Zhou Qing-Feng, Luo Xue-Gang, Ye Liang, Xi Tao
Department of Marine Biochemistry Engineer, School of Life Science and Technology China Pharmaceutical University, Nanjing, 210009, People's Republic of China.
Curr Microbiol. 2007 May;54(5):366-70. doi: 10.1007/s00284-006-0466-y. Epub 2007 May 4.
Perinerin is a small antimicrobial peptide (AMP) isolated from an Asian marine clamworm, Perinereis aibuhitensis Grube. It shows marked activity in vitro against both Gram-negative and Gram-positive bacteria. To obtain it in large amounts, the coding sequence of perinerin was cloned into pET32a(+) vector and expression as a Trx fusion protein in Escherichia coli. The soluble fusion protein collected from the supernatant of the cell lyste was separated by Ni(2+)-chelating chromatography. The purified protein was then cleaved by Factor Xa protease to release mature perinerin. Final purification was achieved by ion-exchange chromatography. Recombinant perinerin exhibited a similar antimicrobial activity to the native perinerin. These works might provide a significant foundation for the following research on the action of mechanism of marine AMPs.
围沙蚕抗菌肽是从亚洲海沙蚕（Perinereis aibuhitensis Grube）中分离出的一种小抗菌肽（AMP）。它在体外对革兰氏阴性菌和革兰氏阳性菌均表现出显著活性。为大量获得该抗菌肽，将围沙蚕抗菌肽的编码序列克隆到pET32a(+)载体中，并在大肠杆菌中表达为Trx融合蛋白。从细胞裂解液上清中收集的可溶性融合蛋白通过Ni(2+)螯合层析进行分离。然后用因子Xa蛋白酶切割纯化后的蛋白以释放成熟的围沙蚕抗菌肽。最终通过离子交换层析实现纯化。重组围沙蚕抗菌肽表现出与天然围沙蚕抗菌肽相似的抗菌活性。这些工作可能为后续关于海洋抗菌肽作用机制的研究提供重要基础。
