Wei Xiumei, Xu Jie, Yang Jianmin, Liu Xiangquan, Zhang Ranran, Wang Weijun, Yang Jialong
Shandong Provincial Key Laboratory of Marine Ecology Restoration, Shandong Marine Resource and Environment Research Institute, Yantai 264006, China.
Yantai Institute of Coastal Zone Research, Chinese Academy of Sciences, Yantai 264003, China.
Fish Shellfish Immunol. 2015 Jan;42(1):79-87. doi: 10.1016/j.fsi.2014.10.028. Epub 2014 Oct 30.
Serpin is an important member of serine protease inhibitors (SPIs), which is capable of regulating proteolytic events and involving in a variety of physiological processes. In present study, a Serpin homolog was identified from Octopus ocellatus (designated as OoSerpin). Full-length cDNA of OoSerpin was of 1735 bp, containing a 5' untranslated region of 214 bp, a 3' UTR of 282 bp, and an open reading frame of 1239 bp. The open reading frame encoded a polypeptide of 412 amino acids which has a predicted molecular weight of 46.5 kDa and an isoelectric point of 8.52. The OoSerpin protein shares 37% sequence identity with other Serpins from Mus musculus (NP_941373) and Ixodes scapularis (XP_002407493). The existence of a conserved SERPIN domain strongly suggested that OoSerpin was a member of the Serpin subfamily. Expression patterns of OoSerpin, both in tissues and towards bacterial stimulation, were then characterized. The mRNA of OoSerpin was constitutively expressed at different levels in all tested tissues of untreated O. ocellatus, including mantle (lowest), muscle, renal sac, gill, hemocyte, gonad, systemic heart, and hepatopancreas (highest). The transcriptional level of OoSerpin was significantly up-regulated (P<0.01) in O. ocellatus upon bacterial challenges with Vibrio anguillarum and Micrococcus luteus, indicating its involvement in the antibacterial immune response. Furthermore, rOoSerpin, the recombinant protein of OoSerpin, exhibited strong abilities to inhibit proteinase activities of trypsin and chymotrypsin as well as the growth of Escherichia coli. Our results demonstrate that OoSerpin is a potential antibacterial factor involved in the immune response of O. ocellatus against bacterial infection.
丝氨酸蛋白酶抑制剂（Serpin）是丝氨酸蛋白酶抑制剂（SPIs）的重要成员，能够调节蛋白水解事件并参与多种生理过程。在本研究中，从短蛸中鉴定出一个Serpin同源物（命名为OoSerpin）。OoSerpin的全长cDNA为1735 bp，包含一个214 bp的5'非翻译区、一个282 bp的3'非翻译区和一个1239 bp的开放阅读框。该开放阅读框编码一个由412个氨基酸组成的多肽，预测分子量为46.5 kDa，等电点为8.52。OoSerpin蛋白与小家鼠（NP_941373）和肩突硬蜱（XP_002407493）的其他Serpin蛋白具有37%的序列同一性。保守的SERPIN结构域的存在强烈表明OoSerpin是Serpin亚家族的成员。随后对OoSerpin在组织中的表达模式以及对细菌刺激的反应进行了表征。在未处理的短蛸的所有测试组织中，OoSerpin的mRNA均以不同水平组成性表达，包括外套膜（最低）、肌肉、肾囊、鳃、血细胞、性腺、体心脏和肝胰腺（最高）。在用鳗弧菌和藤黄微球菌对短蛸进行细菌攻击后，OoSerpin的转录水平显著上调（P<0.01），表明其参与了抗菌免疫反应。此外，OoSerpin的重组蛋白rOoSerpin表现出强大的抑制胰蛋白酶和糜蛋白酶的蛋白酶活性以及抑制大肠杆菌生长的能力。我们的结果表明，OoSerpin是参与短蛸抗细菌感染免疫反应的潜在抗菌因子。
