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首页> 外文期刊>Journal of Molecular Biology >STRUCTURAL TRANSITIONS DURING BACTERIOPHAGE HK97 HEAD ASSEMBLY
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STRUCTURAL TRANSITIONS DURING BACTERIOPHAGE HK97 HEAD ASSEMBLY

机译：噬菌体HK97头组装过程中的结构转变。

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摘要

Bacteriophage HK97 builds its head shell from a 42 kDa major head protein, but neither this 42 kDa protein nor its processed, 31 kDa form is found in the mature head. Instead, each of the major head-protein subunits is covalently cross-linked into oligomers of five, six or more by a protein cross-linking reaction that occurs both in vivo and in vitro. Mutants that block prohead maturation lead to the accumulation of one of two types of proheads, termed Prohead I and Prohead II. Prohead I is assembled from about 415 copies of the 42 kDa (384 amino acids) protein subunit and accumulates in infections by mutant amU4. Following assembly, the N-terminal 102 amino acids of each subunit are removed, leaving a prohead shell constructed of 31 kDa subunits, called Prohead II, which accumulates in infections by mutant amC2. During DNA packaging, when the prohead shell expands, all of the head protein subunits become covalently cross-linked to other subunits. Purified Prohead II (or, less completely, Prohead I) becomes cross-linked ill vitro in response to any of a number of conditions that induce shell expansion, including conditions commonly used for protein analysis. In vitro cross-linking occurs efficiently in the absence of added cofactors of enzymes, and we propose that cross-linking is catalyzed by shell subunits themselves. Shell expansion is easily monitored by observing a decrease in electrophoretic mobility of Prohead II in agarose gels. Using the mobility shift in agarose gel to monitor expansion and SDS/gel electrophoresis to monitor cross-linking in vitro, we find that expansion precedes and is required for cross-linking, and we propose that expansion triggers the cross-linking reaction. Comparison of peptides isolated from Prohead II and in vitro cross-linked Prohead II shows a single altered major cross-link peptide in which a lysine, originating from lysine169 of the protein sequence, is linked to asparagine356, presumably derived from the neighboring subunit. Examination of the cross-link-containing peptide by mass spectrometry shows that the cross-link bond is an amide between the side-chains of the lysine and the asparagine residues. [References: 31]
机译：噬菌体HK97用42 kDa的主要头部蛋白构建头壳，但是在成熟的头部中既没有发现这种42 kDa的蛋白质，也没有加工出的31 kDa形式。相反，每个主要的头蛋白亚基通过体内和体外发生的蛋白质交联反应共价交联成五个，六个或更多的寡聚物。阻碍前额成熟的突变体导致两种前额累积之一，称为前额I和前额II。 Prohead I由约415个拷贝的42 kDa（384个氨基酸）蛋白亚基组装而成，并在突变amU4的感染中积累。组装后，去除每个亚基的N端102个氨基酸，剩下一个由31 kDa亚基构成的前额壳，称为Prohead II，它在突变amC2的感染中积累。在DNA包装过程中，当前额壳扩张时，所有头蛋白质亚基都与其他亚基共价交联。纯化的Prohead II（或更简而言之，Prohead I）在体外会因多种诱导壳膨胀的条件（包括通常用于蛋白质分析的条件）而发生交联。在没有添加酶的辅助因子的情况下，体外交联有效地发生，我们提出交联是由壳亚基本身催化的。通过观察琼脂糖凝胶中Prohead II电泳迁移率的降低，可以轻松监测壳的膨胀。使用琼脂糖凝胶中的迁移率变化来监测扩展，并使用SDS /凝胶电泳来监测体外交联，我们发现扩展先于交联并且是交联所必需的，并且我们建议扩展触发交联反应。从Prohead II和体外交联的Prohead II分离的肽的比较显示单个改变的主要交联肽，其中源自蛋白序列的lysine169的赖氨酸与天冬酰胺356相连，而天冬酰胺356可能源自邻近的亚基。通过质谱检查包含交联的肽表明，交联键是赖氨酸的侧链与天冬酰胺残基之间的酰胺。 [参考：31]

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