Identification and characterization of the domain structure of bacteriophage P22 coat protein
Identification and characterization of the domain structure of bacteriophage P22 coat protein
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DOI：
10.1021/bi9915420
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发表时间：
1999-11-02
期刊：
影响因子：
2.9
通讯作者：
Prevelige, PE
中科院分区：
文献类型：
--
作者：
Lanman, J;Tuma, R;Prevelige, PE
噬菌体P22是组装二十面体dsDNA病毒的模型。P22蛋白是DNA包装的前体，它是由420份单一外壳蛋白在化学计量比的支架蛋白的帮助下构建的。一旦DNA进入，原衣壳就会膨胀并成熟为稳定的病毒粒子。有人认为，膨胀是由铰链弯曲和磁区移动所介导的。我们使用有限的蛋白分解来绘制外壳蛋白结构域结构的动态稳定性。外壳蛋白单体对蛋白水解性很敏感，但用少量的弹性蛋白酶或胰凝乳酶进行限制的蛋白水解会产生两个亚稳片段(结构域)。N-末端结构域(残基1-180)通过对蛋白酶敏感的环(残基180-205)连接到C-末端结构域(残基205-429)。在环切割后，这两个结构域保持关联。虽然环裂解只导致二级结构的微小变化，但三级结构域间的接触和亚基的热稳定性都降低了。完整的环路也是单体外壳蛋白组装成Proapsids所必需的。一旦组装，外壳蛋白在很大程度上变得对蛋白酶具有抵抗力，在大约一半的亚基的环区内暴露出裂解。环状裂解降低了前衣壳的稳定性，促进了热诱导的壳层膨胀。一旦扩张，这个环就变得对蛋白酶具有抵抗力。我们的数据表明，环区在组装和成熟过程中变得更加有序，从而在这两个阶段都发挥着重要作用。
The bacteriophage P22 serves as a model for assembly of icosahedral dsDNA viruses. The P22 procapsid, which constitutes the precursor for DNA packaging, is built from 420 copies of a single coat protein with the aid of stoichiometric amounts of scaffolding protein. Upon DNA entry, the procapsid shell expands and matures into a stable virion. It was proposed that expansion is mediated by hinge bending and domain movement. We have used limited proteolysis to map the dynamic stability of the coat protein domain structures. The coat protein monomer is susceptible to proteolytic digestion, but limited proteolysis by small quantities of elastase or chymotrypsin yielded two metastable fragments (domains). The N-terminal domain (residues 1-180) is linked to the C-terminal domain (residues 205-429) by a protease-susceptible loop (residues 180-205). The two domains remain associated after the loop cleavage. Although only a small change of secondary structure results from the loop cleavage, both tertiary interdomain contacts and subunit thermostability are diminished. The intact loop is also required for assembly of the monomeric coat protein into procapsids. Upon assembly, coat protein becomes largely protease-resistant, baring cleavage within the loop region of about half of the subunits. Loop cleavage decreases the stability of the procapsids and facilitates heat-induced shell expansion. Upon expansion, the loop becomes protease-resistant. Our data suggest the loop region becomes more ordered during assembly and maturation and thereby prays an important role in both of these stages.