Mass Spectral Analyses of Salmonella Myovirus SPN3US Reveal Conserved and Divergent Themes in Proteolytic Maturation of Large Icosahedral Capsids.
Mass Spectral Analyses of Salmonella Myovirus SPN3US Reveal Conserved and Divergent Themes in Proteolytic Maturation of Large Icosahedral Capsids.
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DOI：
10.3390/v15030723
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发表时间：
2023-03-10
期刊：
影响因子：
--
通讯作者：
Thomas JA
中科院分区：
文献类型：
--
作者：
Scheuch A;Moran SAM;Faraone JN;Unwin SR;Vu G;Benítez AD;Mohd Redzuan NH;Molleur D;Pardo S;Weintraub ST;Thomas JA
沙门氏菌肌病毒SPN3US有一个T=27的衣壳，由50种不同的基因产物组成，其中包括许多与240kb基因组一起包装并喷射到宿主细胞中的基因产物。最近，我们发现在SPN3US头部组装过程中，一个必需的噬菌体编码的前头蛋白水解酶gp245负责蛋白质的切割。这一蛋白分解成熟步骤诱导前体头部颗粒发生重大变化，使它们能够扩张并经历基因组包装。为了全面确定成熟SPN3US头部的组成，并阐明它在组装过程中是如何被蛋白质降解修饰的，我们对纯化的病毒粒子和无尾头部进行了串联质谱分析。在9种蛋白质中确定了14个蛋白酶切割位点，其中8个在活体中没有发现。其中包括gp245的成熟裂解位点，它与我们先前在纯化的重组gp245中鉴定的自切位点相同。我们的发现强调了采用多个基于质谱学的实验策略作为一种增强尾部噬菌体头部蛋白裂解位点检测的方法的价值。此外，我们的结果还在相关的巨噬菌体中发现了一组保守的头部蛋白，这些蛋白类似地被各自的Prohead蛋白酶切割，表明这些蛋白在调控大的二十面体衣壳的形成和功能方面发挥着重要作用。
Salmonella myovirus SPN3US has a T = 27 capsid composed of >50 different gene products, including many that are packaged along with the 240 kb genome and ejected into the host cell. Recently, we showed that an essential phage-encoded prohead protease gp245 is responsible for cleavage of proteins during SPN3US head assembly. This proteolytic maturation step induces major changes in precursor head particles, enabling them to expand and undergo genome packaging. To comprehensively define the composition of the mature SPN3US head and elucidate how it is modified by proteolysis during assembly, we conducted tandem mass spectrometry analysis of purified virions and tailless heads. Fourteen protease cleavage sites were identified in nine proteins, including eight sites not previously identified in head proteins in vivo. Among these was the maturation cleavage site of gp245 which was identical to the autocleavage site we had previously identified in purified recombinant gp245. Our findings underscore the value of employing multiple mass spectrometry-based experimental strategies as a way to enhance the detection of head protein cleavage sites in tailed phages. In addition, our results have identified a conserved set of head proteins in related giant phages that are similarly cleaved by their respective prohead proteases, suggesting that these proteins have important roles in governing the formation and function of large icosahedral capsids.
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DOI：
10.1073/pnas.0400444101
发表时间：
2004-04-20
影响因子：
11.1
作者：
Fokine, A;Chipman, PR;Rossmann, MG
通讯作者：
Rossmann, MG
影响因子：
5.4
作者：
Thomas JA;Benítez Quintana AD;Bosch MA;Coll De Peña A;Aguilera E;Coulibaly A;Wu W;Osier MV;Hudson AO;Weintraub ST;Black LW
通讯作者：
Black LW
影响因子：
5.6
作者：
Maurer, Joshua B.;Oh, Bonnie;Duda, Robert L.
通讯作者：
Duda, Robert L.
影响因子：
5.6
作者：
SHOWE, MK;ISOBE, E;ONORATO, L
通讯作者：
ONORATO, L
影响因子：
3.7
作者：
Sycheva, Lada V.;Shneider, Mikhail M.;Leiman, Petr G.
通讯作者：
Leiman, Petr G.