Yousuf Mohd, Shamsi Anas, Anjum Farah, Shafie Alaa, Islam Asimul, Haque Qazi Mohd Rizwanul, Elasbali Abdelbaset Mohamed, Yadav Dharmendra Kumar, Hassan Md Imtaiyaz
Department of Biosciences, Jamia Millia Islamia, New Delhi, India.
Centre for Interdisciplinary Research in Basic Sciences, Jamia Millia Islamia, New Delhi, India.
PLoS One. 2022 Feb 11;17(2):e0263693. doi: 10.1371/journal.pone.0263693. eCollection 2022.
Cyclin-dependent kinase 6 (CDK6) is an important protein kinase that regulates cell growth, development, cell metabolism, inflammation, and apoptosis. Its overexpression is associated with reprogramming glucose metabolism through alternative pathways and apoptosis, which ultimately plays a significant role in cancer development. In the present study, we have investigated the structural and conformational changes in CDK6 at varying pH employing a multi-spectroscopic approach. Circular dichroism (CD) spectroscopy revealed at extremely acidic conditions (pH 2.0-4.0), the secondary structure of CDK6 got significantly disrupted, leading to aggregates formation. These aggregates were further characterized by employing Thioflavin T (ThT) fluorescence. No significant secondary structural changes were observed over the alkaline pH range (pH 7.0-11.0). Further, fluorescence and UV spectroscopy revealed that the tertiary structure of CDK6 was disrupted under extremely acidic conditions, with slight alteration occurring in mild acidic conditions. The tertiary structure remains intact over the entire alkaline range. Additionally, enzyme assay provided an insight into the functional aspect of CDK at varying pH; CDK6 activity was optimal in the pH range of 7.0-8.0. This study will provide a platform that provides newer insights into the pH-dependent dynamics and functional behavior of CDK6 in different CDK6 directed diseased conditions, viz. different types of cancers where changes in pH contribute to cancer development.
细胞周期蛋白依赖性激酶6（CDK6）是一种重要的蛋白激酶，可调节细胞生长、发育、细胞代谢、炎症和细胞凋亡。其过表达与通过替代途径的葡萄糖代谢重编程和细胞凋亡相关，这最终在癌症发展中起重要作用。在本研究中，我们采用多光谱方法研究了不同pH值下CDK6的结构和构象变化。圆二色性（CD）光谱显示，在极端酸性条件下（pH 2.0 - 4.0），CDK6的二级结构明显破坏，导致聚集体形成。通过硫黄素T（ThT）荧光对这些聚集体进行了进一步表征。在碱性pH范围（pH 7.0 - 11.0）内未观察到明显的二级结构变化。此外，荧光和紫外光谱显示，在极端酸性条件下CDK6的三级结构被破坏，在轻度酸性条件下有轻微改变。在整个碱性范围内三级结构保持完整。此外，酶活性测定提供了不同pH值下CDK功能方面的见解；CDK6活性在pH 7.0 - 8.0范围内最佳。本研究将提供一个平台，为不同CDK6相关疾病状态下，即不同类型癌症中pH变化促进癌症发展的情况下，CDK6的pH依赖性动力学和功能行为提供新的见解。
