The Multifunctional Staphylococcus aureus Autolysin Aaa Mediates Adherence to Immobilized Fibrinogen and Fibronectin

Abstract

Staphylococci can cause a wide spectrum of infections, including endocarditis, osteomyelitis, and sepsis, which is reflected by the numerous virulence factors they produce, among them a recently identified new class of adhesins, namely, the multifunctional autolysins/adhesins. Here we report the identification and molecular characterization of Aaa, a novel autolysin/adhesin from Staphylococcus aureus. The gene encoding Aaa was cloned from the clinical isolate Staphylococcus aureus 4074. DNA sequence analysis revealed that aaa encodes a deduced protein of 334 amino acids with a predicted molecular mass of 35.8 kDa. Aaa contains three N-terminal repetitive sequences that comprise features of a peptidoglycan-binding domain, the LysM domain. The expression of aaa by Escherichia coli and its subsequent characterization revealed that Aaa possesses bacteriolytic activity as well as adhesive properties, such as binding to extracellular matrix proteins. Real-time biomolecular interaction analysis demonstrated that the interaction of Aaa with fibrinogen, fibronectin, and vitronectin is dose dependent and saturable and occurs with a high affinity. Furthermore, we demonstrate that Aaa binds to the A␣ and B␤ chains of fragment D of fibrinogen. Immunofluorescence microscopy revealed that Aaa is located at the cell surface. Finally, an aaa knockout mutant showed reduced adherence to surfaceadsorbed fibrinogen and fibronectin, strongly suggesting a role for Aaa in the colonization of host factor-coated polymer surfaces and/or host tissue.

Relative Quantitative Comparisons of the Extracellular Protein Profiles ofStaphylococcus aureusUAMS-1 and ItssarA,agr, andsarA agrRegulatory Mutants Using One-Dimensional Polyacrylamide Gel Electrophoresis and Nanocapillary Liquid Chromatography Coupled with Tandem Mass Spectrometry

“…One (Q6GJK9, aaa [sle1]) of these proteins has been identified as an N-acetylmuramyl-L-alanine amidase and is involved with cell-to-cell separation (33) and adherence to the extracellular matrix proteins, fibrinogen, fibronectin, and vitronectin (31). A second protein (Q6GED5) was originally identified as an immunodominant staphylococcal antigen (IsaA) reactive with antisera taken from patients with MRSA sepsis (39).…”

Essential Role for the Major Autolysin in the Fibronectin-Binding Protein-Mediated Staphylococcus aureus Biofilm Phenotype

“…This mechanism explains why the Atl-derived peptidoglycan hydrolases are primarily active at the site of cell separation and why rates of autolysis are higher in WTA mutants (22,32,38,65). The repeat domains of Atl have also been shown to bind various host extracellular matrix proteins, including vitronectin and fibronectin (1,(24)(25)(26).…”

Bacterial Virulence in the Moonlight: Multitasking Bacterial Moonlighting Proteins Are Virulence Determinants in Infectious Disease

“…A number of the autolysins of the staphylococci have been shown to function also as fibronectin binding proteins. These include Aaa (autolysin/adhesion of Staphylococcus aureus) which binds fibronectin with high affinity (K d of 30 nM) and which is involved in bacterial adherence to fibronectin (71). Staphylococcus epidermidis Aae (autolysin/adhesin in S. epidermidis) is homologous to S. aureus Aaa and binds to the 29-kDa heparin-binding module of fibronectin (70).…”