Department of Pharmacy, Uppsala University, SE-75232, Uppsala, Sweden.
Department of Biophysics, Bose Institute, P-1/12 CIT Scheme VII (M), Kolkata, 700054, India.
Sci Rep. 2017 Mar 16;7(1):212. doi: 10.1038/s41598-017-00188-7.
The objective of the present study is the investigation of possibilities for boosting peptide anti-inflammatory effects by tryptophan end-tagging, including identification of underlying mechanisms for this. In doing so, effects of tryptophan end-tagging of KYE21 (KYEITTIHNLFRKLTHRLFRR), a peptide derived from heparin co-factor II, on membrane and lipopolysaccharide (LPS) interactions were investigated by ellipsometry, NMR, fluorescence spectroscopy, and circular dichroism measurements. Through its N-terminal W stretch, WWWKYE21 displays higher membrane binding, liposome rupture, and bacterial killing than unmodified KYE21. Analogously, W-tagging promotes binding to E. coli LPS and to its endotoxic lipid A moiety. Furthermore, WWWKYE21 causes more stable peptide/LPS complexes than KYE21, as evidenced by detailed NMR studies, adopting a pronounced helical conformation, with a large hydrophobic surface at the N-terminus due to the presence of W-residues, and a flexible C-terminus due to presence of several positively charged arginine residues. Mirroring its increased affinity for LPS and lipid A, WWWKYE21 displays strongly increased anti-inflammatory effect due to a combination of direct lipid A binding, peptide-induced charge reversal of cell membranes for LPS scavenging, and peptide-induced fragmentation of LPS aggregates for improved phagocytosis. Importantly, potent anti-inflammatory effects were observed at low cell toxicity, demonstrated for both monocytes and erythrocytes.
本研究的目的是探讨通过色氨酸末端标记来增强肽类抗炎作用的可能性，包括确定其潜在机制。为此，通过椭圆测量法、NMR、荧光光谱法和圆二色性测量法研究了肝素辅因子 II 衍生肽 KYE21（KYEITTIHNLFRKLTHRLFRR）的色氨酸末端标记对膜和脂多糖（LPS）相互作用的影响。通过其 N 端的 W 伸展，与未修饰的 KYE21 相比，WWWKYE21 显示出更高的膜结合、脂质体破裂和杀菌能力。类似地，W 标记促进了与大肠杆菌 LPS 及其内毒素脂质 A 部分的结合。此外，详细的 NMR 研究表明，WWWKYE21 比 KYE21 能形成更稳定的肽/LPS 复合物，采用明显的螺旋构象，由于存在 W 残基，N 端具有较大的疏水面，由于存在几个带正电荷的精氨酸残基，C 端具有柔韧性。由于其与 LPS 和脂质 A 的亲和力增加，WWWKYE21 由于直接结合脂质 A、肽诱导细胞膜电荷反转以清除 LPS 以及肽诱导 LPS 聚集物碎裂以提高吞噬作用的综合作用，表现出强烈的抗炎作用。重要的是，在低细胞毒性下观察到了强大的抗炎作用，在单核细胞和红细胞中均得到了证明。
