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Specificity of the hyaluronate lyase of group-B streptococcus toward unsulphated regions of chondroitin sulphate.

ABSTRACT: The purification and properties of a hyaluronate lyase secreted by Streptococcus agalactiae, which is believed to facilitate the invasion of host tissues by the organism, have been described previously [Pritchard, Lin, Willingham and Baker (1994) Arch. Biochem. Biophys. 315, 431-436]. The specificity of the limited cleavage of chondroitin sulphate by the enzyme is the subject of this report. To simplify the task, a chondroitin sulphate from the Swarm rat chondrosarcoma, which contains only 4-sulphated and unsulphated disaccharide repeats, was used in this study. Tetrasaccharides from an ovine testicular hyaluronidase digest of the chondroitin sulphate were isolated, identified and tested as substrates of the streptococcal hyaluronate lyase. Only tetrasaccharides with an unsulphated disaccharide at the reducing end were cleaved (by elimination at the N-acetylgalactosaminidic bond). Thus chondroitin sulphate chains are cleaved by the action of this lyase at every unsulphated disaccharide repeat, but release of unsaturated unsulphated disaccharides only occurs from sites where two or more sequential unsulphated disaccharide repeats are present. Analysis of the chondrosarcoma chondroitin sulphate showed that of approximately five unsulphated disaccharide repeats per chain, two are clustered. The ability of group-B streptococcal hyaluronate lyase to cleave chondroitin sulphate may allow the organisms to invade tissues more efficiently. The demonstrated specific and highly limited cleavage of chondroitin sulphate by this bacterial lyase promises to be a useful tool in the determination of chondroitin sulphate structure and variability.

SUBMITTER: Baker JR

PROVIDER: S-EPMC1218764 | biostudies-other | 1997 Oct

REPOSITORIES: biostudies-other

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Project description:Streptococcus dysgalactiae subsp. equisimilis (SDSE) causes cellulitis, bacteremia, and invasive diseases, such as streptococcal toxic shock syndrome. Although SDSE infection is more prevalent among elderly individuals and those with diabetes mellitus than infections with Streptococcus pyogenes (Group A streptococci; GAS) and Streptococcus agalactiae (Group B streptococci; GBS), the mechanisms underlying the pathogenicity of SDSE remain unknown. SDSE possesses a gene hylD encoding a hyaluronate lyase (HylD), whose homologue (HylB) is involved in pathogenicity of GBS, while the role of HylD has not been characterized. In this study, we focused on the enzyme HylD produced by SDSE; HylD cleaves hyaluronate (HA) and generates unsaturated disaccharides via a ?-elimination reaction. Hyaluronate-agar plate assays revealed that SDSE promoted dramatic HA degradation. SDSE expresses both HylD and an unsaturated glucuronyl hydrolase (UGL) that catalyzes the degradation of HA-derived oligosaccharides; as such, SDSE was more effective at HA degradation than other ?-hemolytic streptococci, including GAS and GBS. Although HylD shows some homology to HylB, a similar enzyme produced by GBS, HylD exhibited significantly higher enzymatic activity than HylB at pH 6.0, conditions that are detected in the skin of both elderly individuals and those with diabetes mellitus. We also detected upregulation of transcripts from hylD and ugl genes from SDSE wild-type collected from the mouse peritoneal cavity; upregulated expression of ugl was not observed in ?hylD SDSE mutants. These results suggested that disaccharides produced by the actions of HylD are capable of triggering downstream pathways that catalyze their destruction. Furthermore, we determined that infection with SDSE?hylD was significantly less lethal than infection with the parent strain. When mouse skin wounds were infected for 2 days, intensive infiltration of neutrophils was observed around the wound areas infected with SDSE wild-type but not SDSE?hylD. Our investigation suggested that HylD and UGL play important roles in nutrient acquisition from hosts, followed by the bacterial pathogenicity damaging host tissues.

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Specificity of the hyaluronate lyase of group-B streptococcus toward unsulphated regions of chondroitin sulphate.

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