Li Xia, Li Yifeng, Han Huiyun, Miller Donald W, Wang Guangshun
Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, 986805 Nebraska Medical Center, Omaha, Nebraska 68198-6805, USA.
J Am Chem Soc. 2006 May 3;128(17):5776-85. doi: 10.1021/ja0584875.
To understand the structure and activity relationship of human LL-37, a series of peptide fragments was designed. The N-terminal fragment, LL-37(1-12), was not active, while the C-terminal fragment, LL-37(13-37), killed Escherichia coli, as well as drug-sensitive and drug-resistant cancer cells. A 13-residue core antibacterial and anticancer peptide, corresponding to residues 17-29 of LL-37, was identified based on total correlated spectroscopy by trimming nonessential regions (TOCSY-trim). Because LL-37 acts on bacterial membranes, three-dimensional structures of its fragments were determined in micelles by NMR, including structural refinement by natural abundance 15N and 13C chemical shifts. Aromatic-aromatic interactions in the N-terminal fragment were proposed to be essential for LL-37 aggregation. The LL-37 core peptide adopts a similar structure in the micelles of SDS or dioctanoyl phosphatidylglycerol. This structure is retained in the C-terminal fragment LL-37(13-37) and very likely in intact LL-37 based on peptide-aided signal assignments. The higher antibacterial activity of the LL-37 core peptide than aurein 1.2 was attributed to additional cationic residues. To achieve selective membrane targeting, D-amino acids were incorporated into LL-37(17-32). While the D-peptide showed similar antibacterial activity to the L-diastereomer, it lost toxicity to human cells. Structural analysis revealed hydrophobic defects in the new amphipathic structure of the D-peptide, leading to a much shorter retention time on a reversed-phase HPLC column. It is proposed that hydrophobic defects as a result of incoherent hydrophobic packing provide a structural basis for the improvement in cell selectivity of the LL-37 fragment.
为了解人LL-37的结构与活性关系，设计了一系列肽片段。N端片段LL-37(1-12)无活性，而C端片段LL-37(13-37)可杀死大肠杆菌以及药敏和耐药癌细胞。通过修剪非必需区域的全相关谱（TOCSY-trim）鉴定出一个13个残基的核心抗菌和抗癌肽，对应于LL-37的17-29位残基。由于LL-37作用于细菌膜，通过核磁共振在胶束中测定了其片段的三维结构，包括利用天然丰度的15N和13C化学位移进行结构优化。有人提出N端片段中的芳香-芳香相互作用对LL-37的聚集至关重要。LL-37核心肽在十二烷基硫酸钠（SDS）或二辛酰磷脂酰甘油的胶束中具有相似的结构。基于肽辅助信号归属，这种结构保留在C端片段LL-37(13-37)中，并且很可能也保留在完整的LL-37中。LL-37核心肽比奥瑞因1.2具有更高的抗菌活性归因于额外的阳离子残基。为了实现选择性膜靶向，将D-氨基酸掺入LL-37(17-32)中。虽然D-肽显示出与L-非对映异构体相似的抗菌活性，但它对人细胞失去了毒性。结构分析揭示了D-肽新的两亲结构中的疏水缺陷，导致其在反相高效液相色谱柱上的保留时间短得多。有人提出，由于疏水堆积不一致导致的疏水缺陷为LL-37片段细胞选择性的提高提供了结构基础。
