We have previously identified that Leishmania mexicana cysteine proteases (CPs) are virulence factors. We have now produced a recombinant L. mexicana CP, CPB2.8, which has similar enzymatic activity to native enzyme. Inoculation of CPB2.8 (< or =5 microg) into the footpads of BALB/c mice not only up-regulated mRNA transcripts for IL-4 and IL-4 production in the draining popliteal lymph nodes, but also polarized splenocyte anti-CD3 stimulated responses toward a Th2 bias as measured by increased IL-5 production compared with controls. In agreement with promoting a Th2 response, CPB2.8 also induced strong specific IgE responses in treated mice as well as increasing whole IgE levels. Inhibition of the enzyme activity of CPB2.8 by treatment with E-64 ablated the enzyme's ability to induce IgE. Significantly, infection of mice with CPB-deficient parasites failed to stimulate production of IgE, unlike infection with wild-type parasites. Furthermore, enzymatically active (<0.1 U/ml) but not E-64-inactivated CPB2.8 was able to proteolytically cleave CD23 and CD25, although not B220 or CD4 from murine lymphocytes. These properties are similar to those demonstrated by the house dust mite allergen Der p I and provide an explanation for the immunomodulatory activity of the CPB2.8 virulence factor. Vaccination with CPB2.8 enhanced L. mexicana lesion growth compared with control animals. Nevertheless, vaccination with IL-12 and CPB2.8 resulted in a degree of protection associated with inhibition of lesion growth and a Th1 response. Thus, CPB2.8 is a potent Th2-inducing molecule capable of significant vaccine potential if administered with a suitable adjuvant.