靶向膜型1基质金属蛋白酶反义肽的虚拟筛选与分子模拟

Virtual screening and molecular simulations of antisense peptides targeting MT1-MMP

摘要:

膜型1基质金属蛋白酶 (Membrane type-1 matrix metalloproteinase, MT1-MMP, MMP14) 在肿瘤的发生发展及转移中起着重要作用，是肿瘤潜在理想的药物靶标。为了获得MT1-MMP结合肽，我们首先采用生物信息学方法分析MMPs序列，获得MT1-MMP差异大且特异的序列。以此为正义肽靶标，设计反义肽库，然后通过分子对接、分子动力学模拟以及体外细胞实验等多种方法，进行靶向MT1-MMP反义肽的筛选与活性研究。多序列比对确定了位于MT1-MMP环区的特异序列AYIREGHE (简称MT1-loop)，并构建1 536条反义肽。经两轮虚拟筛选，选取打分位于前五的反义肽用于后续研究。该五条反义肽与MT1-MMP存在较强的相互作用且能很好地对接于正义肽区域。进一步分析其与MMPs其他家族成员 (MMP1-3, MMP7-13, MMP14 HPX, MMP16) 的亲和力，发现反义肽FVTFPYIR对MT1-MMP具有更强的特异性。分子动力学模拟表明，反义肽FVTFPYIR可能是通过影响受体MT1-MMP的构象稳定性，进而影响其功能活性。体外细胞实验初步确定反义肽FVTFPYIR可选择性地抑制表达MT1-MMP的人成骨肉瘤细胞MG63和乳腺癌MDA-MB-231细胞的增殖。本研究为抗肿瘤反义肽先导药物的研发提供了一种新的思路与途径。

Abstract:

Membrane type-1 matrix metalloproteinase (MT1-MMP or MMP14) plays the pivotal role in tumor development and metastasis, so it is a promising drug target in malignancy. To acquire MT1-MMP specific binding peptides, we first analyzed MMPs sequences to find the divergent and specific sequence of MT1-MMP by bioinformatics approach, then set the specific sequence as the sense peptide target and designed antisense peptide library. Finally, by means of molecular docking, molecular dynamics simulation and in vitro cell assays, we screened the antisense peptide library against MT1-MMP and further studied the obtained specific peptides. Here, we identified the divergent and specific sequence of AYIREGHE (Named MT1-loop) located in MT1-MMP loop by multiple sequence alignment and established the antisense peptides library with capacity of 1 536 sequences. After two rounds of virtual screening, we obtained five antisense peptides with Rerankscores in the top for further screening. They all interacted with MT1-MMP, and docked well at the active site composed of MT1-loop sequence. Analysis of the affinities of these five antisense peptides to other MMPs (MMP1-3, MMP7-13, MMP14 HPX, MMP16) revealed that the peptide FVTFPYIR was more specific to MT1-MMP. Molecular dynamics simulation showed that the peptide FVTFPYIR might affect the stability of MT1-MMP and thus have effects on its activities. Meanwhile, the peptide FVTFPYIR could specifically inhibit the growth of MG63 and MDA-MB-231 tumor cells both of which expressed MT1-MMP. The work provides a new insight and way for the development of antitumor lead peptides targeting MT1-MMP.