Characterization of the peroxidase activity of CYP119, a thermostable P450 from Sulfolobus acidocaldarius.
We report the cloning, expression, and purification of CYP119, a thermostable enzyme previously thought to derive from Sulfolobus solfataricus. Sequence analysis suggested that, in contrast to the conclusions of earlier studies, the enzyme stems from the closely related Sulfolobus acidocaldarius, and we were indeed able to clone the gene from the genomic DNA of this organism. For the first time, we report here on the peroxidase activity of this enzyme and the optimization of the associated reaction parameters. The optimized reaction conditions were then applied to the biocatalytic epoxidation of styrene. The values obtained for k(cat) (78.2+/-20.6 min(-1)) and K(M) (9.2+/-4.3 mM) indicated an approximately 100-fold increased catalytic activity over previously reported results.
D019843:Archaeal Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D016945:Sulfolobus acidocaldarius / Q000201:enzymology*; D013696:Temperature
D019843:Archaeal Proteins; D017931:DNA Primers; D003577:Cytochrome P-450 Enzyme System; C408874:CYP119 protein, Sulfolobus solfataricus