RECOMBINANT BOVINE CONGLUTININ, LACKING THE N-TERMINAL AND COLLAGENOUS DOMAINS, HAS LESS CONGLUTINATION ACTIVITY BUT IS ABLE TO INHIBIT HEMAGGLUTINATION BY INFLUENZA-A VIRUS

Conglutinin is a bovine serum protein which was first described as a v ertebrate lectin. This protein belongs to the family of C-type lectins . These lectins are composed of four characteristic domains: (1) an N- terminal cysteine-rich domain, (2) a collagen-like domain, (3) a neck domain and (4) a carbohydrate recognition domain (CRD). Recently lecti ns have been shown to function as immunoglobulin-independent defence m olecules due to a complement-mediated mechanism or opsonization. Our p revious study showed that bovine conglutinin can inhibit haemagglutina tion by influenza A viruses and act by directly neutralizing them due to its lectin properties. In order to elucidate the biological role of the collagen-like domain, a recombinant partial conglutinin lacking t his collagen-like domain was produced in an Escherichia coli system an d its biological activities were examined. A 497 bp sequence, consisti ng of a short collagen region (two repeats of G-X-Y amino acid sequenc es), the neck domain, and the CRD of conglutinin cDNA, was amplified b y the reverse-transcriptase PCR technique. The cDNA was transferred to a bacterial expression vector system (pRSET-A) and stable transfectan ts with a high level of conglutinin production were obtained. SDS/PAGE and Western blotting analyses showed a recombinant fusion protein of 27 kDa. Results of a cross-linking study and gel-filtration assay indi cated that the recombinant conglutinin can form a trimeric structure a nd that it has sugar binding activity and specificity similar to that of native conglutinin. The recombinant conglutinin was also found to i nhibit haemagglutination caused by influenza A virus as well as to pos sess less conglutination activity. These results suggest that in order for conglutinin to inhibit haemagglutination caused by the influenza virus, as well as to have sugar binding activity or to form trimers, i t does not require the N-terminal and collagenous domains; however, th ey are essential for full conglutination activity.