Cloning, functional characterization, and mode of action of a novel insecticidal pore-forming toxin, sphaericolysin, produced by Bacillus sphaericus
Cloning, functional characterization, and mode of action of a novel insecticidal pore-forming toxin, sphaericolysin, produced by Bacillus sphaericus
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DOI：
10.1128/aem.00021-07
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发表时间：
2007-05-01
影响因子：
4.4
通讯作者：
Matsuda, Kazuhiko
中科院分区：
文献类型：
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作者：
Nishiwaki, Hisashi;Nakashima, Kenta;Matsuda, Kazuhiko
对球形芽孢杆菌A3-2产生的一种杀虫蛋白进行了纯化，以阐明其结构和作用方式。从A3-2的培养液中纯化的活性成分是一种分子量为53 kDa的蛋白质，当注射时，该蛋白质以约100 ng的剂量迅速使德国小蠊（Blattela germanica）中毒。该杀虫蛋白具有胆固醇依赖性溶细胞素的十一肽基序，并且具有独特的N端序列。在大肠杆菌中表达的重组蛋白与天然蛋白一样有效。溶血素诱导的溶血作用是由于该蛋白的成孔作用。这种活性以及杀虫活性被Y159 A突变显著降低。此外，共同应用的球孢菌素与胆固醇废除杀虫行动，表明胆固醇结合在杀虫活性中起着重要作用。从德国小蠊胸神经节分离的Sphaericolysin裂解的神经元。此外，Sphaericolysin在神经节中的活性被Y159 A突变抑制。的sphaericolysin诱导的蟑螂神经节的损害是大于常见的切根虫（斜纹夜蛾），这占，至少部分，为sphaericolysin显示的蟑螂比显示的切根虫的神经节的损害。
An insecticidal protein produced by Bacillus sphaericus A3-2 was purified to elucidate its structure and mode of action. The active principle purified from the culture broth of A3-2 was a protein with a molecular mass of 53 kDa that rapidly intoxicated German cockroaches (Blattela germanica) at a dose of about 100 ng when injected. The insecticidal protein sphaericolysin possessed the undecapeptide motif of cholesterol-dependent cytolysins and had a unique N-terminal sequence. The recombinant protein expressed in Escherichia coli was equally as potent as the native protein. Sphaericolysin-induced hemolysis resulted from the protein's pore-forming action. This activity as well as the insecticidal activity was markedly reduced by a Y159A mutation. Also, coapplication of sphaericolysin with cholesterol abolished the insecticidal action, suggesting that cholesterol binding plays an important role in insecticidal activity. Sphaericolysin-lysed neurons dissociated from the thoracic ganglia of the German cockroaches. In addition, sphaericolysin's activity in ganglia was suppressed by the Y159A mutation. The sphaericolysin-induced damage to the cockroach ganglia was greater than the damage to the ganglia of common cutworms (Spodoptera litura), which accounts, at least in part, for the higher sensitivity to sphaericolysin displayed by the cockroaches than that displayed by cutworms.