Formation of a new O-polysaccharide in Escherichia coli O86 via disruption of a glycosyltransferase gene involved in O-unit assembly.
The majority of hetero-polysaccharide biosynthesis in Gram-negative bacteria utilizes the wzy-dependent pathway, in which repeating O-units are first synthesized in the cytosol and then subsequently translocated to the periplasmic face of the inner membrane where polymerization is initiated by the Wzy polymerase. Wzy proteins share little primary sequence homology and are specific for their cognate O-unit structures. Our previous studies on O-polysaccharide biosynthesis in Escherichia coli O86 identified the wbnI gene, which encodes a galactosyltransferase responsible for the introduction of alpha-(1-->3)-Galp residues as side chains of the polysaccharide. In this work, we functionally inactivated the wbnI gene and showed that the mutant strain produced a different polysaccharide without the side chain Galp residue. The yield of the polysaccharide was substantially lower than the one produced by the wild-type strain. This study indicated that the complete O-unit structure is the preferred substrate for the polymerization, thus further confirming the specificity of Wzy. On the other hand, these studies also suggest that the Wzy polymerase might have moderate tolerance of side-chain truncated O-unit substrates.
Yi
Wen
W
Department of Biochemistry, The Ohio State University, Columbus, OH 43210, USA.
Zhu
Lizhi
L
Guo
Hongjie
H
Li
Mei
M
Li
Jianjun
J
Wang
Peng George
PG
D002240:Carbohydrate Sequence; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D019075:Electrophoresis, Capillary; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics* / Q000378:metabolism; D017353:Gene Deletion; D016695:Glycosyltransferases / Q000235:genetics* / Q000378:metabolism; D006602:Hexosyltransferases / Q000235:genetics* / Q000378:metabolism; D015151:Immunoblotting; D008070:Lipopolysaccharides / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008956:Models, Chemical; D008969:Molecular Sequence Data; D015394:Molecular Structure; D019081:O Antigens / Q000096:biosynthesis* / Q000737:chemistry / Q000378:metabolism
D029968:Escherichia coli Proteins; D008070:Lipopolysaccharides; D019081:O Antigens; D015500:Chloramphenicol O-Acetyltransferase; D016695:Glycosyltransferases; D006602:Hexosyltransferases; C085652:O-antigen polymerase; C513753:Wzy polymerase, E coli