RESUMO
D-Galacturonic acid reductase was purified from a psychrophilic yeast strain of Cryptococcus diffluens, which was isolated from Satho, a traditional alcohol drink in Thailand. This enzyme, named Cd-GalUAR, assimilates D-galacturonic acid and requires NADPH as a cofactor. Cd-GalUAR is about 45 kDa and stable from pH 6.5 to 7.5 and up to 35°C. Its optimum pH and temperature are pH 7.0 and 40°C, respectively. However, 80% of its maximum activity remained at 4°C. The reaction of Cd-GalUAR from D-galacturonic acid produces L-galactonic acid, which was identified by (13)C NMR and LC-MS. Three amino acid sequences were determined from trypsin-digested peptides of Cd-GalUAR. Similar sequences are found in many NAD or NADP oxidoreductases, including some D-galacturonate reductases. Our results suggest that Cd-GalUAR is the first D-galacturonate reductase identified in yeast.
Assuntos
Oxirredutases do Álcool/metabolismo , Cryptococcus/enzimologia , Proteínas Fúngicas/metabolismo , Oxirredutases do Álcool/isolamento & purificação , Bebidas Alcoólicas/microbiologia , Clonagem Molecular , Proteínas Fúngicas/isolamento & purificação , Ácidos Hexurônicos/metabolismo , Concentração de Íons de Hidrogênio , Peso Molecular , Álcool Oxidorredutases Dependentes de NAD(+) e NADP(+) , NADP/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Análise de Sequência de Proteína , Especificidade por Substrato , Açúcares Ácidos/metabolismo , Temperatura , TailândiaRESUMO
One psychrophilic yeast strain, that grew well in a cold environment such as in a refrigerator, was isolated from the yeast starter (Loog-pang) of a traditional alcohol drink in Thailand. The isolated strain OPU-FC11 was identified as Cryptococcus diffluens by the assay for 26S ribosomal DNA and the test for carbon source assimilation. OPU-FC11 showed a good amount of growth at 4 degrees 0 at which a commonly found yeast like Saccharomyces cerevisiae did not grow, and produced cold-adapted enzymes that showed a relatively high activity at lower temperatures.