Pyruvate dehydrogenase kinase-3 (PDK3) plays important role in the glucose metabolism and is associated with cancer progression, and thus being considered as an attractive target for cancer therapy. In this study, we employed spectroscopic techniques to study the structural and conformational changes in the PDK3 at varying pH conditions ranging from pH 2.0 to 12.0. UV/Vis, fluorescence and circular dichroism spectroscopic measurements revealed that PDK3 maintains its native-like structure (both secondary and tertiary) in the alkaline conditions (pH 7.0-12.0). However, a significant loss in the structure was observed under acidic conditions (pH 2.0-6.0). The propensity of aggregate formation at pH 4.0 was estimated by thioflavin T fluorescence measurements. To further complement structural data, kinase activity assay was performed, and maximum activity of PDK3 was observed at pH 7.0-8.0 range; whereas, its activity was lost under acidic pH. To further see conformational changes at atomistic level we have performed all-atom molecular dynamics at different pH conditions for 150 ns. A well defined correlation was observed between experimental and computational studies. This work highlights the significance of structural dependence of pH for wide implications in protein-protein interaction, biological function and drug design procedures.

中文翻译：

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pH对丙酮酸脱氢酶激酶3结构和功能的影响：结合光谱和MD模拟研究。

丙酮酸脱氢酶激酶3（PDK3）在葡萄糖代谢中起重要作用，并且与癌症的发展有关，因此被认为是癌症治疗的诱人靶标。在这项研究中，我们采用光谱技术研究了在pH 2.0至12.0范围内的各种pH条件下PDK3的结构和构象变化。UV / Vis，荧光和圆二色性光谱测量表明，PDK3在碱性条件（pH 7.0-12.0）下保持其天然结构（二级和三级）。然而，在酸性条件（pH 2.0-6.0）下观察到结构的重大损失。通过硫黄素T荧光测量估计在pH 4.0下聚集体形成的倾向。为了进一步补充结构数据，我们进行了激酶活性测定，在pH 7.0-8.0范围内观察到PDK3的最大活性。而在酸性pH下其活性丧失。为了进一步查看原子级的构象变化，我们在不同的pH条件下进行了150 ns的全原子分子动力学研究。在实验研究和计算研究之间观察到明确定义的相关性。这项工作强调了pH的结构依赖性对于蛋白质-蛋白质相互作用，生物学功能和药物设计程序的广泛影响的重要性。