海水养殖真鲷弧菌病病原菌外毒素的理化特性

THE CHARACTERISTICS OF THE EXOTOXIN Vm-Pm PRODUCED BY VIBRIO MIMICUS

对最小弧菌Vm Pm毒素的分子量、分子结构及氨基酸组分进行了测定 ,同时还研究了温度、酸碱度及胰酶等因子对该毒素生物学活性的影响。研究结果表明 ,该毒素为单一的多肽 ,不具备典型的细菌毒素A、B亚基结构 ,分子量为 30 4 5 6kD ;毒素蛋白含天冬氨酸、苏氨酸、丝氨酸及半胱氨酸等 15种氨基酸 ,其中天冬氨酸的含量最多 ,半胱氨酸的含量最少 ,酸性氨基酸含量为 2 6 6 % ,碱性氨基酸含量为 14 4 % ,疏水性氨基酸含量为 33 4 % ;毒素活性的最适pH为 6— 8,对胰酶有抗性 ;不耐热 ,10 0℃下作用 1min ,其毒性完全消失 ,而在 5 6℃下 ,毒素完全失去活性需要 4 0min。

Toxin Vm-Pm produced by Vibrio mimicus was purified in procedures as follows:salting out of ammonium sulfate,anion exchange chromatography on DEAE-cellulose and gel filtration chromatography of Sephadex G100.The purified toxin Vm-Pm gave a single band with a molecular weight of 30.456kD following SDS-PAGE with or without β-mercaptoethanol,indicating that the toxin Vm-Pm did not contain inter-chain disulphide bonds in its conformation and was a single-chain polypeptide.Amino acid composition analysis demonstrated that the toxin protein contained fifteen different kinds of amino acids and revealed a comparatively higher percentage of acidic amino acids(26.6%)than alkalic amino acids(14.4%).Hydrophobic amino acids corresponded to 33.4% of total amino acid residues.The optimal pH for the toxin was 6-8 and resistant to trypsase. Heat treatment at 56℃ and 100℃ abolished the hemolytic activity of the toxin.The toxin completely lost its activity when incubated at temperatures of 56℃ and 100℃ for 40min and 1min,respectively. The toxin showed unique characteristics as compared with other known bacterial toxins.