Molecular analysis of the O-antigen gene cluster of Escherichia coli O86:B7 and characterization of the chain length determinant gene (wzz).
Escherichia coli O86:B7 has long been used as a model bacterial strain to study the generation of natural blood group antibody in humans, and it has been shown to possess high human blood B activity. The O-antigen structure of O86:B7 was solved recently in our laboratory. Comparison with the published structure of O86:H2 showed that both O86 subtypes shared the same O unit, yet each of the O antigens is polymerized from a different terminal sugar in a different glycosidic linkage. To determine the genetic basis for the O-antigen differences between the two O86 strains, we report the complete sequence of O86:B7 O-antigen gene cluster between galF and hisI, each gene was identified based on homology to other genes in the GenBank databases. Comparison of the two O86 O-antigen gene clusters revealed that the encoding regions between galF and gnd are identical, including wzy genes. However, deletion of the two wzy genes revealed that wzy in O86:B7 is responsible for the polymerization of the O antigen, while the deletion of wzy in O86:H2 has no effect on O-antigen biosynthesis. Therefore, we proposed that there must be another functional wzy gene outside the O86:H2 O-antigen gene cluster. Wzz proteins determine the degree of polymerization of the O antigen. When separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the lipopolysaccharide (LPS) of O86:B7 exhibited a modal distribution of LPS bands with relatively short O units attached to lipid A-core, which differs from the LPS pattern of O86:H2. We proved that the wzz genes are responsible for the different LPS patterns found in the two O86 subtypes, and we also showed that the very short type of LPS is responsible for the serum sensitivity of the O86:B7 strain.
Guo
Hongjie
H
Department of Biochemistry and Chemistry, The Ohio State University, Columbus, OH 43210, USA.
Yi
Wen
W
Shao
Jun
J
Lu
Yuquan
Y
Zhang
Wenpeng
W
Song
Jing
J
Wang
Peng George
PG
D002240:Carbohydrate Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D006801:Humans; D008070:Lipopolysaccharides / Q000737:chemistry; D008957:Models, Genetic; D008969:Molecular Sequence Data; D005810:Multigene Family / Q000235:genetics*; D016296:Mutagenesis; D019081:O Antigens / Q000235:genetics*; D015183:Restriction Mapping
D008070:Lipopolysaccharides; D019081:O Antigens