中科院大连化学物理研究所复杂分子体系反应动力学研究组（1101组）与该所高级伙伴研究员、美国肯塔基大学湛昌国教授合作，在细胞色素P450 2A6催化尼古丁反应机理研究方面取得了新进展。
酶催化反应机理是当今计算化学研究中的前沿领域和热门课题。理论计算作为一种有力的手段，能够研究酶催化反应的过程，解释实验现象，阐明反应机理，并为进一步的实验研究提供理论基础。尼古丁是烟草中的主要成瘾物质，它在人体中的代谢主要是由细胞色素P450 2A6（CYP2A6）催化的，研究尼古丁和CYP2A6的结合方式以及CYP2A6催化尼古丁的反应机理能够为合理设计CYP2A6抑制剂提供理论支持，为戒烟药物的研发提供一定的理论基础。
细胞色素P450 2A6（CYP2A6）
大连化物所1101组自2006年以来一直致力于酶催化反应机理的理论研究，并相继在J. Phys. Chem. B; ChemBioChem; J. Biol. Inorg. Chem.; Dalton Trans.等杂志上发表了一系列文章。近期，韩克利研究员等人与湛昌国教授合作，利用分子对接、分子动力学模拟、QM/MM计算等方法，研究了尼古丁与CYP2A6的结合方式、CYP2A6催化的尼古丁5'-羟基化反应的机理以及5'-羟基化反应的立体选择性。结果表明，虽然尼古丁在溶液中主要以质子化的形式存在，但是它以非质子化的形式(SRt和SRc)结合到CYP2A6的活性中心。在CYP2A6-尼古丁复合物中，尼古丁吡啶环的N与CYP2A6的Asn297形成氢键，吡啶环与周围的苯丙氨酸存在π-π相互作用，吡咯烷环的trans-5'-H或者cis-5'-H指向CYP2A6的活性中心Cpd I的O，从而利于5'-羟基化反应的发生。
研究发现，CYP2A6催化的尼古丁5'-羟基化反应由两步组成：第一步是尼古丁的5'-H传递到Cpd I的O上（氢传递）；第二步是OH再与尼古丁的5'-位置的C结合，形成5'-羟基尼古丁（氧反弹）。其中，氢传递是决速步骤。计算结果显示，CYP2A6催化的尼古丁5'-羟基化反应容易发生在trans-5'-H上，反应的立体选择性为97%，与实验报导的立体选择性相符。（生物谷Bioon.com）
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American Chemical Society DOI: 10.1021/ja111657j
Catalytic Mechanism of Cytochrome P450 for 5′-Hydroxylation of Nicotine: Fundamental Reaction Pathways and Stereoselectivity
Dongmei Li, Xiaoqin Huang, Keli Han, and Chang-Guo Zhan
A series of computational methods were used to study how cytochrome P450 2A6 (CYP2A6) interacts with (S)-(-)-nicotine, demonstrating that the dominant molecular species of (S)-(-)-nicotine in CYP2A6 active site exists in the free base state (with two conformations, SRt and SRc), despite the fact that the protonated state is dominant for the free ligand in solution. The computational results reveal that the dominant pathway of nicotine metabolism in CYP2A6 is through nicotine free base oxidation. Further, first-principles quantum mechanical/molecular mechanical free energy (QM/MM-FE) calculations were carried out to uncover the detailed reaction pathways for the CYP2A6-catalyzed nicotine 5′-hydroxylation reaction. In the determined CYP2A6–(S)-(-)-nicotine binding structures, the oxygen of Compound I (Cpd I) can abstract a hydrogen from either the trans-5′- or the cis-5′-position of (S)-(-)-nicotine. CYP2A6-catalyzed (S)-(-)-nicotine 5′-hydroxylation consists of two reaction steps, that is, the hydrogen transfer from the 5′-position of (S)-(-)-nicotine to the oxygen of Cpd I (the H-transfer step), followed by the recombination of the (S)-(-)-nicotine moiety with the iron-bound hydroxyl group to generate the 5′-hydroxynicotine product (the O-rebound step). The H-transfer step is rate-determining. The 5′-hydroxylation proceeds mainly with the stereoselective loss of the trans-5′-hydrogen, that is, the 5′-hydrogen trans to the pyridine ring. The calculated overall stereoselectivity of 97% favoring the trans-5′-hydroxylation is close to the observed stereoselectivity of 89–94%. This is the first time it has been demonstrated that a CYP substrate exists dominantly in one protonation state (cationic species) in solution, but uses its less-favorable protonation state (neutral free base) to perform the enzymatic reaction.
