Membrane Biology Laboratory, Interdisciplinary Research Program , Rajiv Gandhi Centre for Biotechnology , Thiruvananthapuram 695014 , India.
J Am Chem Soc. 2019 Feb 20;141(7):2949-2959. doi: 10.1021/jacs.8b09973. Epub 2019 Feb 12.
The porinACj is an α-helical porin that spans the mycolic acid outer membrane of Gram-positive mycolate, Corynebacterium jeikeium. Here, we report that a 40-amino acid, synthetic peptide, pPorA corresponding to porin PorACj, inserts into the lipid bilayers and forms well-defined pores. By electrical recordings, we measured the single-channel properties that revealed the autonomous assembly of large conductance ion-selective synthetic pores. Further, we characterized the functional properties by blocking the peptide pores by cyclodextrins of different charge and symmetry. We deduced the subunit stoichiometry and putative structure of the pore by site-specific chemical modification in single-channel electrical recordings and gel electrophoresis. On the basis of these findings, we suggest that this is a large functional uniform transmembrane pore built entirely from short synthetic α-helical peptides. Accordingly, we propose a model demonstrating structural assembly of large α-helix-based peptide pores for understanding the action of antimicrobial peptides and for the design of pores with applications in biotechnology.
PorinACj 是一种α-螺旋孔蛋白，跨越革兰氏阳性分枝杆菌的类脂酸外膜，来自 Corynebacterium jeikeium。在这里，我们报告了一个 40 个氨基酸的合成肽 pPorA，它对应于孔蛋白 PorACj，插入脂质双层并形成了明确的孔。通过电记录，我们测量了单通道特性，揭示了大电导离子选择性合成孔的自主组装。此外，我们通过不同电荷和对称性的环糊精来阻断肽孔，从而对其功能特性进行了表征。我们通过单通道电记录和凝胶电泳中的定点化学修饰推导出了亚基的化学计量和孔的假设结构。基于这些发现，我们认为这是一个完全由短的合成α-螺旋肽组成的大型功能均一的跨膜孔。因此，我们提出了一个模型，展示了基于大α-螺旋肽的孔的结构组装，以理解抗菌肽的作用，并设计具有生物技术应用的孔。
