Contribution of the tyrosines to the structure and function of the human U1A N-terminal RNA binding domain
Contribution of the tyrosines to the structure and function of the human U1A N-terminal RNA binding domain
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DOI：
10.1002/pro.5560050812
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发表时间：
1996-08-01
期刊：
影响因子：
8
通讯作者：
Hall, KB
中科院分区：
文献类型：
--
作者：
Kranz, JK;Lu, JR;Hall, KB
RNA结合结构域（RBD）是共享最小序列保守性但采用α β夹心全局折叠的蛋白质大家族的成员。确定特定氨基酸对RBD结构和RNA结合的贡献对于理解这些蛋白质的功能至关重要。在使用人U1 A N-末端RNA结合结构域（RBD 1）的这些实验中，测量了其四个酪氨酸中的每一个对蛋白质结构、稳定性和RNA结合的贡献。每个酪氨酸被苯丙氨酸和另一个选定的残基取代，并通过化学变性来测量其解折叠自由能，通过将自由能结合到野生型RNA发夹上，以及通过F-19 NMR来探测结构变化来表征所得蛋白质。在这些实验中鉴定的蛋白质的特征包括α-螺旋中可能的酪氨酸/赖氨酸接触，这可能是能量上有利的芳香族/氨基侧链相互作用的实例。蛋白质的一个长环，显示出不寻常的N-15骨架和酪氨酸侧链动力学，与蛋白质：蛋白质缔合有关。RBD 1组织中四个酪氨酸残基的不同相互作用说明了这个蛋白质家族的每个成员如何具有独特的分子细节，有助于功能。
RNA binding domains (RBDs) are members of a large family of proteins that share minimal sequence conservation but adopt an alpha beta sandwich global fold. Defining the contributions of specific amino acids to RBD structure and RNA binding is critical to understanding the functions of these proteins. In these experiments with the human U1A N-terminal RNA binding domain (RBD1), the contributions from each of its four tyrosines to protein structure, stability, and RNA binding were measured. Each tyrosine was substituted with phenylalanine and one other selected residue, and the resulting proteins were characterized by chemical denaturation to measure their unfolding free energy, by binding free energies to the wild-type RNA hairpin, and by F-19 NMR to probe for structural changes. Features of the protein identified in these experiments include a possible tyrosine/lysine contact in an alpha-helix, which may be an example of an energetically favorable aromatic/amino side chain interaction. One long loop of the protein, which shows unusual N-15 backbone and tyrosine side-chain dynamics, is implicated in protein:protein association. The diverse interactions of the four tyrosine residues in the organization of RBD1 illustrate how each member of this family of proteins will have unique molecular details that contribute to function.