Thiamine diphosphate binds to intermediates in the assembly of adenovirus fiber knob trimers in Escherichia coli.
Thiamine diphosphate binds to intermediates in the assembly of adenovirus fiber knob trimers in Escherichia coli.
复制标题
二磷酸硫胺素与大肠杆菌中腺病毒纤维旋钮三聚体组装的中间体结合。
DOI：
10.1110/ps.072805007
复制
发表时间：
2007
期刊：
影响因子：
--
通讯作者：
Freimuth,Paul
中科院分区：
文献类型：
--
作者：
Schulz,Ryan;Zhang,Yian-Biao;Liu,Chang-Jun;Freimuth,Paul
腺病毒（Ad）同源三聚体纤维蛋白的组装通过其C末端结结构域成核，当表达为重组蛋白片段时，其本身可以三聚化。球三聚体中亚基的非互锁球形结构意味着三聚体通过二聚体中间体从预折叠单体组装，但尚未观察到这些中间体，因此组装机制仍未表征。在这里，我们报告说，表达的Ad血清型2（Ad 2）球是有毒的fori-菌株的大肠杆菌，这是缺陷的从头合成硫胺素（维生素B1）。从athi+菌株中分离的Ad 2球三聚体通过多个色谱步骤与质量相当于二磷酸硫胺素（ThDP）的小分子共纯化。突变体分析没有涉及任何特定的网站ThDP结合。我们的研究结果表明，ThDP可能与组装中间体相关联，并被困在组装的三聚体中，可能在几个大的空腔中的一个中，这些空腔部分溶剂可进入或完全埋在三聚体内部。
Assembly of the adenovirus (Ad) homotrimeric fiber protein is nucleated by its C‐terminal knob domain, which itself can trimerize when expressed as a recombinant protein fragment. The non‐interlocked, globular structure of subunits in the knob trimer implies that trimers assemble from prefolded monomers through a dimer intermediate, but these intermediates have not been observed and the mechanism of assembly therefore remains uncharacterized. Here we report that expression of the Ad serotype 2 (Ad2) knob was toxic forthi− strains ofEscherichia coli, which are defective in de novo synthesis of thiamine (vitamin B1). Ad2 knob trimers isolated from athi+ strain copurified through multiple chromatography steps with a small molecule of mass equivalent to that of thiamine diphosphate (ThDP). Mutant analysis did not implicate any specific site for ThDP binding. Our results suggest that ThDP may associate with assembly intermediates and become trapped in assembled trimers, possibly within one of several large cavities that are partially solvent‐accessible or buried completely within the trimer interior.