Hwang Jee Youn, Ohira Tsuyoshi, Hirono Ikuo, Aoki Takashi
Laboratory of Genome Science, Graduate School of Marine Science and Technology, Tokyo University of Marine Science and Technology, Konan 4-5-7, Minato, 108-8477, Tokyo, Japan.
Immunogenetics. 2004 Aug;56(5):360-7. doi: 10.1007/s00251-004-0688-8. Epub 2004 Jul 29.
A perforin cDNA of Japanese flounder, Paralichthys olivaceus, was cloned from a cDNA library of kidney stimulated with ConA/PMA. The full-length cDNA is 2,157 bp, which encodes 587 amino acids. The Japanese flounder perforin gene consists of five exons and four introns, with a length of approximately 3 kb. The amino acid sequence of the Japanese flounder perforin is 36% identical to that of rat perforin and 37% identical to amino acid sequences of mouse and human perforin. The Japanese flounder perforin also showed low homology to human and mouse complement components (C6, C7, C8 and C9), ranging from 19% to 24%. However, the membrane attack complex/perforin domain is conserved. A phylogenetic analysis placed the Japanese flounder perforin in the same cluster with other known mammalian perforins. RT-PCR analysis revealed that the perforin gene was expressed in the peripheral blood leukocytes, head kidney, trunk kidney, spleen, heart, gill and intestine of healthy fish. Recombinant perforin produced in insect cells using the baculovirus expression system showed calcium-dependent hemolytic activity.
从经刀豆蛋白A/佛波酯刺激的牙鲆（Paralichthys olivaceus）肾脏cDNA文库中克隆到了牙鲆穿孔素cDNA。该全长cDNA为2157 bp，编码587个氨基酸。牙鲆穿孔素基因由5个外显子和4个内含子组成，长度约为3 kb。牙鲆穿孔素的氨基酸序列与大鼠穿孔素的氨基酸序列一致性为36%，与小鼠和人穿孔素的氨基酸序列一致性为37%。牙鲆穿孔素与人和小鼠补体成分（C6、C7、C8和C9）的同源性也较低，在19%至24%之间。然而，膜攻击复合物/穿孔素结构域是保守的。系统发育分析表明，牙鲆穿孔素与其他已知的哺乳动物穿孔素位于同一簇中。RT-PCR分析显示，穿孔素基因在健康鱼类的外周血白细胞、头肾、躯干肾、脾脏、心脏、鳃和肠道中表达。使用杆状病毒表达系统在昆虫细胞中产生的重组穿孔素表现出钙依赖性溶血活性。
