Purification of 7 alpha-hydroxysteroid dehydrogenase from Escherichia coli strain 080.
Purification studies of 7 alpha-hydroxysteroid dehydrogenase (7 alpha-HSDH) (EC 1.1.1.159) from Escherichia coli 080 showed that 1.59-fold purification could be achieved by heating (60 degrees C for 10 min) the ultracentrifuged enzyme preparation, and 6.46-fold purification was achieved by subsequent precipitation with ammonium sulfate. Further purification on Sephadex G-100 gel gave 10.1-fold purification. After pooling and concentrating the active fractions obtained from the Sephadex G-100 filtration, an 11.1-fold purification was achieved using DEAE-cellulose chromatography. The purified enzyme produced a single band on polyacrylamide gel electrophoresis and its molecular weight was determined to be 54,000. The enzyme was immunogenic and showed immunoprecipitation with homologus antisera.
Prabha
V
V
Department of Microbiology, Panjab University, Chandigarh, India.
Gupta
M
M
Seiffge
D
D
Gupta
K G
KG
D002848:Chromatography, DEAE-Cellulose; D002850:Chromatography, Gel; D004926:Escherichia coli / Q000201:enzymology*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D006913:Hydroxysteroid Dehydrogenases / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D008970:Molecular Weight; D010084:Oxidation-Reduction; D011233:Precipitin Tests; D014461:Ultracentrifugation
D006913:Hydroxysteroid Dehydrogenases; C025376:7 alpha-hydroxysteroid dehydrogenase