The Journal of Membrane Biology ( IF 2.3 ) Pub Date : 2021-02-10 , DOI: 10.1007/s00232-021-00174-1 Charles H Chen 1, 2, 3, 4 , Charles G Starr 5 , Shantanu Guha 5 , William C Wimley 5 , Martin B Ulmschneider 1, 2, 3 , Jakob P Ulmschneider 6
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The use of designed antimicrobial peptides as drugs has been impeded by the absence of simple sequence-structure–function relationships and design rules. The likely cause is that many of these peptides permeabilize membranes via highly disordered, heterogeneous mechanisms, forming aggregates without well-defined tertiary or secondary structure. We suggest that the combination of high-throughput library screening with atomistic computer simulations can successfully address this challenge by tuning a previously developed general pore-forming peptide into a selective pore-former for different lipid types. A library of 2916 peptides was designed based on the LDKA template. The library peptides were synthesized and screened using a high-throughput orthogonal vesicle leakage assay. Dyes of different sizes were entrapped inside vesicles with varying lipid composition to simultaneously screen for both pore size and affinity for negatively charged and neutral lipid membranes. From this screen, nine different LDKA variants that have unique activity were selected, sequenced, synthesized, and characterized. Despite the minor sequence changes, each of these peptides has unique functional properties, forming either small or large pores and being selective for either neutral or anionic lipid bilayers. Long-scale, unbiased atomistic molecular dynamics (MD) simulations directly reveal that rather than rigid, well-defined pores, these peptides can form a large repertoire of functional dynamic and heterogeneous aggregates, strongly affected by single mutations. Predicting the propensity to aggregate and assemble in a given environment from sequence alone holds the key to functional prediction of membrane permeabilization.
Graphic Abstract
中文翻译：
膜穿孔抗菌肽的调谐选择性靶向不同脂质成分的膜
由于缺乏简单的序列-结构-功能关系和设计规则，设计的抗菌肽作为药物的使用受到了阻碍。可能的原因是许多这些肽通过高度无序的异质机制渗透膜，形成没有明确定义的三级或二级结构的聚集体。我们建议将高通量文库筛选与原子计算机模拟相结合，通过将先前开发的通用成孔肽调整为针对不同脂质类型的选择性成孔剂，可以成功应对这一挑战。基于 LDKA 模板设计了 2916 个肽库。使用高通量正交囊泡渗漏测定法合成和筛选文库肽。不同大小的染料被包裹在具有不同脂质成分的囊泡内，以同时筛选孔径和对带负电荷和中性脂质膜的亲和力。从这个筛选中，选择、测序、合成和表征了九种具有独特活性的不同 LDKA 变体。尽管序列变化很小，但这些肽中的每一种都具有独特的功能特性，形成小孔或大孔，并对中性或阴离子脂质双层具有选择性。大规模、无偏的原子分子动力学 (MD) 模拟直接表明，这些肽不是刚性的、明确定义的孔，而是可以形成大量功能动态和异质聚集体，受单突变的强烈影响。
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