粉纹夜蛾颗粒体病毒增强蛋白锌离子结合域定点突变

粉纹夜蛾颗粒体病毒（Trichoplusia ni granulovirus, TnGV）增强蛋白（enhancin）具有增强病毒感染力的作用。该蛋白包含一个多种杆状病毒增强蛋白都具有的保守结构域HELGH，是典型的金属蛋白酶锌离子结合域，但该结构域对增强蛋白生物活性的重要性尚未得到研究。本研究通过定点突变构建了该结构域的5个氨基酸分别突变为2种不同氨基酸的共10种增强蛋白突变体基因，并用杆状病毒载体进行了重组表达。活性测定发现，10种突变型增强蛋白大部分都丧失了野生型增强蛋白所具有的降解粉纹夜蛾幼虫围食膜粘蛋白的生物学功能，只有1种（第4位G突变为A）保留该生物学活性。这一结果表明锌离子结合域对增强蛋白生物活性具有重要作用，也提示增强蛋白确是一种金属蛋白酶。

Site-directed mutagenesis of the zinc-binding domain of Trichoplusia ni granulovirus enhancin

Trichoplusia ni granulovirus (TnGV) enhancin can enhance the infectivity of virus in several insect hosts. It has a conserved zinc-binding domain commonly found in metalloproteases, HELGH, which is also present in enhancin proteins from other baculoviruses. In the current work, each of the five amino acids in the domain was mutated to be two different amino acids, and the total 10 mutant enhancin genes were used to construct recombinant AcMNPV. The expression of enhancin proteins was observed in virus-infected cells. Peritrophic membrane (PM) assay indicated all but one mutant lost the ability to degrade the insect intestinal mucin in PM. The only exception was the mutant in which glycine in position 4 was mutated to alanine. The results showed that the zinc-binding domain in enhancin was essential for the activity of the protein, and also suggest that baculovirus enhancin is a metalloprotease.