REQUIMTE, Departamento de Química e Bioquímica, Faculdade de Ciências, Universidade do Porto , s/n, Rua do Campo Alegre, 4169-007 Porto, Portugal.
J Phys Chem B. 2018 Jan 11;122(1):77-85. doi: 10.1021/acs.jpcb.7b09645. Epub 2018 Jan 2.
Human fatty acid synthase (hFAS) is a megasynthase whose main function is de novo biosynthesis of saturated fatty acids. Interest has been drawn to this enzyme beyond its physiological role due to the association between high levels of hFAS and clinical conditions such as obesity, diabetes, and cancer. Thus, it has become an undeniably attractive pharmacological target. Until now, no crystal structure of the complete hFAS is available, hindering attempts to fully understand this protein. Using homology modeling, we built a model of the entire megasynthase, encompassing all of its domains, including the acyl carrier protein (ACP) and thioesterase (TE) mobile domains absent in the crystal structure of mammalian fatty acid synthase (FAS). On a second stage, we used data-driven protein-protein docking between the substrate shuttling domain ACP and every catalytic domain in the protein. We also propose sets of amino acids at the interface of each domain that we believe are important to favor the interaction between ACP and each domain of hFAS. After inspection, we validated each complex between ACP and MAT/KS/KR/DH/ER domains through classical molecular dynamics simulations and RMSd analysis. Additionally, we mapped the interactions between the residues at the active site of each catalytic domain and its intermediaries. In every docking, we ensured that the distance between catalytic residues and the intermediaries was maintained. Until now, there was not a complete 3D model of this megasynthase. This study is the first to present a homology model for the whole hFAS, including its two mobile domains and possible poses of ACP throughout the cycle of fatty acid biosynthesis, thus mapping obligatory checkpoints in its trajectory. Hence, we believe that these structural insights will allow for future studies of the catalytic mechanism of the overall hFAS.
人脂肪酸合酶（hFAS）是一种超大合酶，其主要功能是从头合成饱和脂肪酸。由于 hFAS 水平与肥胖症、糖尿病和癌症等临床状况之间存在关联，人们对这种酶的兴趣超出了其生理作用。因此，它已成为一个不可否认的有吸引力的药理靶点。到目前为止，还没有完整的 hFAS 晶体结构，这阻碍了对该蛋白的充分理解。我们使用同源建模构建了整个超大合酶的模型，包括其所有结构域，包括晶体结构中缺失的酰基载体蛋白（ACP）和硫酯酶（TE）可移动结构域。在第二阶段，我们使用底物穿梭结构域 ACP 与蛋白质中每个催化结构域之间的数据驱动蛋白-蛋白对接。我们还提出了每个结构域界面的氨基酸集合，我们认为这些氨基酸对于促进 ACP 与 hFAS 的每个结构域之间的相互作用很重要。经过检查，我们通过经典分子动力学模拟和 RMSd 分析验证了 ACP 与 MAT/KS/KR/DH/ER 结构域之间的每个复合物。此外，我们还绘制了每个催化结构域的活性位点处的残基与中间体之间的相互作用。在每次对接中，我们都确保了催化残基与中间体之间的距离保持不变。到目前为止，还没有这种超大合酶的完整 3D 模型。这项研究首次提出了 hFAS 的同源模型，包括其两个可移动结构域和 ACP 在脂肪酸生物合成循环中的可能构象，从而绘制了其轨迹中的强制性检查点。因此，我们相信这些结构上的见解将为整个 hFAS 的催化机制的未来研究提供参考。
