Shi Yong-Xia, Zeng Shao-Ling, Yuan Mei-Jin, Sun Fan, Pang Yi
State Key Laboratory for Biocontrol, Zhongshan(Sun Yat-sen) University, Guangzhou 510275, China.
Wei Sheng Wu Xue Bao. 2006 Jun;46(3):353-7.
p19 gene, cry11Aa gene and p20 gene from Bacillus thuringienesis subsp. israelensis are organized as a single operon. It is reported that P20 polypeptide is not required for high-level expression of Cry11Aa and crystal formation in B. thuringiensis. It is deduced that P19 might relate to Cry11Aa crystallization. In this study, two recombinant plasmids pHcy1 and pHcy3 containing cryllAa gene were constructed, the latter absent from p19 gene encoding a possible accessory protein between cry11Aa promoter and cry11Aa gene. The recombinant plasmids were introduced into an acrystalliferous mutant 4Q7 of B. thuringiensis subsp. israelensis. SDS-PAGE showed that Cry11Aa protein per unit of culture medium had a higher expression level in 4Q7(pHcy1) with p19 and cry11Aa genes than in 4Q7(pHcy3) with only cry11Aa gene. Both two B. thuringiensis strains formed Cry11Aa crystals in a similar size and shape during sporulation. Toxicity bioassay showed 4Q7 (pHcy1) and 4Q7 (pHcy3) exhibited a comparable mosquito-larvicidal activity against 3rd-instar Culex quinquefasciatus. It indicated that accessory protein P19 did not have an effect on cry11Aa crystallization and high mosquitocidal toxicity. However, it could enhance Cry11Aa expression amount to a certain extent.
来自苏云金芽孢杆菌以色列亚种的p19基因、cry11Aa基因和p20基因组成一个单一操纵子。据报道，在苏云金芽孢杆菌中，Cry11Aa的高水平表达和晶体形成不需要P20多肽。推测P19可能与Cry11Aa的结晶有关。在本研究中，构建了两个含有cryllAa基因的重组质粒pHcy1和pHcy3，后者缺少在cry11Aa启动子和cry11Aa基因之间编码一种可能辅助蛋白的p19基因。将重组质粒导入苏云金芽孢杆菌以色列亚种的无晶体突变体4Q7中。SDS-PAGE分析表明，与仅含有cry11Aa基因的4Q7(pHcy3)相比，含有p19和cry11Aa基因的4Q7(pHcy1)中每单位培养基的Cry11Aa蛋白具有更高的表达水平。在芽孢形成过程中，这两株苏云金芽孢杆菌都形成了大小和形状相似的Cry11Aa晶体。毒力生物测定表明，4Q7(pHcy1)和4Q7(pHcy3)对致倦库蚊三龄幼虫表现出相当的杀蚊活性。这表明辅助蛋白P19对Cry11Aa的结晶和高杀蚊毒性没有影响。然而，它可以在一定程度上提高Cry11Aa的表达量。
