Testosterone dehydrogenase activity in koala liver: characterisation of cofactor and steroid substrate differences
I. Stupans a,*, S. Kong a, A. Kirlich a, R.A. McKinnon a, M. Murray b
a Centre for Pharmaceutical Research, School of Pharmacy and Medical Sciences, University of South Australia, North Terrace, Adelaide, SA, 5000 , Australia
b School of Physiology and Pharmacology, University of New South Wales, NSW, 2052 , Australia
We have studied the hepatic microsomal 17β-hydroxysteroid dehydrogenase (17β-HSD) capacity of koala (Phascolarctos cinereus) and tammar wallaby (Macropus eugenii). A detailed comparison of the activity in hepatic fractions from koala and rat was made. Hepatic microsomal NADP-supported 17β-HSD activity was signiﬁcantly higher in koala (11.64 ± 3.35 nmoles/mg protein/min), (mean ± S.D.) than in tammar wallaby liver (1.52 ± 0.79 nmoles/mg protein/min). However, when NAD was utilised as cofactor the activity was similar in both marsupial species (2.83 ± 2.03 nmoles/mg protein/min, koala; 0.70 ± 0.71 nmoles/mg protein/min, tammar wallaby). Data for rat indicated a cofactor preference for NAD rather than NADP (17.94 ± 6.40 nmoles/mg protein/min, NAD; 2.18 ± 1.04 nmoles/mg protein/min, NADP). Michaelis–Menten parameters for the kinetics of 17β-HSD testosterone oxidation by NADP and NAD were determined in the koala. The Km for testosterone was of the order of 10.0–24.0 µM (n=6) irrespective of the cofactor used, whilst the Km for NADP was 0.28–0.43 µM (n=2) and for NAD was 13.9–18.5 µM (n=2). 17β-estradiol was found to be an inhibitor of both NAD- and NADP- supported 17β-HSD activity. These ﬁndings indicate that NADP-mediated, but not NAD-mediated testosterone dehydrogenation is a major pathway of steroid biotransformation in koala liver; the reaction is less extensive in fractions from wallaby, human and rat. Such species-related differences in cofactor preference may contribute along with species differences in gene expression to observed rates of 17β-HSD activity in mammals.