Fucosyltransferase in Klebsiella Pneumoniae---Cloning and Characterization

Previously, our study discovered that the capsule of Klebsiella pneumoniae (KP) that caused liver abscess (LA) contained high amount of fucose. This bacterium caused high fatality rate in infected mice. In contrary, the KP that caused urinary tract infection (UTI) did not contain fucose and caused no fatality in mice. Both strains caused bacteremia. This led us to suspect that fucose (also a component in human tissue) enabled the LA-KP to evade host immune reaction. Our experiment of bacteria-phagocytes interaction in mice peritoneum revealed that the LA-KP rarely interacted with macrophages while the UTI-KP readily attached to macrophage surface indicating the LA-KP could survive and multiply to high concentration in blood to invade various organs without being cleared by phagocytosis. In human colon cancer, studies revealed that cancer cells expressed Lewis antigens are easily metastasize to liver. Lewis antigens contain fucose on their polysaccharide surface. We therefore hypothesize that LA-KPs may contain Lewis antigen for them to migrate to liver and cause liver abscess. Fucosyltransferase (FucT) is needed to form Lewis antigens. Our preliminary results discovered the LA-KP contained Lewis antigen. However, FucT has not been reported in KPs. Our goal in this proposal is to design primers of FucT using human FucT (a total of 12) gene sequences and amplify from KP genome by polymerase chain reaction. Genes cloned will be screened for FucT activity, expressed in protein and analyzed for enzyme activity. The enzyme will be crystallized and the structure of the protein will be analyzed (This part will collaborated with Dr. Lin CH in Academia Sinica). The FucT sequence will be compared with FucT of other source to investigate the origin of the gene. This project may enable us to understand the virulence factor in KP and may serve as a therapeutic target.

Project ID:PC9808-0539
External Project ID:NSC98-2320-B182-032