Specificity characterization of the alpha-mating factor hormone by Kex2 protease

Specificity characterization of the alpha-mating factor hormone by Kex2 protease

Author Manfredi, Marcella Araujo Google Scholar
Antunes, Alyne Alexandrino Google Scholar
Passos Jesus, Larissa de Oliveira Google Scholar
Juliano, Maria Aparecida Autor UNIFESP Google Scholar
Juliano, Luiz Autor UNIFESP Google Scholar
de Souza Judice, Wagner Alves Google Scholar
Abstract Kex2 is a Ca2+-dependent serine protease from S. cerevisiae. Characterization of the substrate specificity of Kex2 is of particular interest because this protease serves as the prototype of a large family of eukaryotic subtilisin-related proprotein-processing proteases that cleave sites consisting of pairs or clusters of basic residues. Our goal was to study the prime region subsite S' of Kex2 because previous studies have only taken into account non-prime sites using AMC substrates but not the specificity of prime sites identified through structural modeling or predicted cleavage sites. Therefore, we used peptides derived from Abz-KR down arrow EADQ-EDDnp and Abz-YKR down arrow EADQ-EDDnp based on the pro-a-mating factor sequence. The specificity of Kex2 due to basic residues at P-1' is affected by the type of residue in the P-3 position. Some residues in P-1' with large or bulky side chains yielded poor substrate specificity. The k(cat)/K-M values for peptides with P-2' substitutions containing Tyr in P-3 were higher than those obtained for the peptides without Tyr. In fact, P' and P modifications mainly promoted changes in k(cat) and K-M, respectively. The pH profile of Kex2 was fit to a double-sigmoidal pH-titration curve. The specificity results suggest that Kex2 might be involved in the processing of the putative cleavage sites in a poly peptide involved in cell elongation, hyphal formation and the processing of a toxin, which result in host cell lysis. In summary, the specificity of Kex2 is dependent on the set of interactions with prime and non-prime subsites, resulting in synergism. (C) 2016 Elsevier B.V. and Societe Francaise de Biochimie et Biologie Moleculaire (SFBBM). All rights reserved.
Keywords Specificity
Proprotein-processing proteases
alpha-mating factor hormone
xmlui.dri2xhtml.METS-1.0.item-coverage Paris
Language English
Sponsor Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
Grant number FAPESP: 2014/02205-1
Date 2016
Published in Biochimie. Paris, v. 131, p. 149-158, 2016.
ISSN 0300-9084 (Sherpa/Romeo, impact factor)
Publisher Elsevier France-Editions Scientifiques Medicales Elsevier
Extent 149-158
Origin http://dx.doi.org/10.1016/j.biochi.2016.10.003
Access rights Closed access
Type Article
Web of Science ID WOS:000390513800015
URI https://repositorio.unifesp.br/handle/11600/56619

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