CLONING AND HETEROLOGOUS EXPRESSION OF AN ALPHA-1,3-FUCOSYL-TRANSFERASE GENE FROM THE GASTRIC PATHOGEN HELICOBACTER-PYLORI

Helicobacter pylori is an important human pathogen which causes both g astric and duodenal ulcers and is also associated with gastric cancer and lymphoma. This microorganism has been shown to express cell surfac e glycoconjugates including Lewis X (Le(x)) and Lewis Y. These bacteri al oligosaccharides are structurally similar to tumor-associated carbo hydrate antigens found in mammals, In this study, we report the clonin g of a novel alpha 1,3-fucosyltransferase gene (HpfucT) involved in th e biosynthesis of Le(x) within H. pylori. The deduced amino acid seque nce of HpfucT consists of 478 residues with the calculated molecular m ass of 56,194 daltons, which is approximately 100 amino acids longer t han known mammalian alpha 1,3/1,4-fucosyltransferases. The similar to 52-kDa protein encoded by HpfucT was expressed in Escherichia coli CSR DE3 cells and gave rise to alpha 1,3-fucosyltransferase activity but n either alpha 1,4-fucosyltransferase nor alpha 1,2-fucosyltransferase a ctivity as characterized by radiochemical assays and capillary zone el ectrophoresis, Truncation of the C-terminal 100 amino acids of HpFuc-T abolished the enzyme activity, An approximately 72-amino acid region of HpFuc-T exhibits significant sequence identity (40-45%) with the hi ghly conserved C-terminal catalytic domain among known mammalian and c hicken alpha 1,3-fucosyltransferases. These lines of evidence indicate that the HpFuc-T represents the bacterial alpha 1,3-fucosyltransferas e. In addition, several structural features unique to HpFuc-T, includi ng 10 direct repeats of seven amino acids and the lack of the transmem brane segment typical for known eukaryotic alpha 1,3-fucosyltransferas es, were revealed, Notably, the repeat region contains a leucine zippe r motif previously demonstrated to be responsible for dimerization of various basic region-leucine zipper proteins, suggesting that the HpFu c-T protein could form dimers.