Biosynthesis of the Escherichia coli K4 Capsule Polysaccharide
A PARALLEL SYSTEM FOR STUDIES OF GLYCOSYLTRANSFERASES IN CHONDROITIN FORMATION*
- From the Department of Medical and Physiological Chemistry, University of Uppsala, The Biomedical Center, S-751 23 Uppsala, Sweden
- ‡ To whom correspondence should be addressed: Dept. of Medical and Physiological Chemistry, The Biomedical Center, Box 575, S-751 23 Uppsala, Sweden. Tel.: 46-18-17-44-43; Fax: 46-18-17-42-09; E-mail: Kerstin.Lidholt{at}medkem.uu.se
Abstract
Escherichia coli K4 bacteria synthesize a capsule polysaccharide (GalNAc-GlcA(fructose))n with the carbohydrate backbone identical to chondroitin. GlcA- and GalNAc-transferase activities from the bacterial membrane were assayed with acceptors derived from the capsule polysaccharide and radiolabeled UDP-[14C]GlcA and UDP-[3H]GalNAc, respectively. It was shown that defructosylated oligosaccharides (chondroitin) could serve as substrates for both the GlcA- and the GalNAc-transferases. The radiolabeled products were completely degraded with chondroitinase AC; the [14C]GlcA unit could be removed by β-D-glucuronidase, and the [3H]GalNAc could be removed by β-N-acetylhexosaminidase. A fructosylated oligosaccharide acceptor tested for GlcA-transferase activity was found to be inactive. These results indicate that the chain elongation reaction of the K4 polysaccharide proceeds in the same way as the polymerization of the chondroitin chain, by the addition of the monosaccharide units one by one to the nonreducing end of the polymer. This makes the biosynthesis of the K4 polysaccharide an interesting parallel system for studies of chondroitin sulfate biosynthesis. In the biosynthesis of capsule polysaccharides from E. coli, a similar mechanism has earlier been demonstrated for polysialic acid (NeuNAc)n (Rohr, T. E., and Troy, F. A. (1980) J. Biol. Chem. 255, 2332-2342) and for the K5 polysaccharide (GlcAβ1-4GlcNAcα1-4)n (Lidholt, K., Fjelstad, M., Jann, K., and Lindahl, U. (1994) Carbohydr. Res. 255, 87-101). In contrast, chain elongation of hyaluronan (GlcAβ1-3GlcNAcβ1-4)n is claimed to occur at the reducing end (Prehm, P. (1983) Biochem. J. 211, 181-189).
Footnotes
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↵* This work was supported by Grants 10440, 10155, and 2309 from the Swedish Medical Research Council and by Polysackaridforskning AB (Uppsala, Sweden). The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵1 The abbreviation used is:
- Fru
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fructose.
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- Received September 19, 1996.
- © 1997 by The American Society for Biochemistry and Molecular Biology, Inc.
