We have continued to investigate the structure-function relationships and genetic regulation of bacterial ADP-ribosylating toxins using site-directed mutagenesis and recombinant expression systems. Studies of the amino acids essential to the enzymatic/toxic activity of the diarrheagenic toxin from enterotoxingenic Escherichia coli (heat-labile enterotoxin) have identified several residues that are likely to be involved in the active site of this toxin. These residues appear to be suitable sites for efforts directed at creating inactive holotoxoid forms of the heat-labile enterotoxin and perhaps of the homologous toxin from Vibrio cholerae (cholera toxin) for vaccine use. Additional studies have allowed the detection of an intrinsic regulatory mechanism associated with the expression of the pertussis toxin gene. This mechanism, which is continuing to be defined imparts negative (or down) regulation of the first gene in the operon. Precise definition of the mechanism may be of some importance in the construction of Bordetella pertussis strains which are pertussis toxin hyper-producers for vaccine purposes. Other collaborative studies have characterized the biologic and enzymatic properties of mutant forms of pertussis toxin and have permitted the precise evaluation of their suitability for vaccine use.
