与凡纳滨对虾抗副溶血弧菌有关的6.0 kD血蓝蛋白降解肽段  

A novel 6.0 kD fragment resistant to Vibrio parahaemolyticus derived from shrimp,Litopenaeus vannamei,hemocyanin

作　　者：张泽蕙 黄河[1] 文英[1] 钟名其[1] 伦镜盛[1] 章跃陵[1] 

机构地区：[1]汕头大学理学院生物学系广东省海洋生物技术重点实验室,广东汕头515063

出　　处：《中国水产科学》2016年第1期218-224,共7页Journal of Fishery Sciences of China

基　　金：国家自然科学基金项目(31372558);教育部新世纪优秀人才支持计划项目(NCET-11-0922);广东省自然科学基金项目(S2013010015190);广东省高等学校科技创新重点项目(2012CXZD0025)

摘　　要：以凡纳滨对虾（Litopenaeus vannamei）为研究对象，采用人工感染、Tricine．SDS．PAGE、Western．blotting、MALDI-TOF／TOF、抑菌实验等方法对血蓝蛋白小分子降解肽段进行研究。结果发现，副溶血弧菌（Vibriopara-haemolyticus）刺激对虾24h后，与对照组相比，其血淋巴中新出现5个分子量为6．0～31．0kD的与兔抗血蓝蛋白抗体呈阳性的小分子条带。其中，分子量为6．0kD的a条带（命名为HMCp6肽段）与凡纳滨对虾血蓝蛋白具有高度同源性，对副溶血弧菌具有明显的抑菌活性，与对照组相比，存在极显著性差异（P〈0．01）。由此说明，HMCp6应该是对虾感染副溶血弧菌后所产生的一种新的小分子降解肽段，推测其在对虾抗感染防御中发挥重要作用。Recent reports have revealed that hemocyanin in shrimp and other invertebrates is a novel multifunctional protein with immunological activities, such as phenoloxidase, antiviral, antibacterial, hemolytic, and antitumor activities Moreover, several studies--including our own previous findings--indicate that hemocyanin may produce some func- tional fragments involved in pathogen resistance in shrimp. However, no study has comprehensively evaluated these fragments. We conducted a comparative analysis in the proteome of the shrimp Litopenaeus vannamei after the shrimp were infected with Vibrio parahaemolyticus for 24 h to search for novel hemocyanin fragments. Tricine-sodium dodecyl sulfate-potyacrylamide gel electrophoresis and Western blotting analyses of hemolymph drawn directly from the peri- cardial sinus revealed five peptide fragments （bands a, c, e, f, g）, ranging from 6.0 to 31.0 kD, that bound to anti-shrimp hemocyanin antibodies in plasma, compared with those in the control group, suggesting that they might be hemocyanin degradation products. Moreover, a 6.0 kD peptide fragment （named HMCp6） was subjected to matrix-assisted laser desorption/ionization-tandem time-of-flight mass spectrometry analysis, and the database search results indicated that the HMCp6 peptide fragment was highly homologous with two subunits ofL. vannamei hemocyanin （gil325462537 and gil854403）. This result is similar to the finding that the C- and N-terminal hemocyanin fragments are signifi- cantly altered in Penaeus vannamei hemocytes during Taura syndrome virus infection. The HMCp6 peptide fragment possessed clear antibacterial activities （antibacterial rate, 98%） against V. parahaemolytieus. This result agrees with a previous finding that a 1.9 kD antibacterial peptide from Pacifastacus leniusculus hemocyanin possesses clear antibac- terial activities against Gram-positive bacteria. In conclusion, our data suggest that the HMCp6 peptide fragment might be a novel fragment derived from hemocyanin with a pathogen-resis

关 键 词：凡纳滨对虾 血蓝蛋白 降解肽段 HMCp6 抑菌活性 

分 类 号：S917[农业科学—水产科学]