Isolation and Characterization of the Epoxide Hydrolase-Encoding Gene from Xanthophyllomyces dendrorhous

Abstract

The epoxide hydrolase (EH)-encoding gene (EPH1) from the basidiomycetous yeast Xanthophyllomyces dendrorhous was isolated. The genomic sequence has a 1,236-bp open reading frame which is interrupted by eight introns that encode a 411-amino-acid polypeptide with a calculated molecular mass of 46.2 kDa. The amino acid sequence is similar to that of microsomal EH and belongs to the α/β hydrolase fold family. The EPH1 gene was not essential for growth of X. dendrorhous in rich medium under laboratory conditions. The Eph1-encoding cDNA was functionally expressed in Escherichia coli. A sixfold increase in specific activity was observed when we used resting cells rather thanX. dendrorhous. The epoxides 1,2-epoxyhexane and 1-methylcyclohexene oxide were substrates for both native and recombinant Eph1. Isolation and characterization of the X. dendrorhous EH-encoding gene are essential steps in developing a yeast EH-based epoxide biotransformation system.

Estudio de la producción de astaxantina por Xanthophyllomyces dendrorhous = Study of astaxanthin production by Xanthophyllomyces dendrorhous

“…La comparación de la secuencia de ADN genómico del gen crtS con la secuencia del ADNc del gen ast descrita por Hoshino y colaboradores (2000) puso de manifiesto que el gen constaba de 18 exones (en las posiciones 1-118, 201-249, 365-481, 554-645, 769-983, 1067-1100, 1190-1332, 1406-1465, 1548-1647, 1724-1820, 1898-2008, 2085-2118, 2209-2280, 2429-2516, 2580-2637, 2714-2787, 2865-2889 y 2980-3166) y que estaba interrumpido por 17 intrones (cuyos tamaños oscilan entre 64 y 149 pb distribuidos de forma homogénea a lo largo de todo el gen (Figura 1.6.). Este elevado número de intrones es típico de los genes de X. dendrorhous, incluidos aquellos implicados en la carotenogénesis (crtI: 11 intrones [Verdoes et al, 1999b]; crtYB: 4 intrones [Verdoes et al, 1999a]; eph1: 8 intrones [Visser et al, 1999]; gpd: 6 intrones [Verdoes et al, 1997]).…”