Interactions within the yeast Sm core complex:: from proteins to amino acids
Interactions within the yeast Sm core complex:: from proteins to amino acids
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DOI：
10.1128/mcb.18.4.1956
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发表时间：
1998-04-01
影响因子：
5.3
通讯作者：
Bordonné, R
中科院分区：
文献类型：
--
作者：
Camasses, A;Bragado-Nilsson, E;Bordonné, R
Sm 核心蛋白通过与小的不清楚的 RNA 结合并参与蛋白质相互作用网络，在小核核糖核蛋白颗粒 (snRNP) 的形成中发挥重要作用。双杂交系统用于识别 SmE 相互作用蛋白并测试酵母 Sm 蛋白的所有成对组合之间的相互作用。我们观察到 SmB 和 SmD3、SmE 和 SmF、SmE 和 SmG 之间的相互作用。对于这些相互作用，直接生化测定证实了体内获得的结果的有效性。为了绘制 Sm 蛋白的蛋白质-蛋白质相互作用表面图，我们生成了 SmE 突变体文库，并研究了它们在双杂交系统中与 SmF 和/或 SmG 蛋白相互作用的能力。观察到几类突变体：一些突变体不能与 SmF 或 SmG 蛋白相互作用，同样与 SmG 相互作用但不与 SmF 相互作用，而其他突变体与 SmF 适度相互作用但不与 SmG 相互作用。我们对酵母 SmE 蛋白的突变分析表明，保守的疏水残基对于与 SmF 和 SmG 的相互作用以及活力至关重要。令人惊讶的是，我们观察到其他进化上保守的位置能够耐受突变，替换仅轻微影响与 SmF 和 SmG 的结合，并赋予野生型生长表型。
Sm core proteins play an essential role in the formation of small nuclear ribonucleoprotein particles (snRNPs) by binding to small unclear RNAs and participating in a network of protein interactions, The two-hybrid system was used to identify SmE interacting proteins and to test for interactions between ail pairwise combinations of yeast Sm proteins. We observed interactions between SmB and SmD3, SmE and SmF, and SmE, and SmG. For these interactions, a direct biochemical assay confirmed the validity of the results obtained in vivo., To map the protein-protein interaction surface of Sm proteins, we generated a library of SmE mutants and investigated their ability to interact with SmF and/or SmG proteins in the two-hybrid system. Several classes of mutants were observed: some mutants are unable to interact with either SmF or SmG proteins, same interact with SmG but not with SmF, while others interact moderately viith SmF but not with SmG. Our mutational analysis of yeast SmE protein shows that conserved hydrophobic residues are essential for interactions with SmF and SmG as well as for viability. Surprisingly, we observed that other evolutionarily conserved positions are tolerant to mutations, with substitutions affecting binding to SmF and SmG only mildly and conferring a wild-type growth phenotype.