CLONING, SEQUENCING AND EXPRESSION OF AN EIKENELLA-CORRODENS GENE ENCODING A COMPONENT PROTEIN OF THE LECTIN-LIKE ADHESIN COMPLEX

A lectin-like substance (LS), that was isolated from Eikenella corrode ns (Ec) 1073, migrated as proteins of about 300 and 45 kDa upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing con ditions. In this study, we cloned the gene encoding the 45-kDa protein and predicted its structure and function. Based on the N-terminal 23- amino acid (aa) sequence of this protein, we cloned the region for its N-terminus. We cloned the entire gene by means of gene walking using polymerase chain reaction and Southern hybridization. The nucleotide s equences of cloned fragments revealed an open reading frame encoding a polypeptide of 330 aa (M(r) 35 748). This ORF displayed high homology to those of porins of Neisseria species. Using the T7-expression syst em, the 45-kDa protein was produced in E. coli. Our results suggested that the 45-kDa protein of Ec 1073 is a component of the EcLS complex, and that it is the major outer membrane protein.