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Characterization of two aminotransferases from Candida albicans

Abstract

Aminoadipate aminotransferase (AmAA) is an enzyme of α-aminoadipate pathway (AAP) for l-lysine biosynthesis. AmAA may also participated in biosynthesis or degradation of aromatic amino acids and in d-tryptophan based pigment production. The AAP is unique for fungal microorganisms. Enzymes involved in this pathway have specific structures and properties. These features can be used as potential molecular markers. Enzymes catalyzing reactions of l-lysine biosynthesis in Candida albicans may also become new targets for antifungal chemotherapy. Search of the NCBI database resulted in identification of two putative aminoadipate aminotransferase genes from Candida albicans: ARO8 (ORFs 19.2098 and 19.9645) and YER152C (ORFs 19.1180 and 19.8771). ARO8 from C. albicans exhibits 53% identity to ARO8 from S. cerevisiae, while YER152C exhibits 30% identity to ARO8 and 45% to YER152C from S. cerevisiae. We amplified two genes from the C. albicans genome: ARO8 and YER152C. Both were cloned and expressed as His-tagged fusion proteins in E. coli. The purified Aro8CHp gene product revealed aromatic and α-aminoadipate aminotransferase activity. Basic molecular properties of the purified protein were determined. We obtained catalytic parameters of Aro8CHp with aromatic amino acids and aminoadipate (AA) (Km(L-Phe) 0.05±0.003 mM, Km(L-Tyr) 0.1±0.008 mM, Km(L-AA) 0.02±0.006 mM) and confirmed the enzyme broad substrate spectrum. The assays also demonstrated that this enzyme may use 2-oxoadipate and 2-oxoglutarate (2-OG) as amino acceptors. Aro8CHp exhibited pH optima range of 8, which is similar to AmAA from S. cerevisiae. Our results also indicate that CaYer152Cp has a possible role only in aromatic amino acids degradation, in contrast to CaAro8CHp.

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DOI:
Digital Object Identifier (open in new tab) 10.18388/abp.2015_1158
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Category:
Articles
Type:
artykuł w czasopiśmie wyróżnionym w JCR
Published in:
Acta Biochimica Polonica no. 62, pages 903 - 912,
ISSN: 0001-527X
Language:
English
Publication year:
2015
Bibliographic description:
Rząd K., Gabriel I.: Characterization of two aminotransferases from Candida albicans// Acta Biochimica Polonica. -Vol. 62, iss. 4 (2015), s.903-912
DOI:
Digital Object Identifier (open in new tab) 10.18388/abp.2015_1158
Verified by:
Gdańsk University of Technology

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