The cell surface expression of group 2 capsular polysaccharides in Escherichia coli: the role of KpsD, RhsA and a multi-protein complex at the pole of the cell.
The export of large negatively charged capsular polysaccharides across the outer membrane represents a significant challenge to Gram negative bacteria. In the case of Escherichia coli group 2 capsular polysaccharides, the mechanism of export across the outer membrane was unknown, with no identified candidate outer membrane proteins. In this paper we demonstrate that the KpsD protein, previously believed to be a periplasmic protein, is an outer membrane protein involved in the export of group 2 capsular polysaccharides across the outer membrane. We demonstrate that KpsD and KpsE are located at the poles of the cell and that polysaccharide biosynthesis and export occurs at these polar sites. By in vivo chemical cross-linking and MALDI-TOF-MS analysis we demonstrate the presence of a multi-protein biosynthetic/export complex in which cytoplasmic proteins involved in polysaccharide biosynthesis could be cross-linked to proteins involved in export across the inner and outer membranes. In addition, we show that the RhsA protein, of previously unknown function, could be cross-linked to the complex and that a rhsA mutation reduces K5 biosynthesis suggesting a role for RhsA in coupling biosynthesis and export.
McNulty
Clodagh
C
Faculty of Life Sciences, 1.800 Stopford Building, The University of Manchester, Oxford Road, Manchester M13 9PT, UK.
Thompson
James
J
Barrett
Brendan
B
Lord
Liz
L
Andersen
Christian
C
Roberts
Ian S
IS
D016667:Bacterial Capsules / Q000032:analysis / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D016764:Cell Polarity; D004926:Escherichia coli / Q000166:cytology / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism; D009154:Mutation; D033903:Periplasmic Proteins / Q000032:analysis / Q000378:metabolism*; D011135:Polysaccharides, Bacterial / Q000032:analysis / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D029968:Escherichia coli Proteins; C088016:KpsD protein, E coli; D046912:Multiprotein Complexes; D033903:Periplasmic Proteins; D011135:Polysaccharides, Bacterial; C509296:RhsA protein, E coli