Structural analyses of the Haemophilus influenzae peptidoglycan synthase activator LpoA suggest multiple conformations in solution [Microbiology]

In many Gram-negative bacteria, the peptidoglycan synthase PBP1A requires the outer membrane lipoprotein LpoA for constructing a functional peptidoglycan required for bacterial viability. Previously, we have shown that the C-terminal domain of Haemophilus influenzae LpoA (HiLpoA) has a highly conserved, putative substrate-binding cleft between two α/β lobes. Here, we report a 2.0-Å-resolution crystal structure of the HiLpoA N-terminal domain. Two subdomains contain tetratricopeptide-like motifs that form a concave groove, but their relative orientation differs by ~45° from that observed in an NMR structure of the Escherichia coli LpoA N domain. We also determined three 2.0-2.8-Å-resolution crystal structures containing four independent full-length HiLpoA molecules. In contrast to an elongated model previously suggested for E. coli LpoA, each HiLpoA formed a U-shaped structure with a different C-domain orientation. This resulted from both N-domain twisting and rotation of the C domain (up to 30°) at the end of the relatively immobile interdomain linker. Moreover, a previously predicted hinge between the lobes of the LpoA C domain exhibited variations of up to 12°. Small-angle X-ray scattering (SAXS) data revealed excellent agreement with a model calculated by normal mode analysis (NMA) from one of the full-length HiLpoA molecules, but even better agreement with an ensemble of this molecule and two of the partially extended NMA-predicted models. The different LpoA structures helped explain how an outer membrane−anchored LpoA can either withdraw from or extend toward the inner membrane−bound PBP1A through peptidoglycan gaps and hence regulate the synthesis of peptidoglycan necessary for bacterial viability.

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