ИСТИНА

D-amino acid oxidase (EC 1.4.3.3., DAAO) is FAD-containing flavoprotein catalyzing stereospecific oxidative deamination of various D-amino acids to corresponding alpha-keto-acids and ammonia. DAAO plays important role in living cells and the enzyme is widely used as biocatalysts in several biotech and biosensor applications. We have identified 4 new orthologous genes encoding potential DAAOs in the genome oа thermotolerant methylotrophic yeast Ogataea parapolymorpha DL-1. Transcriptome analysis showed marked differences in their expression in glucose- and methanol- grown cells. Gene disruptions, in situ enzymatic screens and heterologous expression in E.coli strains were used to determine substrate specificity and in vivo functions of the encoded proteins. We found that HPODL_02914 and HPODL_02165 genes encode DAAO paralogs with increased expression on glucose. Both genes are capable to support the growth of O.parapolymorpha on the medium with D-alanine as sole nitrogen source. The HPDL_400 gene encodes D-aspartate oxidase. The major DAAO overexpressed on methanol is encoded by HPODL_02082 gene. Comparative analysis of the substrate specificity of the encoded protein with published data on substrate specificity of DAAO from Candida boidinii showed notable differences in their relative activity towards D-Met, D-Ser and D-Phe. Studies are in progress towards identification if subcellular localization, physiological role, enzymatic and physico-chemical properties of encoded DAAO/DDO. Supported by RFBR grant 17-04-01487