Azhar Nur Alya Amirah, Chua Yung-An, Nawawi Hapizah, Jusoh Siti Azma
Faculty of Pharmacy, Universiti Teknologi MARA, Puncak Alam Campus, 42300, Bandar Puncak Alam, Selangor, Malaysia.
Institute of Pathology, Laboratory and Forensic Medicine (I-PPerForm), Universiti Teknologi MARA, 47000, Sungai Buloh, Selangor, Malaysia.
J Mol Model. 2025 May 19;31(6):161. doi: 10.1007/s00894-025-06380-1.
The low-density lipoprotein receptor (LDLR) regulates cholesterol uptake by mediating the hepatic clearance of plasma low-density lipoprotein cholesterol (LDL-C). Proprotein convertase subtilisin/kexin type-9 (PCSK9) attenuates LDLR function by binding to the LDLR, leading to its lysosomal degradation and preventing the total depletion of circulating LDL-C. However, pathogenic PCSK9 variants can reduce LDLR availability, significantly increase plasma LDL-C levels. Despite this understanding, the detailed molecular mechanism of LDLR-PCSK9 interaction remains unclear due to the incomplete LDLR structure. This study uses molecular dynamics (MD) simulations to predict LDLR structural dynamics upon binding to PCSK9. Furthermore, PCSK9 variants, E498A and R499G, that were identified in Malaysian FH patients were investigated for their mutational effects. Throughout the simulations, PCSK9 remained stable, while LDLR explored a larger conformational space. The LDLR-PCSK9 wild-type (WT) complex showed minimal changes, while the LDLR-PCSK9(R499G) complex exhibited pronounced conformational rearrangement. The MM/GBSA analysis revealed that the LDLR-PCSK9(E498A) complex had the highest binding affinity (- 63.81 kcal/mol), followed by the WT complex (- 33.07 kcal/mol), and LDLR-PCSK9(R499G) (- 24.21 kcal/mol). These findings offer novel insights into the dynamic interactions between LDLR and PCSK9, highlighting the role of structural flexibility in their relationship. Further MD simulation studies with the complete LDLR structure as well as experimental validation are needed to elucidate the molecular mechanisms underlying LDLR-PCSK9-mediated cholesterol homeostasis.
The initial structure of the wild-type (WT) LDLR-PCSK9 complex was obtained from PDB ID 3P5C, and the PCSK9 mutant structures (E498A and R499G) were modeled using the SPDBV program. MD simulations for each complex-LDLR-PCSK9 WT, LDLR-PCSK9(E498A), and LDLR-PCSK9(R499G)-were conducted using the GROMACS package with the CHARMM36m force field. The simulations were performed at 310.15 K with 2-fs timesteps under the isothermal-isobaric (NPT) ensemble, with each run lasting 500 ns. Including triplicates, the total duration of MD simulation time for all complexes amounted to 3.5 μs.
低密度脂蛋白受体（LDLR）通过介导血浆低密度脂蛋白胆固醇（LDL-C）的肝脏清除来调节胆固醇摄取。前蛋白转化酶枯草杆菌蛋白酶/kexin 9型（PCSK9）通过与LDLR结合来减弱LDLR功能，导致其溶酶体降解并防止循环LDL-C完全耗尽。然而，致病性PCSK9变体可降低LDLR的可用性，显著提高血浆LDL-C水平。尽管有此认识，但由于LDLR结构不完整，LDLR-PCSK9相互作用的详细分子机制仍不清楚。本研究使用分子动力学（MD）模拟来预测LDLR与PCSK9结合后的结构动力学。此外，还研究了在马来西亚FH患者中鉴定出的PCSK9变体E498A和R499G的突变效应。在整个模拟过程中，PCSK9保持稳定，而LDLR探索了更大的构象空间。LDLR-PCSK9野生型（WT）复合物显示出最小的变化，而LDLR-PCSK9（R499G）复合物表现出明显的构象重排。MM/GBSA分析表明，LDLR-PCSK9（E498A）复合物具有最高的结合亲和力（-63.81 kcal/mol），其次是WT复合物（-33.07 kcal/mol）和LDLR-PCSK9（R499G）（-24.21 kcal/mol）。这些发现为LDLR与PCSK9之间的动态相互作用提供了新的见解，突出了结构灵活性在它们关系中的作用。需要进一步以完整的LDLR结构进行MD模拟研究以及实验验证，以阐明LDLR-PCSK9介导的胆固醇稳态的分子机制。
野生型（WT）LDLR-PCSK9复合物的初始结构来自PDB ID 3P5C，PCSK9突变体结构（E498A和R499G）使用SPDBV程序进行建模。使用带有CHARMM36m力场的GROMACS软件包对每个复合物——LDLR-PCSK9 WT、LDLR-PCSK9（E498A）和LDLR-PCSK9（R499G）——进行MD模拟。模拟在310.15 K下进行，时间步长为2 fs，处于等温等压（NPT）系综，每次运行持续500 ns。包括重复实验，所有复合物的MD模拟总时长达到3.5 μs。
