The nucleotide sequence determination of catalases of three medically important yeasts using newly designed degenerated primers

Abstract

We have developed degenerated primers for the isolation of several fungal species catalases, based on the known catalase genes of several yeast species. Using a combination of degenerated primers and the nested polymerase chain reaction (PCR) method, we were able to obtain PCR products from Candida dubliniensis, C. tropicalis, and C. glabrata. The nucleotide sequence of the PCR products amplified showed that those fragments contained sequences homologous with the known Candida catalases, indicating the usefulness of the designed primers. We determined the nucleotide sequences of the open reading frames and respective 5′ untranscribed regions of these yeasts and compared each sequence with that of the respective related species. The difference between the deduced amino acid sequence of catalase of C. dubliniensis and C. albicans was 5 in 485 amino acids. The nucleotide sequence of C. glabrata catalase was identical to the sequence results from the genome sequence project which was recently released, whereas that of the catalase of the C. tropicalis clinical isolate was not the same as the strain Pk233, n-alkane-utilizing C. tropicalis. The catalase activities of all the strains tested so far were activated by short-term hydrogen peroxide treatment, suggesting that common mechanisms were involved in the induction of catalase activity, although the nucleotide sequences of the 5′ untranscribed region of these yeasts were diversified.