喹啉加氧酶中氧气扩散的通道分析

doi:10.7503/cjcu20170202

摘要/Abstract

摘要：

通过生物信息学分析、量化计算优化、 CAVER和MDpocket预测、随机加速分子动力学及伞状抽样动力学模拟等方法, 对喹啉加氧酶(HOD)中的氧气扩散途径进行了计算预测. 结果表明, 氧气在HOD中的反应位点包埋在蛋白内部, 而HOD中有数条可能的通道供氧气进出, 其中长度最短的通道具有最高的优先度, 不仅在随机加速动力学模拟中具有最高的氧气逸出概率, 而且伞状抽样方法计算得到的自由能也最低. 此通道的内端位于底物Re面的氧气结合位点, 较好地解释了HOD的相关实验数据.

关键词: 喹啉加氧酶, 氧气扩散, 随机加速分子动力学, 伞状抽样

Abstract:

Oxygen diffusion pathways of quinoline oxygenase(HOD) were investigated by bioinformatics analysis, quantum chemical optimization, CAVER and MD pocket prediction, random accelerated molecular dynamics simulations and umbrella sampling. The results show that the pre-reaction site of oxygen does locate inside the active pocket, and there exists several possible oxygen tunnels in HOD, in which the one with shortest length has the highest priority. Random accelerated molecular dynamics proved that oxygen has the highest probability to exit from this tunnel, and umbrella sampling further verified its free energy for oxygen passing by is the lowest. The tunnel ended at the binding site of oxygen located on the Re-face of the substrate, which is consistent with previous studies. These results better illuminate the kinetics and the stereo-selectivity of the catalytic reaction in HOD, and also provide theoretical guidance to further experimentally improve the efficiency of oxygen usage by enzyme.

Key words: Quinoline oxygenase, Oxygendiffusion, Random accelerated molecular dynamics, Umbrella sampling

中图分类号:

O641

TrendMD:

周超, 石婷, 赵一雷, 王晓雷. 喹啉加氧酶中氧气扩散的通道分析. 高等学校化学学报, 2017, 38(10): 1813.

ZHOU Chao, SHI Ting, ZHAO Yilei, WANG Xiaolei. Theoretical Analysis of Oxygen Diffusion in the Micro-channels of Quinoline Oxygenase^†. Chem. J. Chinese Universities, 2017, 38(10): 1813.

图/表 11

Scheme 1 Degradation reaction catalyzed by HOD and its analogue enzymes, QDO and QD(A) and catalytic cycle of HOD(B)

Fig.1 Location of Cl- in the crystal structure of HODCl- is buried deeper inside the active pocket, beneath the substrate QND, which is also the putative binding site of O2. Re face and Si face of QND are labelled according to the carbonyl group.

Fig.2 Sequence alignment of HOD, QDO and QD

Fig.3 Proportion of the two pre-reaction oxygen orientations(red and purple) in total 26 optimizations(A) and their 3D structure in the active site of HOD(B)

Fig.4 25 Tunnels predicted by CAVER(A) and top 5 tunnels in rank score(B)

Table 1 25 Predicted oxygen tunnels and their bottleneck radius, length, curvature and priority

ID	Bottleneck radius/nm	Length/nm	Curvature	Priority	ID	Bottleneck radius/nm	Length/nm	Curvature	Priority
1	0.095	1.674	1.38	0.42	14	0.074	3.365	1.35	0.07
2	0.070	2.213	1.52	0.30	15	0.077	4.507	3.28	0.06
3	0.079	2.624	1.34	0.18	16	0.061	3.293	2.09	0.06
4	0.070	2.419	1.36	0.14	17	0.067	3.664	1.52	0.05
5	0.064	2.705	1.56	0.14	18	0.066	3.852	2.32	0.05
6	0.062	2.054	1.70	0.13	19	0.063	3.437	2.21	0.05
7	0.077	3.221	1.42	0.13	20	0.064	3.517	2.30	0.05
8	0.074	2.875	1.41	0.12	21	0.064	4.117	1.73	0.04
9	0.077	3.380	3.34	0.12	22	0.062	3.565	2.22	0.04
10	0.064	2.353	1.31	0.12	23	0.064	3.962	1.58	0.04
11	0.064	2.712	1.38	0.11	24	0.061	4.564	2.44	0.04
12	0.064	2.524	1.55	0.09	25	0.064	4.137	1.86	0.02
13	0.066	2.970	1.73	0.09

Fig.5 4 Pockets in HOD predicted by MDpocket(A) and the positional relationship between tunnel 1 and pocket 2(B)

Fig.6 Active site in pocket 1(A), amino acid compositions of pocket 2(B), pocket 3(C) and pocket 4(D)

Table 2 Results of the RAMD simulation

Tunnel	Time step			Sum
Tunnel	10	30	50	Sum
1	9	8	11	28
2	2	1	0	3
3	1	2	0	3
4	1	1	0	2
5	1	3	0	4
8	5	4	1	10
Others	1	1	8	10
Sum	20	20	20	60

Fig.7 Sampling distribution of the three tunnels along the reaction coordinate(A) Tunnel 1; (B) tunnel 2; (C) tunnel 8.

Fig.8 Potential mean force calculated by WHAM program

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