CHARACTERIZATION OF THE VIBRIO-ANGUILLARUM FUR GENE - ROLE IN REGULATION OF EXPRESSION OF THE FATA OUTER-MEMBRANE PROTEIN AND CATECHOLS

The chromosomally encoded Vibrio anguillarum fur gene was characterize d. The amino acid sequence of the Fur protein showed a very high degre e of homology with those of V. cholerae and V. vulnificus. The degree of homology was lower, although still high,,vith the Escherichia coli and Yersinia pestis Fur amino acid sequences, while the lowest degree of homology was found with the Pseudomonas aeruginosa Fur protein. The C-terminal portion of Fur is the least conserved region among these P ur proteins. Within this portion, Cove regions spanning amino acids 10 5 to 121 and 132 to the end are the least conserved. A certain degree of variation is also present in the N termini spanning amino acids 28 to 46. Regulation of expression of the V. anguillarum fur gene hy iron was not detected by immunoblot analysis. Mutations in the cloned fur gene were generated either by site-directed mutagenesis (the Lys-77 wa s changed to a Gly to generate the derivative FurG77) or by insertion of a DNA fragment harboring the aph gene in the same position. FurG77 was impaired in its ability to regulate a reporter gene with the Fur b ox in its promoter, while the insertion mutant was completely inactive . V. anguillarum fur mutants were obtained by isolating manganese-resi stant derivatives. In one of these mutants, which encoded a Fur protei n with an apparent lower molecular weight, the regulation of the produ ction of catechols add synthesis of the outer membrane protein FatA we re partially lost. In the case of another mutant, no protein was detec ted by anti-Fur serum. This derivative shelved a total lack of regulat ion of biosynthesis of catechols and FatA protein by iron.