Prieß Marten, Göddeke Hendrik, Groenhof Gerrit, Schäfer Lars V
Theoretical Chemistry, Faculty of Chemistry and Biochemistry, Ruhr-University Bochum, D-44780 Bochum, Germany.
Department of Chemistry and Nanoscience Center, University of Jyväskylä, P.O. Box 35, FI-40014 Jyväskylä, Finland.
ACS Cent Sci. 2018 Oct 24;4(10):1334-1343. doi: 10.1021/acscentsci.8b00369. Epub 2018 Oct 5.
Hydrolysis of nucleoside triphosphate (NTP) plays a key role for the function of many biomolecular systems. However, the chemistry of the catalytic reaction in terms of an atomic-level understanding of the structural, dynamic, and free energy changes associated with it often remains unknown. Here, we report the molecular mechanism of adenosine triphosphate (ATP) hydrolysis in the ATP-binding cassette (ABC) transporter BtuCD-F. Free energy profiles obtained from hybrid quantum mechanical/molecular mechanical (QM/MM) molecular dynamics (MD) simulations show that the hydrolysis reaction proceeds in a stepwise manner. First, nucleophilic attack of an activated lytic water molecule at the ATP γ-phosphate yields ADP + HPO as intermediate product. A conserved glutamate that is located very close to the γ-phosphate transiently accepts a proton and thus acts as catalytic base. In the second step, the proton is transferred back from the catalytic base to the γ-phosphate, yielding ADP + HPO . These two chemical reaction steps are followed by rearrangements of the hydrogen bond network and the coordination of the Mg ion. The rate constant estimated from the computed free energy barriers is in very good agreement with experiments. The overall free energy change of the reaction is close to zero, suggesting that phosphate bond cleavage itself does not provide a power stroke for conformational changes. Instead, ATP binding is essential for tight dimerization of the nucleotide-binding domains and the transition of the transmembrane domains from inward- to outward-facing, whereas ATP hydrolysis resets the conformational cycle. The mechanism is likely relevant for all ABC transporters and might have implications also for other NTPases, as many residues involved in nucleotide binding and hydrolysis are strictly conserved.
核苷三磷酸（NTP）的水解在许多生物分子系统的功能中起着关键作用。然而，就与催化反应相关的结构、动力学和自由能变化的原子水平理解而言，催化反应的化学过程往往仍不为人知。在此，我们报告了ATP结合盒（ABC）转运蛋白BtuCD - F中三磷酸腺苷（ATP）水解的分子机制。从混合量子力学/分子力学（QM/MM）分子动力学（MD）模拟获得的自由能分布表明，水解反应以逐步方式进行。首先，一个活化的裂解水分子对ATP的γ - 磷酸进行亲核攻击，产生ADP + HPO作为中间产物。一个位于非常靠近γ - 磷酸的保守谷氨酸短暂接受一个质子，从而作为催化碱。在第二步中，质子从催化碱转移回γ - 磷酸，产生ADP + HPO 。这两个化学反应步骤之后是氢键网络的重排和Mg离子的配位。根据计算出的自由能垒估计的速率常数与实验结果非常吻合。反应的总体自由能变化接近零，这表明磷酸键断裂本身并不为构象变化提供动力冲程。相反，ATP结合对于核苷酸结合结构域的紧密二聚化以及跨膜结构域从内向型向外向型的转变至关重要，而ATP水解则重置构象循环。该机制可能与所有ABC转运蛋白相关，并且可能对其他NTP酶也有影响，因为许多参与核苷酸结合和水解的残基是严格保守的。
