Glucuronyltransferase activity of KfiC from Escherichia coli strain K5 requires association of KfiA: KfiC and KfiA are essential enzymes for production of K5 polysaccharide, N-acetylheparosan.

The Journal of Biological Chemistry
Nobuo SugiuraHideto Watanabe

Abstract

Heparan sulfate is a ubiquitous glycosaminoglycan in the extracellular matrix of most animals. It interacts with various molecules and exhibits important biological functions. K5 antigen produced by Escherichia coli strain K5 is a linear polysaccharide N-acetylheparosan consisting of GlcUA beta1-4 and GlcNAc alpha1-4 repeating disaccharide, which forms the backbone of heparan sulfate. Region 2, located in the center of the K5-specific gene cluster, encodes four proteins, KfiA, KfiB, KfiC, and KfiD, for the biosynthesis of the K5 polysaccharide. Here, we expressed and purified the recombinant KfiA and KfiC proteins and then characterized these enzymes. Whereas the recombinant KfiC alone exhibited no GlcUA transferase activity, it did exhibit GlcUA transferase and polymerization activities in the presence of KfiA. In contrast, KfiA had GlcNAc transferase activity itself, which was unaffected by the presence of KfiC. The GlcNAc and GlcUA transferase activities were analyzed with various truncated and point mutants of KfiA and KfiC. The point mutants replacing aspartic acid of a DXD motif and lysine and glutamic acid of an ionic amino acid cluster, and the truncated mutants deleting the C-terminal and N-terminal sites, revealed the...Continue Reading

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Citations

Aug 24, 2010·The Journal of Biological Chemistry·Hiroyasu OgawaHideto Watanabe
Dec 27, 2011·Applied Microbiology and Biotechnology·Anaïs A E ChavarocheGerrit Eggink
Jun 4, 2011·Bioengineered Bugs·Zhenyu WangRobert J Linhardt
Mar 8, 2013·Carbohydrate Polymers·Anaïs A E ChavarocheGerrit Eggink
Jun 23, 2020·Annual Review of Biochemistry·Christopher A CaffaletteJochen Zimmer
Mar 10, 2021·Applied Biochemistry and Biotechnology·Ganesh NehruSenthilkumar Sivaprakasam
May 24, 2020·International Journal of Biological Macromolecules·Ganesh NehruSenthilkumar Sivaprakasam

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