Rosengren K Johan, Blond Alain, Afonso Carlos, Tabet Jean-Claude, Rebuffat Sylvie, Craik David J
Institute for Molecular Bioscience, University of Queensland, Brisbane, QLD 4072, Australia.
Biochemistry. 2004 Apr 27;43(16):4696-702. doi: 10.1021/bi0361261.
The structure of a two-chain peptide formed by the treatment of the potent antimicrobial peptide microcin J25 (MccJ25) with thermolysin has been characterized by NMR spectroscopy and mass spectrometry. The native peptide is 21 amino acids in size and has the remarkable structural feature of a ring formed by linkage of the side chain of Glu8 to the N-terminus that is threaded by the C-terminal tail of the peptide. Thermolysin cleaves the peptide at the Phe10-Val11 amide bond, but the threading of the C-terminus through the N-terminal ring is so tight that the resultant two chains remain associated both in the solution and in the gas phases. The three-dimensional structure of the thermolysin-cleaved peptide derived using NMR spectroscopy and simulated annealing calculations has a well-defined core that comprises the N-terminal ring and the threading C-terminal tail. In contrast to the well-defined core, the newly formed termini at residues Phe10 and Val11 are disordered in solution. The C-terminal tail is associated to the ring both by hydrogen bonds stabilizing a short beta-sheet and by hydrophobic interactions. Moreover, unthreading of the tail through the ring is prevented by the bulky side chains of Phe19 and Tyr20, which flank the octapeptide ring. This noncovalent two-peptide complex that has a remarkable stability in solution and in highly denaturing conditions and that survives in the gas phase is the first example of such a two-chain peptide lacking disulfide or interchain covalent bonds.
通过核磁共振光谱法和质谱法对强效抗菌肽微菌素J25（MccJ25）经嗜热菌蛋白酶处理后形成的双链肽结构进行了表征。天然肽大小为21个氨基酸，具有显著的结构特征，即Glu8的侧链与N端相连形成一个环，肽的C端尾巴穿过该环。嗜热菌蛋白酶在Phe10-Val11酰胺键处切割该肽，但C端穿过N端环的缠绕非常紧密，以至于所得的两条链在溶液和气相中都保持结合。利用核磁共振光谱法和模拟退火计算得出的嗜热菌蛋白酶切割肽的三维结构具有一个明确的核心，该核心由N端环和缠绕的C端尾巴组成。与明确的核心不同，Phe10和Val11残基处新形成的末端在溶液中是无序的。C端尾巴通过稳定短β-折叠的氢键和疏水相互作用与环相连。此外，位于八肽环两侧 的Phe19和Tyr20的庞大侧链阻止了尾巴从环中解缠。这种在溶液和高度变性条件下具有显著稳定性且能在气相中存活的非共价双链肽复合物是此类缺乏二硫键或链间共价键的双链肽的首个实例。
