species) is exported toward the outer membrane and forms the outer leaflet. In rich media, the modification, whose role is yet to be discovered, occurs in about one third of total lipid A molecules. The activation of the PmrA/PmrB two-component system, by high extracellular concentrations of Fe³⁺, induces the expression of a small-membrane peptide, PmrR, which inhibits LpxT; along with ArnT and EptA, which catalyze the modification of lipid A with 4-amino-4-deoxy-L-arabinose (to the 4′-phosphate group) and phosphoethanolamine (to the 1-phosphate group), respectively. The latter lipid A decorations confer increased resistance to Fe³⁺ and cationic antibacterial peptides (CAMPs) and attenuate the recognition of lipid A by the host immune TLR4 system.