Amino acid polymorphisms in the fibronectin-binding repeats of fibronectin-binding protein A affect bond strength and fibronectin conformation.
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abstract
The Staphylococcus aureus cell surface contains cell wall-anchored proteins such as fibronectin-binding protein A (FnBPA) that bind to host ligands (e.g. fibronectin; Fn) present in the extracellular matrix of tissue or coatings on cardiac implants. Recent clinical studies have found a correlation between cardiovascular infections caused by S. aureus and nonsynonymous SNPs in FnBPA. Atomic force microscopy (AFM), surface plasmon resonance (SPR), and molecular simulations were used to investigate interactions between Fn and each of eight 20-mer peptide variants containing amino acids Ala, Asn, Gln, His, Ile, and Lys at positions equivalent to 782 and/or 786 in Fn-binding repeat-9 of FnBPA. Experimentally measured bond lifetimes (1/koff) and dissociation constants (Kd = koff/kon), determined by mechanically dissociating the Fnpeptide complex at loading rates relevant to the cardiovascular system, varied from the lowest-affinity H782A/K786A peptide (0.011 s, 747 m) to the highest-affinity H782Q/K786N peptide (0.192 s, 15.7 m). These atomic force microscopy results tracked remarkably well to metadynamics simulations in which peptide detachment was defined solely by the free-energy landscape. Simulations and SPR experiments suggested that an Fn conformational change may enhance the stability of the binding complex for peptides with K786I or H782Q/K786I (Kdapp = 0.2-0.5 m, as determined by SPR) compared with the lowest-affinity double-alanine peptide (Kdapp = 3.8 m). Together, these findings demonstrate that amino acid substitutions in Fn-binding repeat-9 can significantly affect bond strength and influence the conformation of Fn upon binding. They provide a mechanistic explanation for the observation of nonsynonymous SNPs in fnbA among clinical isolates of S. aureus that cause endovascular infections.
Casillas-Ituarte, N. N., Cruz, C., Lins, R. D., DiBartola, A. C., Howard, J., Liang, X., ... Lower, S. K.
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Casillas-Ituarte, Nadia N||Cruz, Carlos HB||Lins, Roberto D||DiBartola, Alex C||Howard, Jessica||Liang, Xiaowen||Höök, Magnus||Viana, Isabelle FT||Sierra-Hernández, M Roxana||Lower, Steven K
