This project proposes to study the structure and activities of BJ38, a lectin purified from extracts of Bradyrhizobium japonicum. This protein binds to glycoconjugates containing lactose (Lac) or galactose (Gal). Derivatives of Gal at C-2, galactosamine and N-acetyl-D-galactosamine bound with much lower affinity. This specificity correlated with the behavior of intact Rhizobium in four carbohydrate-specific binding assays: (a) adsorption to Lac-Sepharose beads; (b) heterotypic binding to cultured soybean (SB-1) cells; (c) heterotypic adhesion to soybean roots; and (d) homotypic autoagglutination. These observations indicate the possibility that all of these binding activities were mediated by the same component(s) and the BJ38 is a key candidate for the carbohydrate-specific binding. The specific objectives of the proposed research are: [a] to demonstrate and quantitate the binding of purified BJ38 to soybean cells and to B. japonicum cells; [b] to test the possibility that soluble BJ38 bound to soybean cells will block adhesion of B. japonicum and that BJ38 bound to the bacteria will block autoagglutination; [c] to test whether univalent Fab fragments of antibodies directed against BJ38 will inhibit heterotypic binding to soybean cells and homotypic autoagglutination; [d] to analyze the expression and localization of BJ38 in B. japonicum cells; [e] to carry out molecular cloning of BJ38; [f] to determine the partial amino acid sequence of BJ38 at the protein level and to derive the complete amino acid sequence from the DNA sequence; [g] to test whether carbohydrate-specific binding can be restored in mutant B. japonicum by complementation with the BJ38 gene.
|Ho, S C; Wang, J L; Schindler, M et al. (1994) Carbohydrate binding activities of Bradyrhizobium japonicum. III. Lectin expression, bacterial binding, and nodulation efficiency. Plant J 5:873-84|
|Loh, J T; Ho, S C; Wang, J L et al. (1994) Carbohydrate binding activities of Bradyrhizobium japonicum: IV. Effect of lactose and flavones on the expression of the lectin, BJ38. Glycoconj J 11:363-70|
|Loh, J T; Ho, S C; de Feijter, A W et al. (1993) Carbohydrate binding activities of Bradyrhizobium japonicum: unipolar localization of the lectin BJ38 on the bacterial cell surface. Proc Natl Acad Sci U S A 90:3033-7|