Identification of two genes, kpsM and kpsT, in region 3 of the polysialic acid gene cluster of Escherichia coli K1

Abstract

The polysialic acid capsule of Escherichia coli K1, a causative agent of neonatal septicemia and meningitis, is an essential virulence determinant. The 17-kb kps gene cluster, which is divided into three functionally distinct regions, encodes proteins necessary for polymer synthesis and expression at the cell surface. The central region, 2, encodes products required for synthesis, activation, and polymerization of sialic acid, while flanking regions, 1 and 3, are thought to be involved in polymer assembly and transport. In this study, we identified two genes in region 3, kpsM and kpsT, which encode proteins with predicted sizes of 29.6 and 24.9 kDa, respectively. The hydrophobicity profile of KpsM suggests that it is an integral membrane protein, while KpsT contains a consensus ATP-binding domain. KpsM and KpsT belong to a family of prokaryotic and eukaryotic proteins involved with a variety of biological processes, including membrane transport. A previously described kpsT chromosomal mutant that accumulates intracellular polysialic acid was characterized and could be complemented in trans. Results of site-directed mutagenesis of the putative ATP-binding domain of KpsT are consistent with the view that KpsT is a nucleotide-binding protein. KpsM and KpsT have significant similarity to BexB and BexA, two proteins that are essential for polysaccharide capsule expression in Haemophilus influenzae type b. We propose that KpsM and KpsT constitute a system for transport of polysialic acid across the cytoplasmic membrane.