Center for Molecular Biophysics, University of Tennessee/Oak Ridge National Lab, Oak Ridge, TN 37830, United States of America.
Department of Medicine, Microbiology, Immunology and Biochemistry University of Tennessee Health Science Center, 19 S, Manassas, St. Memphis, TN 38163, United States of America.
Biochim Biophys Acta Gen Subj. 2021 Mar;1865(3):129809. doi: 10.1016/j.bbagen.2020.129809. Epub 2020 Dec 17.
The carboxylation status of Osteocalcin (Ocn) not only influences formation and structure in bones but also has important endocrine functions affecting energy metabolism and expenditure. In this study, the role of γ-carboxylation of the glutamate residues in the structure-dynamics-function relationship in Ocn is investigated.
Three forms of Ocn, differentially carboxylated at the Glu-17, 21 and 24 residues, along with a mutated form of Ocn carrying Glu/Ala mutations, are modeled and simulated using molecular dynamics (MD) simulation in the presence of calcium ions.
Characterization of the global conformational dynamics of Ocn, described in terms of the orientational variations within its 3-helical domain, highlights large structural variations in the non-carboxylated osteocalcin (nOcn). The bi-carboxylated Ocn (bOcn) and tri-carboxylated (tOcn) species, in contrast, display relatively rigid tertiary structures, with the dynamics of most regions strongly correlated. Radial distribution functions calculated for both bOcn and tOcn show long-range ordering of the calcium ion distribution around the carboxylated glutamate (γGlu) residues, likely playing an important role in promoting stability of these Ocns. Additionally, the same calcium ions are observed to coordinate with neighboring γGlu, better shielding their negative charges and in turn stabilizing these systems more than do the singly coordinating calcium ions observed in the case of nOcn. bOcn is also found to exhibit a more helical C-terminal structure, that has been shown to activate its cellular receptor GPRC6A, highlighting the allosteric role of Ocn carboxylation in modulating the stability and binding potential of the active C-terminal.
The carboxylation status of Ocn as well and its calcium coordination appear to have a direct influence on Ocn structure and dynamics, possibly leading to the known differences in Ocn biological function.
Modification of Ocn sequence or its carboxylation state may provide the blueprint for developing high-affinity peptides targeting its cellular receptor GPRC6A, with therapeutic potential for treatment of metabolic disorders.
骨钙素（Ocn）的羧化状态不仅影响骨骼的形成和结构，而且具有重要的内分泌功能，影响能量代谢和消耗。在这项研究中，研究了谷氨酸残基的γ-羧化在 Ocn 的结构-动力学-功能关系中的作用。
使用分子动力学（MD）模拟在钙离子存在的情况下对 Ocn 的 Glu-17、21 和 24 残基差异羧化的三种形式以及携带 Glu/Ala 突变的 Ocn 突变形式进行建模和模拟。
Ocn 的整体构象动力学的特征描述为其 3 螺旋结构域内的取向变化，突出了非羧化骨钙素（nOcn）的大结构变化。相比之下，双羧化 Ocn（bOcn）和三羧化（tOcn）物种显示出相对刚性的三级结构，大多数区域的动力学强烈相关。为 bOcn 和 tOcn 计算的径向分布函数显示出羧化谷氨酸（γGlu）残基周围钙离子分布的长程有序性，这可能在促进这些 Ocn 的稳定性方面发挥重要作用。此外，观察到相同的钙离子与相邻的 γGlu 配位，更好地屏蔽了它们的负电荷，从而比在 nOcn 情况下观察到的仅配位钙离子更稳定这些系统。还发现 bOcn 表现出更螺旋的 C 末端结构，已显示激活其细胞受体 GPRC6A，突出了 Ocn 羧化在调节活性 C 末端稳定性和结合潜力方面的变构作用。
Ocn 的羧化状态及其钙配位似乎直接影响 Ocn 的结构和动力学，可能导致 Ocn 生物学功能的已知差异。
Ocn 序列的修饰或其羧化状态可能为开发针对其细胞受体 GPRC6A 的高亲和力肽提供蓝图，具有治疗代谢紊乱的治疗潜力。
