Park C B, Kim H S, Kim S C
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Taejeon.
Biochem Biophys Res Commun. 1998 Mar 6;244(1):253-7. doi: 10.1006/bbrc.1998.8159.
The mechanism of action of buforin II, which is a 21-amino acid peptide with a potent antimicrobial activity against a broad range of microorganisms, was studied using fluorescein isothiocyanate (FITC)-labeled buforin II and a gel-retardation experiment. Its mechanism of action was compared with that of the well-characterized magainin 2, which has a pore-forming activity on the cell membrane. Buforin II killed Esche-richia coli without lysing the cell membrane even at 5 times minimal inhibitory concentration (MIC) at which buforin II reduced the viable cell numbers by 6 orders of magnitude. However, magainin 2 lysed the cell to death under the same condition. FITC-labeled buforin II was found to penetrate the cell membrane and accumulate inside E. coli even below its MIC, whereas FITC-labeled magainin 2 remained outside or on the cell wall even at its MIC. The gel-retardation experiment showed that buforin II bound to DNA and RNA of the cells over 20 times strongly than magainin 2. All these results indicate that buforin II inhibits the cellular functions by binding to DNA and RNA of cells after penetrating the cell membranes, resulting in the rapid cell death, which is quite different from that of magainin 2 even though they are structurally similar: a linear amphipathic alpha-helical peptide.
蟾蜍灵II是一种由21个氨基酸组成的肽，对多种微生物具有强大的抗菌活性。本研究使用异硫氰酸荧光素（FITC）标记的蟾蜍灵II并通过凝胶阻滞实验，对其作用机制进行了研究。将其作用机制与已被充分研究的蛙皮素2进行了比较，蛙皮素2对细胞膜具有成孔活性。即使在5倍最小抑菌浓度（MIC）下，蟾蜍灵II也能杀死大肠杆菌，且不会使细胞膜破裂，在此浓度下蟾蜍灵II可使活菌数量减少6个数量级。然而，在相同条件下，蛙皮素2会使细胞裂解死亡。研究发现，即使在低于MIC的浓度下，FITC标记的蟾蜍灵II也能穿透细胞膜并在大肠杆菌内部积累，而即使在MIC浓度下，FITC标记的蛙皮素2仍停留在细胞外部或细胞壁上。凝胶阻滞实验表明，蟾蜍灵II与细胞的DNA和RNA的结合能力比蛙皮素2强20倍以上。所有这些结果表明，蟾蜍灵II通过穿透细胞膜后与细胞的DNA和RNA结合来抑制细胞功能，从而导致细胞迅速死亡，这与蛙皮素2的作用机制截然不同，尽管它们在结构上相似，均为线性两亲性α-螺旋肽。
