TY  - JOUR
A1  - Duchmann, Helmut
A1  - Träger, Lothar
T1  - Purification and characterization of a 3,17 β-hydroxysteroid dehydrogenase from Streptomyces hydrogenans
T2  - Zeitschrift für Naturforschung, C
N2  - 3,17 β-Hydroxysteroid dehydrogenase has been enriched and purified from cytosol of Streptomyces hydrogenans. After ammonium sulfate precipitation and filtration on Sephadex G-100 the enzyme was finally purified by preparative gel electrophoresis and DEAE-Sephadex A-50 chro­matography. Polyacrylamide gel electrophoresis in the presence of sodium dodecylsulfate gave a single band of mobility corresponding to molecular weight of 70 200 ± 2 500. 3 β-. 17 β- as well as 20 β-hydroxy steroids were dehydrogenated by the enzyme in the presence of NAD+. The dehydrogenation proceeded faster than the reduction of the corresponding ketosteroids in the presence of NADH. The enzyme does not accent NADP+ or NADPH as co-substrates. The apparent Km values were calculated to be 11 μᴍ for 5 α-dihydrotestosterone, 20 μᴍ for testosterone ana 68 μᴍ for epiandrosterone in the NAD+-driven reaction, 1.8 x 10-4 m for NADH+ and 1.9 x 10-4 ᴍ for NADH. The catalytic activity was influenced by the ratio of NAD+/ATP. The inhibition by ATP appears to be of a competitive type with respect to NAD+ (Ki 1.15 x 10-3 ᴍ).
After sucrose gradient centrifugation in a preparative ultracentrifuge the enzyme sediments with 4.1 ± 0.1 S as estimated in comparison to other proteins of known sedimentation coefficient. The isoelectric point was determined to be 3.9 with the LKB preparative isoelectric focusing col­umn (pH 2-11) and 4.1 with the analytical flat bed polyacrylamide isofocusing (pH 3 - 5). The number of SH groups was determined to be 2 mol/mol enzyme. In the presence of 6 M urea the fig­ure inceases to 3 mol SH/mol enzyme. In the presence of an excess of p-chloromercuribenzoate the enzyme activity decreases only partially.
KW  - 3,17 β-Hydroxysteroid Dehydrogenase
KW  - Streptomyces hydrogenans
KW  - 20 β-Activity
Y1  - 2014
UR  - http://publikationen.ub.uni-frankfurt.de/frontdoor/index/index/docId/71427
UR  - https://nbn-resolving.org/urn:nbn:de:hebis:30:3-714274
SN  - 0939-5075
SN  - 1865-7125
VL  - 34.1979
IS  - 7-8
SP  - 533
EP  - 540
PB  - Verlag der Zeitschrift für Naturforschung
CY  - Tübingen
ER  -