Structural and Functional Analyses Reveal That Staphylococcus aureus Antibiotic Resistance Factor HmrA Is a Zinc-dependent Endopeptidase

Abstract

HmrA is an antibiotic resistance factor of methicillin-resistant Staphylococcus aureus. Molecular analysis of this protein revealed that it is not a muramidase or ␤-lactamase but a nonspecific double-zinc endopeptidase consisting of a catalytic domain and an inserted oligomerization domain, which probably undergo a relative interdomain hinge rotation upon substrate binding. The active-site cleft is located at the domain interface. Four HmrA protomers assemble to a large ϳ170-kDa homotetrameric complex of 125 Å . All four active sites are fully accessible and ϳ50 -70 Å apart, far enough apart to act on a large meshwork substrate independently but simultaneously. In vivo studies with four S. aureus strains of variable resistance levels revealed that the extracellular addition of HmrA protects against loss of viability in the presence of oxacillin and that this protection depends on proteolytic activity. All of these results indicate that HmrA is a peptidase that participates in resistance mechanisms in vivo in the presence of ␤-lactams. Furthermore, our results have implications for most S. aureus strains of known genomic sequences and several other cocci and bacilli, which harbor close orthologs. This suggests that HmrA may be a new widespread antibiotic resistance factor in bacteria.

The disruption of prenylation leads to pleiotropic rearrangements in cellular behavior in Staphylococcus aureus

“…Most notably, the penicillin‐G MIC for the WT was 617 μg ml −1 , which is 12.3‐fold higher than that of the ispA mutant (50 μg ml −1 ). These sensitivities may be due in part to the down‐regulation of genes with known roles in resistance to antibiotics, including SAUSA300_2087 (a HMRA peptidase) and SAUSA300_1797 (an XRE family regulator) (Botelho et al ., ; Gebhard, ). To determine if the sensitivity to cell wall‐targeting agents was mediated by notable changes in cell ultrastructure, we next analyzed ispA mutants for physically altered cellular morphology using transmission electron microscopy (TEM).…”

Architecture and function of metallopeptidase catalytic domains

“…According to this scheme, MPs are first divided into those that have a single catalytic metal and those that have two. Dimetalate MPs mainly include exopeptidases and these are reviewed elsewhere, so here we focus on structurally characterized members of the subclass of mononuclear MPs (Fig. ).…”

Structure and computational analysis of a novel protein with metallopeptidase-like and circularly permuted winged-helix-turn-helix domains reveals a possible role in modified polysaccharide biosynthesis

“…In contrast, CA_C2195 has one bound Zn ion. In an earlier study, it was found that HmrA [PDB:3ram] [ 25 ], a Peptidase_M20 [Pfam:PF01546] protein (M20 and M28 peptidases are both in the MH clan and closely related to each other), also contained only one Zn ion and that this might have been enough to change its specificity from that of an exopeptidase (aminopeptidase or carboxypeptidase, which are the predominant specificities in both M20 and M28) to that of an endopeptidase. Despite only one Zn ion in HmrA (it is not fully clear whether the HmrA physiologically contains only one Zn ion or whether this was an artifact of the crystallization and that two Zn should be present), all five Zn-coordinating residues expected in Peptidase_M20 are conserved, which is not the case with CA_C2195.…”