A naturally occurring point mutation in the hyaluronidase gene (hysA1) of Staphylococcus aureus UAMS-1 results in reduced enzymatic activity

Abstract

Hyaluronic acid is a high-molecular-weight polysaccharide that is widely distributed in animal tissues. Bacterial hyaluronidases degrade hyaluronic acid as secreted enzymes and have been shown to contribute to infection. Staphylococcus aureus UAMS-1 is a clinical isolate that codes for two hyaluronidases (hysA1 and hysA2). Previous research has shown the presence of a full-length HysA1 protein from the S. aureus UAMS-1 strain with no evidence of enzymatic activity. In this study, the coding and upstream promoter regions of hysA1 from the S. aureus UAMS-1 strain were cloned, sequenced, and compared to the hysA1 gene from the S. aureus Sanger 252 strain. A single base change resulting in an E480G amino acid change was identified in the hysA1 gene from the S. aureus UAMS-1 strain when compared to the hysA1 gene from S. aureus Sanger 252. A plasmid copy of hysA1 from S. aureus Sanger 252 transduced into an S. aureus UAMS-1 hysA2 deletion mutant strain restored near wild-type levels of enzymatic activity. Homology modeling of the HysA1 hyaluronidase was performed with SWISS-MODEL using hyaluronidase from Streptococcus pneumoniae as the template, followed by a series of structural analyses using PyMOL, PLIP, PDBsum, and HOPE servers. This glutamic acid is highly conserved among hyaluronidases from Staphylococcus and other gram-positive bacteria. A series of structural analyses suggested that Glu-480 in HysA1 is critically responsible for maintaining the structural and functional ensemble of the catalytic and tunnel-forming residues, which are essential for enzyme activity. The missense mutation of Glu-480 to Gly introduces a loss of side chain hydrogen bond interactions with key residues Arg-360 and Arg-364, which are responsible for the tunnel topology, resulting in displacement of the substrate from an ideal position for catalysis through a localized conformational change of the active site. There is a high degree of relatedness among several gram-positive bacterial hyaluronidases; the loss of enzymatic activity of HysA1 in the S. aureus UAMS-1 strain is most likely caused by the mutation identified in our study. The role of hyaluronidase in staphylococcal infection and the redundancy of this gene are yet to be determined.