Department of Physics, Colorado School of Mines, 1500 Illinois Street, Golden, CO, 80401, USA.
Warwick Medical School, University of Warwick, Coventry, CV4 7AL, UK.
Sci Rep. 2020 Nov 26;10(1):20615. doi: 10.1038/s41598-020-77699-3.
The roles of protein conformational dynamics and allostery in function are well-known. However, the roles that interdomain dynamics have in function are not entirely understood. We used matrix metalloprotease-1 (MMP1) as a model system to study the relationship between interdomain dynamics and activity because MMP1 has diverse substrates. Here we focus on fibrin, the primary component of a blood clot. Water-soluble fibrinogen, following cleavage by thrombin, self-polymerize to form water-insoluble fibrin. We studied the interdomain dynamics of MMP1 on fibrin without crosslinks using single-molecule Forster Resonance Energy Transfer (smFRET). We observed that the distance between the catalytic and hemopexin domains of MMP1 increases or decreases as the MMP1 activity increases or decreases, respectively. We modulated the activity using (1) an active site mutant (E219Q) of MMP1, (2) MMP9, another member of the MMP family that increases the activity of MMP1, and (3) tetracycline, an inhibitor of MMP1. We fitted the histograms of smFRET values to a sum of two Gaussians and the autocorrelations to an exponential and power law. We modeled the dynamics as a two-state Poisson process and calculated the kinetic rates from the histograms and autocorrelations. Activity-dependent interdomain dynamics may enable allosteric control of the MMP1 function.
蛋白质构象动力学和变构在功能中的作用是众所周知的。然而，结构域间动力学在功能中的作用还不完全清楚。我们使用基质金属蛋白酶-1（MMP1）作为模型系统来研究结构域间动力学与活性之间的关系，因为 MMP1 有多种底物。在这里，我们专注于纤维蛋白，这是血凝块的主要成分。在凝血酶切割后，水溶性的纤维蛋白原自聚合形成不溶于水的纤维蛋白。我们使用单分子Förster 共振能量转移（smFRET）研究了没有交联的 MMP1 在纤维蛋白上的结构域间动力学。我们观察到，随着 MMP1 活性的增加或降低，MMP1 的催化结构域和血红素结合结构域之间的距离分别增加或减少。我们使用（1）MMP1 的活性位点突变体（E219Q）、（2）MMP9（MMP 家族的另一个成员，可增加 MMP1 的活性）和（3）四环素（MMP1 的抑制剂）来调节活性。我们将 smFRET 值的直方图拟合为两个高斯的和，自相关拟合为指数和幂律。我们将动力学建模为二态泊松过程，并从直方图和自相关中计算出动力学速率。活性依赖性结构域间动力学可能能够实现 MMP1 功能的变构控制。
