Project description:Pseudogymnoascus destructans is a pathogenic fungus responsible for White-nose Syndrome, a disease afflicting multiple species of North American bats. P. destructans infects susceptible bats during hibernation, invading dermal tissue and causing extensive tissue damage. In contrast, other Pseudogymnoascus species are non-pathogenic and cross-species comparisons may therefore reveal factors that contribute to virulence. In this study, we compared the secretome of P. destructans with that from several closely related Pseudogymnoascus species. A diverse set of hydrolytic enzymes were discovered, including a putative serine peptidase, PdCP1 that was unique to the P. destructans secretome. This enzyme was expressed in yeast and the substrate preference was discovered using a multiplexed-substrate profiling method based on enzymatic degradation of a sequence-diverse synthetic peptide library. The cleaved products were identified by mass spectrometry. In general, the peptide substrates were sequentially truncated from the carboxyl terminus revealing that this enzyme is a bona fide carboxypeptidase. Peptides with arginine located in the penultimate position were rapidly cleaved and therefore a fluorescent substrate containing arginine was used to biochemically characterize the enzyme and screen a selection of peptidase inhibitors. Antipain and leupeptin were found to be potent inhibitors of PdCP1 activity.
