Betadoublet: de novo design, synthesis, and characterization of a beta-sandwich protein.
Betadoublet: de novo design, synthesis, and characterization of a beta-sandwich protein.
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Betadoublet：β-夹心蛋白的从头设计、合成和表征。
DOI：
10.1073/pnas.91.19.8747
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发表时间：
1994
影响因子：
11.1
通讯作者：
Richardson,DC
中科院分区：
文献类型：
--
作者：
Quinn,TP;Tweedy,NB;Williams,RW;Richardson,JS;Richardson,DC
氨基酸序列如何编码蛋白质采用独特三级结构所需的信息仍然没有得到解决。我们正在通过“从头开始”设计蛋白质分子来解决这个问题，这些分子将采用预定的三维结构。基于这种策略，设计了两个相同的四链β折叠，使其二聚化并形成β夹心蛋白，称为betadaptolet。在大肠杆菌中表达编码一半β-夹心蛋白的合成基因，并将蛋白纯化至均一。倍他赛在水溶液中的生物物理表征表明，在两个折叠之间形成二硫化物，并且二聚体是紧凑的未聚集的球状蛋白，主要由β-折叠组成并且对热变性稳定。它有一些骨架酰胺质子，其交换足够慢以通过NMR测量，但比折叠良好的蛋白质结合更多的染料1-苯胺基萘-8-磺酸盐。
How an amino acid sequence encodes the information necessary for a protein to adopt a unique tertiary structure remains unresolved. We are addressing this problem by designing "from scratch" protein molecules that will adopt predetermined three-dimensional structures. Based on this strategy, two identical four-stranded beta-sheets were designed to dimerize and form a beta-sandwich protein, called betadoublet. A synthetic gene encoding half the beta-sandwich protein was expressed in Escherichia coli, and the protein was purified to homogeneity. Biophysical characterization of betadoublet in aqueous solution demonstrated that the disulfide formed between the two sheets and that the dimer was a compact unaggregated globular protein, consisting predominantly of beta-sheet and stable to thermal denaturation. It has some backbone amide protons whose exchange is slow enough to be measured by NMR but binds more of the dye 1-anilinonaphthalene-8-sulfonate than a well-folded protein.