Tam J P, Lu Y A, Yang J L, Chiu K W
Department of Microbiology and Immunology, Vanderbilt University, A-5119 MCN, 1161 21st Avenue South, Nashville, TN 37232-2363, USA.
Proc Natl Acad Sci U S A. 1999 Aug 3;96(16):8913-8. doi: 10.1073/pnas.96.16.8913.
Four macrocyclic cystine-knot peptides of 29-31 residues, kalata, circulin A and B (CirA and CirB), and cyclopsychotride, have been isolated from coffee plants but have undetermined physiological functions. These macrocycles and 10 of their analogs prepared by chemical synthesis were tested against nine strains of microbes. Kalata and CirA were specific for the Gram-positive Staphylococcus aureus with a minimum inhibition concentration of approximately 0.2 microM. They were relatively ineffective against Gram-negative bacteria such as Escherichia coli and Pseudomonas aeruginosa. However, CirB and cyclopsychotride were active against both Gram-positive and Gram-negative bacteria. In particular, CirB showed potent activity against E. coli with a minimum inhibitory concentration of 0.41 microM. All four cyclic peptides were moderately active against two strains of fungi, Candida kefyr and Candida tropicalis, but were inactive against Candida albicans. These macrocycles are cytotoxic and lysed human red blood cell with a lethal dose 50% of 400 microM. Modifying the Arg residue in kalata with a keto aldehyde significantly reduced its activity against S. aureus whereas blocking the arg in CirA produced no significant effect. The two-disulfide variants and their scrambled disulfide isomers exhibited antimicrobial profiles and potency similar to their native peptides. However, in high-salt assays (100 mM NaCl), few of these macrocyclic peptides, natives or analogs, retained antimicrobial activity. These results show that the macrocyclic peptides possess specific and potent antimicrobial activity that is salt-dependent and that their initial interactions with the microbial surfaces may be electrostatic, an effect commonly found in defensin antimicrobial peptides. Furthermore, their end-to-end cyclic structure with a cystine-knot motif represents a molecular structure of antimicrobials and may provide a useful template for the design of novel peptide antibiotics.
从咖啡植物中分离出了4种由29 - 31个氨基酸残基组成的大环胱氨酸结肽，即卡拉塔肽、环蛋白A和B（CirA和CirB）以及环精神分裂肽，但它们的生理功能尚未确定。对这些大环化合物及其通过化学合成制备的10种类似物针对9种微生物菌株进行了测试。卡拉塔肽和CirA对革兰氏阳性金黄色葡萄球菌具有特异性，最低抑菌浓度约为0.2微摩尔。它们对革兰氏阴性菌如大肠杆菌和铜绿假单胞菌相对无效。然而，CirB和环精神分裂肽对革兰氏阳性菌和革兰氏阴性菌均有活性。特别是，CirB对大肠杆菌表现出强效活性，最低抑菌浓度为0.41微摩尔。所有这4种环肽对两株真菌，即凯氏假丝酵母和热带假丝酵母有中等活性，但对白色念珠菌无活性。这些大环化合物具有细胞毒性，能使人类红细胞裂解，半数致死剂量为400微摩尔。用酮醛修饰卡拉塔肽中的精氨酸残基显著降低了其对金黄色葡萄球菌的活性，而封闭CirA中的精氨酸则没有显著影响。双二硫键变体及其错配二硫键异构体表现出与天然肽相似的抗菌谱和效力。然而，在高盐试验（100 mM氯化钠）中，这些大环肽（天然的或类似物）很少保留抗菌活性。这些结果表明，大环肽具有特异性且强效的抗菌活性，这种活性依赖于盐，并且它们与微生物表面的初始相互作用可能是静电作用，这是防御素抗菌肽中常见的一种效应。此外，它们具有胱氨酸结基序的端到端环状结构代表了一种抗菌分子结构，可能为新型肽类抗生素的设计提供有用的模板。
