Protein conformational relaxation and ligand migration in myoglobin: a nanosecond to millisecond molecular movie from time-resolved Laue X-ray diffraction.
Protein conformational relaxation and ligand migration in myoglobin: a nanosecond to millisecond molecular movie from time-resolved Laue X-ray diffraction.
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DOI：
10.1021/bi010715u
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发表时间：
2001-10
期刊：
影响因子：
2.9
通讯作者：
V. Šrajer;Z. Ren;T. Teng;Marius Schmidt;T. Ursby;D. Bourgeois;C. Pradervand;W. Schildkamp;M. Wulff;K. Moffat
中科院分区：
文献类型：
--
作者：
V. Šrajer;Z. Ren;T. Teng;Marius Schmidt;T. Ursby;D. Bourgeois;C. Pradervand;W. Schildkamp;M. Wulff;K. Moffat
时间分辨劳厄X-射线衍射技术已被用来探讨蛋白质弛豫和配体迁移在室温下光解的单晶碳单氧肌红蛋白。CO配位体的光解由7.5纳秒的激光脉冲，和随后的结构变化探测由150 ps或1微秒的X-射线脉冲在14激光/X-射线延迟时间，范围从1 ns到1.9毫秒。非常快的血红素和蛋白质松弛涉及的E和F螺旋是显而易见的，从数据在1 ns的时间延迟。光解离的CO分子在两个位置被检测到：在远端口袋对接位点和在近端口袋中的BAR 1结合位点。CO的主要，远端网站的人口峰值在1 ns的时间延迟和衰减到一半的峰值在70 ns。第二个近端对接位点在约100 ns时达到其最高占用率20%，半衰期约为10 μ s。在大约100纳秒，所有的CO分子占内的蛋白质：在这两个对接位点之一或绑定到血红素。此后，CO分子迁移到溶剂中，在双分子过程中与脱氧肌红蛋白重新结合，二级速率系数为4.5 × 10（5）M（-1）s（-1）。我们的研究结果还表明，小至0.2 A和人口的CO对接网站的10%的结构变化可以检测到的时间分辨X-射线衍射。
A time-resolved Laue X-ray diffraction technique has been used to explore protein relaxation and ligand migration at room temperature following photolysis of a single crystal of carbon monoxymyoglobin. The CO ligand is photodissociated by a 7.5 ns laser pulse, and the subsequent structural changes are probed by 150 ps or 1 micros X-ray pulses at 14 laser/X-ray delay times, ranging from 1 ns to 1.9 ms. Very fast heme and protein relaxation involving the E and F helices is evident from the data at a 1 ns time delay. The photodissociated CO molecules are detected at two locations: at a distal pocket docking site and at the Xe 1 binding site in the proximal pocket. The population by CO of the primary, distal site peaks at a 1 ns time delay and decays to half the peak value in 70 ns. The secondary, proximal docking site reaches its highest occupancy of 20% at approximately 100 ns and has a half-life of approximately 10 micros. At approximately 100 ns, all CO molecules are accounted for within the protein: in one of these two docking sites or bound to the heme. Thereafter, the CO molecules migrate to the solvent from which they rebind to deoxymyoglobin in a bimolecular process with a second-order rate coefficient of 4.5 x 10(5) M(-1) s(-1). Our results also demonstrate that structural changes as small as 0.2 A and populations of CO docking sites of 10% can be detected by time-resolved X-ray diffraction.