Novel biophysical tools allow the structural dynamics of proteins, and the regulation of such dynamics by binding partners, to be explored in unprecedented detail. Although this has provided critical insights into protein function, the means by which structural dynamics direct protein evolution remains poorly understood. Here, we investigated how proteins with a bilobed structure, composed of two related domains from the type-II periplasmic binding protein domain family, have undergone divergent evolution leading to modification of their structural dynamics and function. We performed a structural analysis of ~600 bilobed proteins with a common primordial structural core, which we complemented with biophysical studies to explore the structural dynamics of selected examples by single-molecule Foerster resonance energy transfer and Hydrogen-Deuterium exchange mass spectrometry. We show that evolutionary modifications of the structural core, largely at its termini, enables distinct structural dynamics, allowing the diversification of these proteins into transcription factors, enzymes, and extra-cytoplasmic transport-related proteins. Structural embellishments of the core created new interdomain interactions that stabilized structural states, reshaping the active site geometry, and ultimately, altered substrate specificity. Our findings reveal an as yet unrecognized mechanism for the emergence of functional promiscuity during long periods of protein evolution and are applicable to a large number of domain architectures.

中文翻译：

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双叶蛋白结构动力学的演变

新颖的生物物理工具允许对蛋白质的结构动力学以及结合伴侣对这种动力学的调控进行前所未有的详细研究。尽管这提供了对蛋白质功能的关键见解，但对于结构动力学指导蛋白质进化的方法仍然知之甚少。在这里，我们调查了具有双叶结构的蛋白质（由来自II型周质结合蛋白结构域家族的两个相关结构域组成）如何经历发散进化，从而导致其结构动力学和功能的修饰。我们对大约600个具有共同原始结构核心的双叶蛋白进行了结构分析，我们通过生物分子研究相辅相成，通过单分子Foerster共振能量转移和氢-氘交换质谱法研究了选定实例的结构动力学。我们表明，结构核心的进化修饰，主要是在其末端，使独特的结构动力学成为可能，从而使这些蛋白质多样化成转录因子，酶和胞质外运输相关蛋白质。核心的结构修饰创造了新的域间相互作用，从而稳定了结构状态，重塑了活性位点的几何形状，并最终改变了底物特异性。