Plasminogen Recruitment by Staphylococcus aureus SdrC Reveals a Tractable Antivirulence Target.

Surface-associated phosphoglycerate kinase (PGK) of Staphylococcus aureus binds human plasminogen in a partially lysine-independent manner.

Findings suggest that PGK may function as a moonlighting protein, playing an essential role in the interaction of S. aureus with its host through plasminogen binding.

Structural basis for the procoagulant activity of Staphylococcus aureus staphylocoagulase

3D models of the C-terminal SC region suggest that the major C-terminal domain of SC folds into a single-layer β-sheet structurally similar to the membrane occupation and recognition nexus (MORN) family of tandem repeats, which is critical for understanding SC-mediated fibrin generation.

SSR42 Enhances the Hemolytic Capacity of Staphylococcus aureus by Stabilizing the hla mRNA

Staphylococcus aureus utilizes a complex regulatory network to precisely control a vast array of virulence factors and facilitate infection. One such regulator, the long regulatory RNA SSR42, is emerging as a key modulator of virulence factor abundance and pathogenesis. Previous work has shown that a primary role for SSR42 is controlling hemolytic behavior through the positive regulation of α-toxin (hla/Hla). Herein, we confirm that loss of SSR42 limits hemolytic capacity due to reduced Hla production. Others have suggested that this occurs through the SaeRS two-component system; however, using epistasis experiments, we reveal that SSR42 and SaeRS function independently to control Hla activity. Using a two-plasmid system optimized to detect regulatory RNA-mediated control in Gram-positive bacteria, we demonstrate that SSR42 directly enhances Hla production through the hla 5’ untranslated region. Using interaction prediction and targeted mutagenesis, we identify a discrete site upstream of the hla ribosome-binding site that is required for SSR42 binding and hemolytic activity. Transcriptional arrest experiments further show that SSR42 stabilizes the hla mRNA, increasing its half-life and promoting downstream toxin production. Finally, electrophoretic mobility shift assays confirm a specific interaction between SSR42 and the hla 5’ UTR. Collectively, these findings establish SSR42 as a major post-transcriptional regulator of S. aureus virulence and reveal a novel RNA-mediated mechanism that promotes α-toxin production through stabilization of the hla transcript.

Exploiting classical virulence factor: staphylococcal protein a in immunotherapy and immunodiagnosis

Structural and Functional Interrogation of Active Streptococcus pneumoniae Sortase A

Sortase A (SrtA) enzymes covalently anchor surface proteins to Gram-positive bacterial cell walls, promoting colonization and virulence. In Streptococcus pneumoniae, previous studies identified both a domain-swapped dimer and an active refolded monomer, but the active enzyme has not been characterized at the structural and residue-specific level. Here, we performed quantitative proteomic comparisons of wild-type and SrtA knockout strains that confirmed the loss of multiple LPxTG-containing virulence factors, including ZmpB, NanA, and IgA1 protease, consistent with an essential role for SrtA in surface protein anchoring. To enable mechanistic studies, we established a biochemical framework to produce monomeric Streptococcus pneumoniae SrtA by refolding and developed a gel-based assay using recombinant substrates to monitor catalytic activity. The refolded monomer, but not the swapped dimer, catalyzed cleavage and transpeptidation of a canonical LPxTG substrate in a metal-independent manner under the conditions examined. We further report high-resolution NMR backbone assignments for the active monomer and identify substrate-induced chemical shift perturbations that localize to the active site. Together, these findings provide an integrated proteomic, biochemical, and NMR characterization of monomeric, catalytically active Streptococcus pneumoniae SrtA and reveal residue-specific interactions with a canonical LPNTG recognition peptide.

A secreted AIP-Like peptide from Helcococcus kunzii inhibits the Agr quorum sensing system of Staphylococcus aureus

ABSTRACT Staphylococcus aureus is a major human pathogen whose virulence is tightly regulated by the Agr quorum sensing system. In this study, we investigated the impact of Adh2, a secreted protein from the commensal bacterium Helcococcus kunzii, on S. aureus physiology and pathogenicity. Adh2 shares structural similarity with native auto-inducing peptides (AIPs), including the conserved CDFIM motif characteristic of Agr group I. We hypothesized that Adh2 interferes with Agr signaling by competitively binding the AgrC receptor. Exposure to Adh2 significantly repressed agrA and its downstream α-hemolysin hla, while upregulating spa, a gene encoding a surface adhesin. Deletion of an Adh2 region encompassing the conserved CDFIM motif abolished this regulatory effect, indicating that this region is required for Adh2 activity. RNA-Seq analysis revealed global transcriptional reprogramming, with downregulation of virulence and metabolic genes. Proteomic profiling corroborated these findings, showing reduced abundance of proteins involved in metabolic pathways (e.g. carbohydrate, lipid, and nucleotide metabolism), consistent with a shift toward a low-energy, colonization-oriented state. Importantly, Adh2 did not impair S. aureus growth across a wide concentration range (0.01–10 g/L) but significantly enhanced biofilm formation. In vivo, Adh2 administration significantly improved survival in zebrafish embryos infected with S. aureus, validating its anti-virulence potential. Together, these findings demonstrate that Adh2 suppresses Agr signaling and virulence gene expression while promoting a persistent phenotype. By shifting S. aureus toward a metabolically reduced and less pathogenic state, Adh2 emerges as a promising candidate for therapeutic modulation of bacterial behavior, particularly in the context of chronic wound infections.