BACKGROUND Pig aldo-keto reductase family 1 member C1 (AKR1C1) belongs to AKR superfamily which catalyzes the NAD(P)H-dependent reduction of various substrates including steroid hormones. Previously we have reported two paralogous pig AKR1C1s, wild-type AKR1C1 (C-type) and C-terminal-truncated AKR1C1 (T-type). Also, the C-terminal region significantly contributes to the NADPH-dependent reductase activity for 5α-DHT reduction. Molecular modeling studies combined with kinetic experiments were performed to investigate structural and enzymatic differences between wild-type AKR1C1 C-type and T-type. RESULTS The results of the enzyme kinetics revealed that Vmax and kcat values of the T-type were 2.9 and 1.6 folds higher than those of the C-type. Moreover, catalytic efficiency was also 1.9 fold higher in T-type compared to C-type. Since x-ray crystal structures of pig AKR1C1 were not available, three dimensional structures of the both types of the protein were predicted using homology modeling methodology and they were used for molecular dynamics simulations. The structural comparisons between C-type and T-type showed that 5α-DHT formed strong hydrogen bonds with catalytic residues such as Tyr55 and His117 in T-type. In particular, C3 ketone group of the substrate was close to Tyr55 and NADPH in T-type. CONCLUSIONS Our results showed that 5α-DHT binding in T-type was more favorable for catalytic reaction to facilitate hydride transfer from the cofactor, and were consistent with experimental results. We believe that our study provides valuable information to understand important role of C-terminal region that affects enzymatic properties for 5α-DHT, and further molecular mechanism for the enzyme kinetics of AKR1C1 proteins.

中文翻译：

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猪醛酮还原酶家族1成员C1中C末端区域的结构重要性及其对酶活性的影响。

背景技术猪醛酮还原酶家族1成员C1（AKR1C1）属于AKR超家族，它催化NAD（P）H依赖的各种底物（包括类固醇激素）的还原。以前，我们已经报道了两个同源猪AKR1C1，即野生型AKR1C1（C型）和C端截短的AKR1C1（T型）。此外，C末端区域显着有助于5α-DHT还原的NADPH依赖性还原酶活性。进行了分子建模研究和动力学实验，以研究野生型AKR1C1 C型和T型之间的结构和酶促差异。结果酶动力学结果表明，T型的Vmax和kcat值比C型分别高2.9倍和1.6倍。此外，与C型催化剂相比，T型催化剂的催化效率也高1.9倍。由于无法获得猪AKR1C1的X射线晶体结构，因此使用同源建模方法预测了两种蛋白质的三维结构，并将其用于分子动力学模拟。C型和T型之间的结构比较表明5α-DHT与T型中的Tyr55和His117等催化残基形成强氢键。特别地，底物的C 3酮基接近于T型的Tyr55和NADPH。结论我们的结果表明，T型中的5α-DHT结合更有利于催化反应，以促进氢化物从辅因子的转移，与实验结果一致。我们相信这项研究提供了宝贵的信息，有助于您了解影响5α-DHT酶性质的C末端区域的重要作用，