title: Hyaluronan molecular weight is controlled by UDP-N-acetylglucosamine concentration in Streptococcus zooepidemicus.
abstract: The molecular weight of hyaluronan is important for its rheological and biological function. The molecular mechanisms underlying chain termination and hence molecular weight control remain poorly understood, not only for hyaluronan synthases but also for other beta-polysaccharide synthases, e.g. cellulose, chitin, and 1,3-betaglucan synthases. In this work, we manipulated metabolite concentrations in the hyaluronan pathway by overexpressing the five genes of the hyaluronan synthesis operon in Streptococcus equi subsp. zooepidemicus. Overexpression of genes involved in UDP-glucuronic acid biosynthesis decreased molecular weight, whereas overexpression of genes involved in UDP-N-acetylglucosamine biosynthesis increased molecular weight. The highest molecular mass observed was at 3.4 +/- 0.1 MDa twice that observed in the wild-type strain, 1.8 +/- 0.1 MDa. The data indicate that (a) high molecular weight is achieved when an appropriate balance of UDP-N-acetylglucosamine and UDP-glucuronic acid is achieved, (b) UDP-N-acetylglucosamine exerts the dominant effect on molecular weight, and (c) the wild-type strain has suboptimal levels of UDP-N-acetylglucosamine. Consistent herewith molecular weight correlated strongly (rho = 0.84, p = 3 x 10(-5)) with the concentration of UDP-N-acetylglucosamine. Data presented in this paper represent the first model for hyaluronan molecular weight control based on the concentration of activated sugar precursors. These results can be used to engineer strains producing high molecular weight hyaluronan and may provide insight into similar polymerization mechanisms in other polysaccharides.
authors.affiliation: Australian Institute for Bioengineering and Nanotechnology, University of Queensland, Queensland 4072, Australia.
authors.forename: Wendy Yiting
authors.initials: WY
authors.lastname: Chen
authors.forename: Esteban
authors.initials: E
authors.lastname: Marcellin
authors.forename: Jacky
authors.initials: J
authors.lastname: Hung
authors.forename: Lars Keld
authors.initials: LK
authors.lastname: Nielsen
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects / Q000502:physiology; D005822:Genetic Vectors; D014453:Glucuronosyltransferase / Q000235:genetics / Q000378:metabolism; D000076002:Hyaluronan Synthases; D006820:Hyaluronic Acid / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism*; D013294:Lactococcus lactis / Q000235:genetics / Q000378:metabolism; D008828:Microbiological Techniques; D008970:Molecular Weight; D009561:Nisin / Q000494:pharmacology; D009876:Operon / Q000235:genetics; D010957:Plasmids; D018502:Streptococcus equi / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D014537:Uridine Diphosphate N-Acetylglucosamine / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D009561:Nisin; D014537:Uridine Diphosphate N-Acetylglucosamine; D006820:Hyaluronic Acid; D014453:Glucuronosyltransferase; D000076002:Hyaluronan Synthases