Destructin-1 is a collagen-degrading endopeptidase secreted by Pseudogymnoascus destructans, the causative agent of white-nose syndrome

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 112; no. 24; pp. 7478 - 7483
• Main Authors: O’Donoghue, Anthony J., Knudsen, Giselle M., Beekman, Chapman, Perry, Jenna A., Johnson, Alexander D., DeRisi, Joseph L., Craik, Charles S., Bennett, Richard J.
• Format: Journal Article
• Language: English
• Published: United States National Acad Sciences 16.06.2015; National Academy of Sciences
• Subjects: Amino Acid Sequence; Animal diseases; Animals; Ascomycota - enzymology; Ascomycota - genetics; Ascomycota - pathogenicity; Base Sequence; Bats; Biochemical analysis; Biochemistry; Biological Sciences; Catalytic Domain; Chiroptera; Chiroptera - microbiology; collagen; Collagenases - chemistry; Collagenases - genetics; Collagenases - metabolism; DNA, Fungal - genetics; Enzymes; Fungal Proteins - chemistry; Fungal Proteins - genetics; Fungal Proteins - metabolism; fungi; Low temperature; Mammals; Models, Molecular; Molecular Sequence Data; Mycoses - microbiology; Mycoses - veterinary; Pathogenesis; Proteolysis; Pseudogymnoascus destructans; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Sequence Homology, Amino Acid; serine; serine proteinases; structural proteins; Tissues; Virulence; pathogenesis; secretome; connective tissue; fungi; peptidase
• ISSN: 0027-8424; 1091-6490; 1091-6490
• DOI: 10.1073/pnas.1507082112

This work identifies molecular factors produced by the fungus Pseudogymnoascus destructans , the causative agent of white-nose syndrome in bats. Our study reveals the repertoire of redox enzymes and hydrolytic enzymes secreted by P. destructans . We establish that a secreted serine peptidase, Destructin-1, is a major component of the P. destructans secretome. This peptidase was purified and shown to degrade collagen, the major structural protein in mammalian connective tissue. Furthermore, chemical inhibition of Destructin-1 blocked collagen degradation in conditioned media from P. destructans . We therefore propose that serine endopeptidases aid in invasive growth and tissue destruction by the fungus, and represent potential targets for therapeutic intervention in white-nose syndrome. Pseudogymnoascus destructans is the causative agent of white-nose syndrome, a disease that has caused the deaths of millions of bats in North America. This psychrophilic fungus proliferates at low temperatures and targets hibernating bats, resulting in their premature arousal from stupor with catastrophic consequences. Despite the impact of white-nose syndrome, little is known about the fungus itself or how it infects its mammalian host. P. destructans is not amenable to genetic manipulation, and therefore understanding the proteins involved in infection requires alternative approaches. Here, we identify hydrolytic enzymes secreted by P. destructans , and use a novel and unbiased substrate profiling technique to define active peptidases. These experiments revealed that endopeptidases are the major proteolytic activities secreted by P. destructans , and that collagen, the major structural protein in mammals, is actively degraded by the secretome. A serine endopeptidase, hereby-named Destructin-1, was subsequently identified, and a recombinant form overexpressed and purified. Biochemical analysis of Destructin-1 showed that it mediated collagen degradation, and a potent inhibitor of peptidase activity was identified. Treatment of P. destructans -conditioned media with this antagonist blocked collagen degradation and facilitated the detection of additional secreted proteolytic activities, including aminopeptidases and carboxypeptidases. These results provide molecular insights into the secretome of P. destructans , and identify serine endopeptidases that have the clear potential to facilitate tissue invasion and pathogenesis in the mammalian host.