Engineering of a novel thioether bridge and role of modified residues in the lantibiotic pep5
Engineering of a novel thioether bridge and role of modified residues in the lantibiotic pep5
复制标题
DOI：
10.1128/aem.62.2.385-392.1996
复制
发表时间：
1996-02-01
影响因子：
4.4
通讯作者：
Sahl, HG
中科院分区：
文献类型：
--
作者：
Bierbaum, G;Szekat, C;Sahl, HG
Pep5是一种含有34个氨基酸的抗菌肽，由表皮葡萄球菌5产生，含有硫醚氨基酸衣硫氨酸和甲基衣硫氨酸，它们形成三个分子内环结构，此外，两个二脱氢丁氨酸存在于lantibiotic的中心部分，一个氧丁基残基位于N端，所有罕见的氨基酸都是通过核糖体制造的前体肽的翻译后修饰引入的。为了阐明这些修饰残基对Pep5抗菌作用的作用，通过位点定向诱变产生了去除单个修饰残基的突变肽，这些突变肽的抗菌活性都降低了。此外，那些环结构被删除的肽变得容易被蛋白水解消化。这表明，环状结构可以作为活性必需构象的稳定剂，例如，两亲性，以及保护pep免受产生菌株蛋白酶的侵害。此外，可以作为l抗生素中新修饰氨基酸前体的残基被引入Pep5前体肽，通过这种方式，新的甲基硫氨酸和二脱氢丙氨酸被插入到Pep5的柔性中心部分。证明通过蛋白质工程和使用非抗生素修饰系统生物合成修饰氨基酸是可行的。
Pep5 is a 34-amino-acid antimicrobial peptide, produced by Staphlococcus epidermidis 5, that contains the thioether amino acids lanthionine and methyllanthionine, which form three intramolecular ring structures, In addition, two didehydrobutyrines are present in the central part of the lantibiotic and an oxobutyryl residue is located at the N terminus, All rare amino acids are introduced by posttranslational modifications of a ribosomally made precursor peptide, To elucidate the function of the modified residues for the antimicrobial action of Pep5, mutant peptides, in which single modified residues had been eliminated, were produced by site-directed mutagenesis, All of these peptides showed a reduced antimicrobial activity. In addition, those peptides from which the ring structures had been deleted became susceptible to proteolytic digest. This demonstrates that the ring structures serve as stabilizers of conformations essential for activity, e.g., amphiphilicity, as well as for protecting PepS against proteases of the producing strains, In addition, residues that could serve as precursors of new modified amino acids in lantibiotics were introduced into the Pep5 precursor peptide, This way, a novel methyllanthionine and a didehydroalanine were inserted into the flexible central part of Pep5, demonstrating that biosynthesis of modified amino acids is feasible by protein engineering and use of the lantibiotic modification system.