Department of Chemistry, Graduate School of Sciences, The University of Tokyo, Tokyo 113-0033, Japan.
National Center for Advancing Translational Sciences, National Institutes of Health, Rockville, Maryland 20850, USA.
Nat Commun. 2017 Apr 3;8:14932. doi: 10.1038/ncomms14932.
Glycolytic interconversion of phosphoglycerate isomers is catalysed in numerous pathogenic microorganisms by a cofactor-independent mutase (iPGM) structurally distinct from the mammalian cofactor-dependent (dPGM) isozyme. The iPGM active site dynamically assembles through substrate-triggered movement of phosphatase and transferase domains creating a solvent inaccessible cavity. Here we identify alternate ligand binding regions using nematode iPGM to select and enrich lariat-like ligands from an mRNA-display macrocyclic peptide library containing >10 members. Functional analysis of the ligands, named ipglycermides, demonstrates sub-nanomolar inhibition of iPGM with complete selectivity over dPGM. The crystal structure of an iPGM macrocyclic peptide complex illuminated an allosteric, locked-open inhibition mechanism placing the cyclic peptide at the bi-domain interface. This binding mode aligns the pendant lariat cysteine thiolate for coordination with the iPGM transition metal ion cluster. The extended charged, hydrophilic binding surface interaction rationalizes the persistent challenges these enzymes have presented to small-molecule screening efforts highlighting the important roles of macrocyclic peptides in expanding chemical diversity for ligand discovery.
磷酸甘油酸异构体的糖酵解互变由一种辅助因子非依赖性突变酶（iPGM）催化，该酶在结构上与哺乳动物依赖辅助因子的（dPGM）同工酶不同。iPGM 活性位点通过磷酸酶和转移酶结构域的底物触发运动动态组装，形成不可接近溶剂的空腔。在这里，我们使用线虫 iPGM 来识别其他配体结合区域，从包含 >10 个成员的 mRNA 展示大环肽文库中选择和富集套索样配体。配体的功能分析表明，ipglycermides 对 iPGM 具有亚纳摩尔级的抑制作用，对 dPGM 具有完全选择性。iPGM 大环肽复合物的晶体结构揭示了一种别构的、锁定的开放抑制机制，将环状肽置于双结构域界面。这种结合模式使侧挂套索半胱氨酸硫醇盐与 iPGM 过渡金属离子簇配位。扩展的带电荷、亲水的结合表面相互作用解释了这些酶一直给小分子筛选工作带来的挑战，突出了大环肽在扩大配体发现的化学多样性方面的重要作用。
