Characterizing the membrane-bound state of cytochrome P450 3A4: structure, depth of insertion, and orientation.
Characterizing the membrane-bound state of cytochrome P450 3A4: structure, depth of insertion, and orientation.
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DOI：
10.1021/ja4003525
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发表时间：
2013-06-12
影响因子：
15
通讯作者：
Tajkhorshid, Emad
中科院分区：
文献类型：
--
作者：
Baylon, Javier L.;Lenov, Ivan L.;Sligar, Stephen G.;Tajkhorshid, Emad
细胞色素P450 3A4（CYP3A4）是人类P450家族中最丰富的膜相关亚型，负责体内50%以上代谢药物的生物转化。尽管CYP3A4有大量的晶体学结构，但在原子水平上没有其膜结合态的结构信息。为了表征CYP3A4的结合、插入深度、膜取向和脂质相互作用，我们在本研究中采用了实验和模拟相结合的方法。利用一种新的膜表示，高移动的膜模拟（HMMM），增强脂质的流动性和动力学，我们已经能够捕捉到自发的结合和插入的球状域的酶到膜中的多个独立的，无偏的模拟。尽管不同的初始方向和位置的蛋白质在溶液中，所有的模拟收敛到相同的膜结合的配置方面的膜插入的深度和膜表面上的酶的方向。同时，使用对与Nanodisc膜结合的CYP3A4进行的线性二色性测量来表征酶在其膜结合形式下的实验取向。实验测得的血红素倾斜角与模拟结果的膜结合结构的计算结果非常一致，从而验证了所开发的模型的有效性。CYP3A4中球状结构域的膜结合（似乎与全长酶的跨膜螺旋的存在无关）显著重塑了膜界面处的蛋白质，导致与通向酶活性位点的通道相关的构象变化。
Cytochrome P450 3A4 (CYP3A4) is the most abundant membrane-associated isoform of the P450 family in humans and is responsible for biotransformation of more than 50% of drugs metabolized in the body. Despite the large number of crystallographic structures available for CYP3A4, no structural information for its membrane-bound state at an atomic level is available. In order to characterize binding, depth of insertion, membrane orientation, and lipid interactions of CYP3A4, we have employed a combined experimental and simulation approach in this study. Taking advantage of a novel membrane representation, highly mobile membrane mimetic (HMMM), with enhanced lipid mobility and dynamics, we have been able to capture spontaneous binding and insertion of the globular domain of the enzyme into the membrane in multiple independent, unbiased simulations. Despite different initial orientations and positions of the protein in solution, all the simulations converged into the same membrane-bound configuration with regard to both the depth of membrane insertion and the orientation of the enzyme on the surface of the membrane. In tandem, linear dichroism measurements performed on CYP3A4 bound to Nanodisc membranes were used to characterize the orientation of the enzyme in its membrane-bound form experimentally. The heme tilt angles measured experimentally are in close agreement with those calculated for the membrane-bound structures resulted from the simulations, thereby verifying the validity of the developed model. Membrane binding of the globular domain in CYP3A4, which appears to be independent of the presence of the transmembrane helix of the full-length enzyme, significantly reshapes the protein at the membrane interface, causing conformational changes relevant to access tunnels leading to the active site of the enzyme.
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影响因子：
--
作者：
CROPEK, DM;BOHN, PW
通讯作者：
BOHN, PW
影响因子：
2.4
作者：
Kucerka, N;Tristram-Nagle, S;Nagle, JF
通讯作者：
Nagle, JF
DOI：
10.1021/jp109631y
发表时间：
2011-06-02
期刊：
The journal of physical chemistry. B
影响因子：
--
作者：
Arcario MJ;Ohkubo YZ;Tajkhorshid E
通讯作者：
Tajkhorshid E
影响因子：
4.4
作者：
FELLER, SE;ZHANG, YH;BROOKS, BR
通讯作者：
BROOKS, BR
影响因子：
4.4
作者：
JORGENSEN, WL;CHANDRASEKHAR, J;KLEIN, ML
通讯作者：
KLEIN, ML