Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.36 (hyaluronidase)
 4,606 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An improved assay for hyaluronic acid (HA) synthetase is described that is suitable for rapid processing of large numbers of samples. High background levels of unincorporated radioactivity are removed by passage of the reaction through a Sephadex G-50 spin column. The labeled HA product is then precipitated onto glass fiber filters with cetylpyridinium chloride. Apparent Km values for HA synthetase from Swiss 3T3 fibroblasts are 10.8 and 58.4 microM for UDP-glucuronic acid and UDP-N-acetylglucosamine, respectively. HA synthetase activity of quiescent cells is 4.5% of that found in actively growing cells and is stimulated in response to 10% calf serum. There is a greater than 10-fold increase in HA synthetase activity when cells are harvested with hyaluronidase as compared with trypsin.
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PMID:A filter paper assay for hyaluronic acid synthetase: application to the enzyme from Swiss 3T3 fibroblasts. 754 31

Previous studies reached different conclusions about whether class I hyaluronan synthases (HASs) elongate hyaluronic acid (HA) by addition to the reducing or the nonreducing end. Here we used two strategies to determine the direction of HA synthesis by purified class I HASs from Streptococcus equisimilis and Streptococcus pyogenes. In the first strategy we used each of the two UDP-sugar substrates separately to pulse label either the beginning or the end of HA chains. We then quantified the relative rates of radioactive HA degradation by treatment with beta-glycosidases that act at the nonreducing end. The results with both purified HASs demonstrated that HA elongation occurred at the reducing end. In the second strategy, we used purified S. equisimilis HAS, UDP-glucuronic acid, and UDP[beta-32P]-Glc-NAc to radiolabel nascent HA chains. Under conditions of limiting substrate, the 32P-labeled products were separated from the substrates by paper chromatography and identified as HA-[32P]UDP saccharides based on their degradation by snake venom phosphodiesterase or hyaluronidase and by their binding to a specific HA-binding protein. The 32P radioactivity was chased (released) by incubation with unlabeled UDP-sugars, showing that the HA-UDP linkages turn over during HA biosynthesis. In contrast, HA-[32P]UDP products made by the purified class II Pasteurella multocida HAS were not released by adding unlabeled UDP-sugars, consistent with growth at the nonreducing end for this enzyme. The results demonstrate that the streptococcal class I HAS enzymes polymerize HA chains at the reducing end.
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PMID:Hyaluronan biosynthesis by class I streptococcal hyaluronan synthases occurs at the reducing end. 1566 42