EC Number   |
|---|
    1.14.13.22 | - |
| 1.14.13.22 | alcohol dehydrogenase(Mesotoga infera)/cyclohexanone monooxygenase(Thermocrispum municipale) fusion construct expressed in Escherichia coli cells |
| 1.14.13.22 | cells centrifuged, washed with reaction buffer (0.1 M glycine/NaOH buffer, pH 9.0), disruption by ultrasonic processor, centrifugation, supernatants used as crude enzyme extracts |
| 1.14.13.22 | crude extract of Rhodococcus cells is partially purified using ammonium sulfate fractionation (50-70%), chromatography on Butyl-Toyopearl 650S with a linear gradient of 30%-0% (NH4)2SO4, and gel filtration on a Sephadex G-150 column. Purification of the recombinant enzyme: cells centrifuged and lysed with a French press, crude extract centrifuged, supernatant loaded on a DEAE-Sepharose FF column equilibrated with 50 mM sodium phosphate buffer (pH 7.0), elution with 0-0.2 M NaCl gradient, ammonium sulfate added to active fractions (30%), loaded onto a Butyl-S-Sepharose 6 FF column equilibrated with 30% ammonium sulfate in 50 mM sodium phosphate buffer (pH 7.0), elution with 30-0% ammonium sulfate gradient, active fractions pooled, concentrated, applied to two combined Superose 6 HR (10/300)/Superose 12 HR (10/300) gel filtration columns equilibrated with 50 mM sodium phosphate buffer (pH 7.0) containing 150 mM NaCl, elution with same buffer, pooled, concentrated with YM 10 membrane |
| 1.14.13.22 | industrial scale production |
| 1.14.13.22 | native enzyme and recombinant His-tagged enzyme from Escherichia coli and Saccharomyces cerevisiae |
| 1.14.13.22 | partial purification of recombinant enzyme |
| 1.14.13.22 | partially |
| 1.14.13.22 | partially, recombinant from Escherichia coli |
| 1.14.13.22 | recombinant enzyme 21fold from Escherichia coli and native enzyme to homogeneity by ion exchange and affinity chromatography |
