Lidholt K, Fjelstad M
Department of Medical and Physiological Chemistry, University of Uppsala, The Biomedical Center, S-751 23 Uppsala, Sweden.
J Biol Chem. 1997 Jan 31;272(5):2682-7. doi: 10.1074/jbc.272.5.2682.
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 beta-D-glucuronidase, and the [3H]GalNAc could be removed by beta-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 (GlcAbeta1-4GlcNAcalpha1-4)n (Lidholt, K., Fjelstad, M., Jann, K., and Lindahl, U. (1994) Carbohydr. Res. 255, 87-101). In contrast, chain elongation of hyaluronan (GlcAbeta1-3GlcNAcbeta1-4)n is claimed to occur at the reducing end (Prehm, P. (1983) Biochem. J. 211, 181-189).
大肠杆菌K4细菌合成一种荚膜多糖（GalNAc-GlcA（果糖））n，其碳水化合物主链与软骨素相同。分别用源自荚膜多糖的受体以及放射性标记的UDP-[14C]GlcA和UDP-[3H]GalNAc测定细菌膜中的GlcA-和GalNAc-转移酶活性。结果表明，去果糖基化的寡糖（软骨素）可作为GlcA-和GalNAc-转移酶的底物。放射性标记的产物可被软骨素酶AC完全降解；[14C]GlcA单元可被β-D-葡萄糖醛酸酶去除，[3H]GalNAc可被β-N-乙酰己糖胺酶去除。经测试，一种用于GlcA-转移酶活性的果糖基化寡糖受体无活性。这些结果表明，K4多糖的链延长反应与软骨素链的聚合反应方式相同，即通过将单糖单元逐个添加到聚合物的非还原端。这使得K4多糖的生物合成成为研究硫酸软骨素生物合成的一个有趣的平行系统。在大肠杆菌荚膜多糖的生物合成中，对于聚唾液酸（NeuNAc）n（Rohr，T. E.，和Troy，F. A.（1980）J. Biol. Chem. 255，2332 - 2342）和K5多糖（GlcAbeta1-4GlcNAcalpha1-4）n（Lidholt，K.，Fjelstad，M.，Jann，K.，和Lindahl，U.（1994）Carbohydr. Res. 255，87 - 101）， earlier已经证明了类似的机制。相比之下，透明质酸（GlcAbeta1-3GlcNAcbeta1-4）n的链延长据说是在还原端发生的（Prehm，P.（1983）Biochem. J. 211，181 - 189）。
