Evolutionary Selection on Barrier Activity: Bar1 Is an Aspartyl Protease with Novel Substrate Specificity

ABSTRACT Peptide-based pheromones are used throughout the fungal kingdom for coordinating sexual responses between mating partners. Here, we address the properties and function of Bar1, an aspartyl protease that acts as a “barrier” and antagonist to pheromone signaling in multiple species. Candida a...

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Autores principales:	Stephen K. Jones, Starlynn C. Clarke, Charles S. Craik, Richard J. Bennett
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2015
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/02af93dcc5074eba9b425b0730e6d660
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spelling	oai:doaj.org-article:02af93dcc5074eba9b425b0730e6d6602021-11-15T15:41:23ZEvolutionary Selection on Barrier Activity: Bar1 Is an Aspartyl Protease with Novel Substrate Specificity10.1128/mBio.01604-152150-7511https://doaj.org/article/02af93dcc5074eba9b425b0730e6d6602015-12-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01604-15https://doaj.org/toc/2150-7511ABSTRACT Peptide-based pheromones are used throughout the fungal kingdom for coordinating sexual responses between mating partners. Here, we address the properties and function of Bar1, an aspartyl protease that acts as a “barrier” and antagonist to pheromone signaling in multiple species. Candida albicans Bar1 was purified and shown to exhibit preferential cleavage of native α pheromone over pheromones from related fungal species. This result establishes that protease substrate specificity coevolved along with changes in its pheromone target. Pheromone cleavage by Bar1 occurred between residues Thr-5 and Asn-6 in the middle of the tridecapeptide sequence. Surprisingly, proteolytic activity was independent of the amino acid residues present at the scissile bond and instead relied on residues at the C terminus of α pheromone. Unlike most aspartyl proteases, Bar1 also exhibited a near-neutral pH optimum and was resistant to the class-wide inhibitor pepstatin A. In addition, genetic analysis was performed on C. albicans BAR1 and demonstrated that the protease not only regulates endogenous pheromone signaling but also can limit interspecies pheromone signaling. We discuss these findings and propose that the unusual substrate specificity of Bar1 is a consequence of its coevolution with the α pheromone receptor Ste2 for their shared peptide target. IMPORTANCE Pheromones are important for intraspecies communication across the tree of life. In the fungal kingdom, extracellular proteases play a key role in antagonizing pheromone signaling in multiple species. This study examines the properties and function of Candida albicans Bar1, an aspartyl protease that cleaves and thereby inactivates α pheromone. We demonstrate that Bar1 plays important roles in regulating both intra- and interspecies pheromone signaling. The fungal protease shows preferential activity on the endogenous pheromone, but, surprisingly, cleavage activity is dependent on amino acid residues distal to the scissile bond. We propose that the unusual substrate specificity of Bar1 is a direct result of coevolution with Ste2, the receptor for α pheromone, for recognition of the same peptide target. The novel specificity of Bar1 reveals the complex forces shaping the evolution of mating pathways in fungi and uncovers a protease with potentially important applications in the biotechnology industry.Stephen K. JonesStarlynn C. ClarkeCharles S. CraikRichard J. BennettAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 6, Iss 6 (2015)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Stephen K. Jones Starlynn C. Clarke Charles S. Craik Richard J. Bennett Evolutionary Selection on Barrier Activity: Bar1 Is an Aspartyl Protease with Novel Substrate Specificity
description	ABSTRACT Peptide-based pheromones are used throughout the fungal kingdom for coordinating sexual responses between mating partners. Here, we address the properties and function of Bar1, an aspartyl protease that acts as a “barrier” and antagonist to pheromone signaling in multiple species. Candida albicans Bar1 was purified and shown to exhibit preferential cleavage of native α pheromone over pheromones from related fungal species. This result establishes that protease substrate specificity coevolved along with changes in its pheromone target. Pheromone cleavage by Bar1 occurred between residues Thr-5 and Asn-6 in the middle of the tridecapeptide sequence. Surprisingly, proteolytic activity was independent of the amino acid residues present at the scissile bond and instead relied on residues at the C terminus of α pheromone. Unlike most aspartyl proteases, Bar1 also exhibited a near-neutral pH optimum and was resistant to the class-wide inhibitor pepstatin A. In addition, genetic analysis was performed on C. albicans BAR1 and demonstrated that the protease not only regulates endogenous pheromone signaling but also can limit interspecies pheromone signaling. We discuss these findings and propose that the unusual substrate specificity of Bar1 is a consequence of its coevolution with the α pheromone receptor Ste2 for their shared peptide target. IMPORTANCE Pheromones are important for intraspecies communication across the tree of life. In the fungal kingdom, extracellular proteases play a key role in antagonizing pheromone signaling in multiple species. This study examines the properties and function of Candida albicans Bar1, an aspartyl protease that cleaves and thereby inactivates α pheromone. We demonstrate that Bar1 plays important roles in regulating both intra- and interspecies pheromone signaling. The fungal protease shows preferential activity on the endogenous pheromone, but, surprisingly, cleavage activity is dependent on amino acid residues distal to the scissile bond. We propose that the unusual substrate specificity of Bar1 is a direct result of coevolution with Ste2, the receptor for α pheromone, for recognition of the same peptide target. The novel specificity of Bar1 reveals the complex forces shaping the evolution of mating pathways in fungi and uncovers a protease with potentially important applications in the biotechnology industry.
format	article
author	Stephen K. Jones Starlynn C. Clarke Charles S. Craik Richard J. Bennett
author_facet	Stephen K. Jones Starlynn C. Clarke Charles S. Craik Richard J. Bennett
author_sort	Stephen K. Jones
title	Evolutionary Selection on Barrier Activity: Bar1 Is an Aspartyl Protease with Novel Substrate Specificity
title_short	Evolutionary Selection on Barrier Activity: Bar1 Is an Aspartyl Protease with Novel Substrate Specificity
title_full	Evolutionary Selection on Barrier Activity: Bar1 Is an Aspartyl Protease with Novel Substrate Specificity
title_fullStr	Evolutionary Selection on Barrier Activity: Bar1 Is an Aspartyl Protease with Novel Substrate Specificity
title_full_unstemmed	Evolutionary Selection on Barrier Activity: Bar1 Is an Aspartyl Protease with Novel Substrate Specificity
title_sort	evolutionary selection on barrier activity: bar1 is an aspartyl protease with novel substrate specificity
publisher	American Society for Microbiology
publishDate	2015
url	https://doaj.org/article/02af93dcc5074eba9b425b0730e6d660
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Evolutionary Selection on Barrier Activity: Bar1 Is an Aspartyl Protease with Novel Substrate Specificity

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