Candida albicans Possesses Sap7 as a Pepstatin A-Insensitive Secreted Aspartic Protease

Abstract

Candida albicans exists in most people, but opportunistically causes severe diseases. One of the major virulence factors is secreted aspartic protease family, encoded by 10 SAP genes (1). Sap isozymes are suggested to contribute to many virulence processes. However, there is a controversy whether Sap family is really a major virulence factor or not, because use of pepstatin A showed little protective effect. In this study, we found Sap7 was a pepstatin A‐insensitive protease and tried to determine the mechanism of pepstatin A‐insensitivity. SAP7 gene was cloned from C. albicans and heterogeneously produced by Pichia pastoris. Proteolytic activity of wild type Sap7 and alanine‐substituted mutants was measured using FRETS‐25Ala library. As a result, we found that Sap7 was pepstatin A‐insensitive and that the M242A and T467A mutants had sensitivity to pepstatin A. M242 and T467 were located in the entrance to an active site, and the substitution of alanine made the entrance wider. This alteration might enable pepstatin A accessibility to the center (2). Grants; JSPS for Young Scientists and Regional Innovation Creation R&D Programs.

Characterization of Antimicrobial Peptides toward the Development of Novel Antibiotics

“…For example, Plasmodium spp., the causative organisms of malaria, require proteases for full virulence [158]. C. albicans secretes ten types of aspartic proteases to degrade host tissues and evade immune cells [159,160]. Thus, designing an AMP that is activated by virulent proteases may confer high selectivity toward each pathogenic organism.…”

Comparison of anti-Candida albicans activities of halogenomethylsulfonyl derivatives

“…We previously demonstrated (Staniszewska et al, 2014a) that compensatory up-regulation of SAP7 occurs in the Dsap mutant under serum influence. Moreover, according to (Aoki et al, 2012) pepstatin A did not have any effect on the Sap7 activity despite inhibiting other Saps suggesting a role of this protein in drug resistance. Therefore, in this study we additionally assayed the influence of Compound no.…”

“…Sap7 is the most divergent and least known member of the whole SAP gene family (Dos Santos, 2010). (Aoki et al, 2012) demonstrated that amino acid residues in close proximity to the enzyme's active site affect Sap7 insensitivity to pepstatin A. Among 9 amino acids identified, M242A and T467 prohibit pepstatin A to access to the active site of Sap7.…”

Differential Proteinase Patterns among Candida albicans Strains Isolated from Root Canal and Lingual Dorsum: Possible Roles in Periapical Disease

“…All types of Sap genes are suggested to contribute to various virulence processes, such as adhesion, invasion, degradation of host proteins, and macrophage escape. Therefore, it has been suggested that the virulence of C. albicans strains correlates with the level of proteolysis activity as well as with the number of Sap genes (3)(4)(5)(6).…”