A wide range of mucosal pathogens produce an enzyme, IgA protease, that cleaves human IgA1. We are interested in the role these enzymes play in virulence and are studying a number of aspects of their synthesis, secretion and function. We have cloned IgA protease genes (iga) from Haemophilus influenzae serotypes b, c and d and from Neisseria gonorrhoeae. The cloned genes are expressed in E.coli leading to production of IgA proteases with the same specificity as those produced in the parent organisms. These enzymes cleave the IgA1 substrate at different bonds in the hinge region. We intend to characterize the genes by DNA sequence analysis and to determine the structural basis for their different specificities. The IgA proteases are secreted from the cell and we wish to elucidate the mechanism by which this occurs. We have evidence that the enzymes are synthesized as precursors which are processed during secretion. The identification of intermediates involved in this process and the mechanism of processing is a major goal. In collaboration with Dr. David Stephens we will look at the role of IgA protease in H.influenzae infection using a nasopharyngeal tissue explant model. We have cloned the IgA protease gene from Streptococcus sanguis and will determine the DNA sequence of this gene to see how its product is related to the proteases of the Gram negative organisms indicated above. We will modify the cloned gene in order to generate S. sanguis mutants that lack IgA protease. Such mutants will be used to look at the effect of iga mutations on adherence of S. sanguis in a model system.