Artificial di-iron proteins: solution characterization of four helix bundles containing two distinct types of inter-helical loops
Artificial di-iron proteins: solution characterization of four helix bundles containing two distinct types of inter-helical loops
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DOI：
10.1007/s00775-005-0002-8
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发表时间：
2005-08-01
影响因子：
3
通讯作者：
Lombardi, A
中科院分区：
文献类型：
--
作者：
Maglio, O;Nastri, F;Lombardi, A
基于肽的模型对金属蛋白模型的发展有着巨大的影响，因为它们似乎是模拟天然系统的结构特征和反应性的合适候选者。通过从头设计的四螺旋束，我们开发了DF（Due Ferri）家族的人工蛋白质，作为模型的二铁和二锰金属蛋白。我们的研究目标是阐明静电环境，极性和溶剂的金属结合位点的可及性，影响二铁蛋白的功能特性。DF蛋白家族的前两个子集DF 1和DF 2由两个非共价缔合的螺旋-环-螺旋基序组成，其在结构的中心附近结合双金属辅因子。DF 2子集被设计为改善DF 1的性质：DF 2和DF 2 t在它们的序列中有几个变化，以改善溶解度和金属离子进入，以及连接两个螺旋的环的变化。为了评估这些变化如何影响模型蛋白质的整体结构，我们解决了DF 2和DF 2 t的二-Zn（II）配合物的NMR结构，并将这些结构与最近从X射线晶体学获得的结构进行比较。此外，我们研究了与突变相关的热力学后果，通过测量DF 2 t在不同金属离子存在下的稳定性，并将结果与DF 2已经获得的数据进行比较。综上所述，所有数据的分析显示了转角构象在四螺旋束的设计和稳定性中的重要性。
Peptide-based models have an enormous impact for the development of metalloprotein models, as they seem appropriate candidates to mimic both the structural characteristics and reactivity of the natural systems. Through the de novo design of four-helix bundles, we developed the DF (Due Ferri) family of artificial proteins, as models of di-iron and di-manganese metalloproteins. The goal of our research is to elucidate how the electrostatic environment, polarity and solvent accessibility of the metal-binding site, influence the functional properties of di-iron proteins. The first two subsets of the DF protein family, DF1 and DF2, consist of two non-covalently associated helix-loop-helix motifs, which bind the di-metal cofactor near the center of the structure. The DF2 subset was designed to improve the properties of DF1: DF2 and DF2t have several changes in their sequences to improve solubility and metal ion access, as well as a change in the loop connecting the two helices. In order to evaluate how these changes affect the overall structure of the model proteins, we solved the NMR structures of the di-Zn(II) complexes of DF2 and DF2t, and compared these structures with those recently obtained from X-ray crystallography. Further, we examined the thermodynamic consequences associated with the mutations, by measuring the stability of DF2t in the presence of different metal ions, and comparing the results with the data already obtained for DF2. Taken together, analysis of all the data showed the importance of the turn conformation in the design and stability of four-helix bundle.