Northrup S H, Wensel T G, Meares C F, Wendoloski J J, Matthew J B
Chemistry Department, Tennessee Technological University, Cookeville 38505.
Proc Natl Acad Sci U S A. 1990 Dec;87(23):9503-7. doi: 10.1073/pnas.87.23.9503.
Energy transfer in the "rapid diffusion" limit from electronically excited terbium(III) chelates in three different charge states to horse heart ferricytochrome c was measured as a function of ionic strength. Theoretical rate constants calculated by numerical integration of the Forster integral (containing the Poisson-Boltzmann-generated protein electrostatic potential) were compared with the experimental data to evaluate the accuracy of protein electrostatic field calculations at the protein/solvent interface. Two dielectric formalisms were used: a simple coulombic/Debye-Hückel procedure and a finite difference method [Warwicker, J. & Watson, H. C. (1982) J. Mol. Biol. 157, 671-679] that accounts for the low-dielectric protein interior and the irregular protein/solvent boundary. Good agreement with experiment was obtained and the ionic-strength dependence of the reaction was successfully reproduced. The sensitivity of theoretical rate constants to the choices of effective donor sphere size and the energy transfer distance criterion was analyzed. Electrostatic potential and rate-constant calculations were carried out on sets of structures collected along two molecular dynamics trajectories of cytochrome c. Protein conformational fluctuations were shown to produce large variations in the calculated energy transfer rate constant. We conclude that protein fluctuations and the resulting transient structures can play significant roles in biological or catalytic activities that are not apparent from examination of a static structure. For calculating protein electrostatics, large-scale low-frequency conformational fluctuations, such as charged side-chain reorientation, are established to be as important as the computational method for incorporating dielectric boundary effects.
测量了在“快速扩散”极限下，处于三种不同电荷状态的电子激发铽（III）螯合物向马心铁细胞色素c的能量转移随离子强度的变化。将通过福斯特积分（包含泊松-玻尔兹曼生成的蛋白质静电势）的数值积分计算得到的理论速率常数与实验数据进行比较，以评估蛋白质/溶剂界面处蛋白质静电场计算的准确性。使用了两种介电形式：一种简单的库仑/德拜-休克尔方法和一种有限差分方法[沃里克，J. & 沃森，H. C.（1982）《分子生物学杂志》157，671 - 679]，该方法考虑了低介电常数的蛋白质内部和不规则的蛋白质/溶剂边界。实验结果与理论计算取得了良好的一致性，成功再现了反应的离子强度依赖性。分析了理论速率常数对有效供体球尺寸选择和能量转移距离标准的敏感性。对沿着细胞色素c的两条分子动力学轨迹收集的结构集进行了静电势和速率常数计算。结果表明，蛋白质构象波动会导致计算出的能量转移速率常数产生很大变化。我们得出结论，蛋白质波动以及由此产生的瞬态结构在生物或催化活性中可能发挥重要作用，而这些作用从静态结构检查中并不明显。对于计算蛋白质静电，大规模低频构象波动，如带电侧链重新定向，与纳入介电边界效应的计算方法同样重要。
