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Mechanism of polysialic acid chain elongation in Escherichia coli K1
Author(s) -
Steenbergen S. M.,
Vimr E. R.
Publication year - 1990
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.1990.tb00629.x
Subject(s) - polysialic acid , biology , transposon mutagenesis , sialyltransferase , mutant , complementation , mutagenesis , gene , microbiology and biotechnology , biochemistry , escherichia coli , transposable element , genetics , sialic acid , cell adhesion , cell , neural cell adhesion molecule
Summary Understanding the mechanisms of polysialic acid synthesis in Escherichia coli K1 requires a molecular description of the polymerase complex. Since the number of potential models explaining polysialic acid assembly would be constrained if only one sialyltransferase were required for this process, the phenotypes of a sialyltransferase null mutation generated by transposon mutagenesis were investigated. The chromosomal insertion mutation was mapped by Southern hybridization analysis and by complementation with plasmid subclones, demonstrating that sialyltransferase is encoded by neuS , a gene implicated previously as coding for the polymerase (Vimr et al. , 1989). As expected, if only one gene encoded sialyltransferase, the null mutant had undetectable polymerase activity when assayed with endogenous or exogenous acceptors, and accumulated sugar nucleotide precursors intracellularly. Nested deletion analysis of neuS ruled out polarity effects of transposon insertion mutation and provided more precise mapping of the sialyltransferase structural gene. Maxicell analysis of the nested deletion set implicated a 34000 molecular weight polypeptide as the neuS gene product. These results, together with biochemical characterization of sialyltransferase reaction products in the wild type, indicated that CMP‐sialic acid is the probable sialosyl donor for polysialic acid elongation and that chain growth is by sequential addition of monomeric units.