Abstract
7α-hydroxysteroid dehydrogenase (7α-HSDH) can catalyse the oxidation of C7 α-OH of the steroid nucleus in the bile acid metabolism. In the paper we determined the crystal structure of 7α-HSDH from Clostridium absonum (CA 7α-HSDH) complexed with taurochenodeoxycholic acid (TCDCA) and NADP+ by X-ray diffraction, which, as a tetramer, possesses the typical α/β folding pattern. The four subunits of an asymmetric unit lie in the fact that there are the stable hydrophobic interactions between Q-axis-related subunits. Significantly, we captured an active state of the NADP+, confirming that nicotinamide moiety of NADP+ act as electron carrier in the dehydrogenation. On the basis of crystal structure analysis, site-directed mutagenesis and MD simulation, furthermore, we find that the guanidinium of Arg38 can form the stable cation-π interaction with the adenine ring of NADP+, and the cation-π interaction and hydrogen bonds between Arg38 and NADP+ have a significant anchor effect on the cofactor binding to CA 7α-HSDH.
Highlights
7α-hydroxysteroid dehydrogenase (7α-HSDH) can catalyse the oxidation of C7 α-OH of the steroid nucleus in the bile acid metabolism
We find that the hydrogen bond between Arg[38] and 2′ -phosphate of adenine ribose of NADP+ still exsits, but the H-bond is very weak because the donor-acceptor distances of the H-bond remain mostly between 3.2 Å and 3.8 Å during 5 ns dynamics
The tetramer of cholic acid (CA) 7α -HSDH lies in the fact that there are the stable hydrophobic interactions between subunit A and C (B and D) and the hydrogen bonds between subunit A and D (B and C)
Summary
7α-hydroxysteroid dehydrogenase (7α-HSDH) can catalyse the oxidation of C7 α-OH of the steroid nucleus in the bile acid metabolism. 7α -hydroxysteroid dehydrogenases (7α -HSDH) is responsible for reversibly catalysing the oxidation of C7 α -oriented hydroxyl of the steroid nucleus in the bile acid metabolism and has been detected in numerous genera of bacteria[3,4,5,6,7] and in mammal[8,9]. The first crystal structure of NADP(H)-dependent SDR member (mouse lung carbonyl reductase, PDB code: 1CYD) was determined in 1996 and this study[19] drew a conclusion the key determinant for cofactor specificity were the electrostatic environment surrounding the 2′ -phosphate or hydroxyl of the adenosine ribose moiety of NADP(H) or NAD(H). The three-dimensional structure the 7α -HSDH utilising NADP(H) has not been determined and their mechanism for cofactor specificity is still not clear
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