COMPUTER MODELING OF AN ATP-BINDING PROTEIN

ATP 结合蛋白的计算机建模

• 批准号: 8170538
• 负责人: ANDREW POHORILLE
• 金额: $ 0.71万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8170538
• 关键词: Accidents; Adenine; Adenosine Diphosphate; Amino Acid Sequence; Amino Acids; Binding; Binding Proteins; Biological; Catalytic RNA; Cells; Complex; Computer Retrieval of Information on Scientific Projects Database; Computer Simulation; Cyclic AMP; Cysteine; Cytoplasmic Protein; Elements; Enzymes; Evolution; Exhibits; Family; Frequencies; Funding; Glycine; Grant; Guanosine Triphosphate; Hydrogen Bonding; In Vitro; Institution; Integral Membrane Protein; Ions; Lead; Membrane; Membrane Proteins; Modeling; Mutation; Pattern; Phenylalanine; Protein Databases; Protein Family; Proteins; Research; Research Personnel; Resolution; Resources; Side; Source; Specificity; Techniques; Tyrosine; United States National Institutes of Health; Water; X-Ray Crystallography; Zinc; alpha helix; analog; beta pleated sheet; interest; novel; pointed protein; polypeptide; premature; pressure; protein structure; three dimensional structure

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The origins of water-soluble proteins appear to be considerably more difficult to identify than the origins of membrane-bound proteins. Most transmembrane proteins, even those that are functionally and structurally complex, are built of a small number of structural elements that are shared across protein families. Moreover, there are simple, natural or synthetic models consisting of the same elements that can perform essential membrane-related functions. This is not the case with cytoplasmic proteins. In contemporary cells, they are usually quite large by protobiological standards, but in contrast to membrane proteins they cannot be significantly reduced in size without loss of activity. Using a novel in vitro technique, Keefe and Szostak selected ATP-binding proteins from six trillion random polypeptides. They found four new protein families, each containing proteins with highly similar amino acid sequences that were unrelated to each other or to anything found in the current protein databases. The frequency of finding ATP-binding proteins appears to be similar to the frequency of finding ATP-binding ribozymes. Proteins from one family have been characterized in fair detail. The originally selected protein contained 80 amino acids but deletion studies revealed that the minimal binding unit is less than 50 amino acids long and, thus, is the smallest known ATP-binding protein. The proteins are highly selective towards ATP and its close analog, adenosine diphosphate (ADP), as they bind neither guanosine triphosphate (GTP) nor cyclic AMP. However, their sequences do not contain any already identified ATP-binding motifs. To function, they require zinc ions and contain four conserved cysteine residues. More recently, the high resolution, three-dimensional structure of a protein from the family was solved using X-ray crystallography. As all biological, water-soluble proteins, this structure has a hydrophobic core, but exhibits a novel fold. It consists of a three-stranded antiparallel beta-sheet and two nonadjacent alpha-helices. ADP is stabilized in the binding pocket by stacking interactions with phenylalanine and tyrosine residues and by hydrogen bonds to several side chains in the protein. Selectivity of binding appears to be insured by hydrogen bonds between the N1, N3 and N6 of adenine and methianine-45 and glycine-63. A zinc ion is coordinated by the conserved cysteines in a region not adjacent to the binding pocket. The ATP-binding protein is a very interesting protobiological model because it is the first example of a simple, functional protein that has not been a subject of long evolutionary optimization. However, its folding pattern may be evolutionarily deficient. For example, it may not have the capability to acquire new specificity through mutations. We propose to examine the protein from this point of view and, if necessary, redesign its sequence in an attempt to eliminate the deficiencies without altering the fold. If this task were successful it would lead to the creation of a novel fold that appears to be suitable for evolution, thus providing an empirical argument supporting an "evolutionary accident" hypothesis of the origin of enzymes. If we found that the sequence could not be appropriately redesigned it would suggest that the fold, even if it were present among protobiological proteins, was not likely to survive subsequent evolutionary pressures. Although it would be clearly premature to draw conclusions from a single negative example, this result would hint that a hypothesis about evolutionary pruning of protein structures is worth serious considerations. In either case, we would gain an understanding how to construct and identify good candidate models for evolutionarily viable protobiological enzymes.

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 水溶性蛋白质的起源似乎比膜结合蛋白质的起源更难识别。大多数跨膜蛋白，即使是那些功能和结构复杂的跨膜蛋白，也是由少数跨蛋白质家族共享的结构元件组成的。此外，有简单的，天然的或合成的模型，由相同的元素组成，可以执行基本的膜相关功能。这不是细胞质蛋白的情况。在现代细胞中，它们通常以原始生物学标准来看相当大，但与膜蛋白相比，它们不能在不丧失活性的情况下显著减小尺寸。 Keefe和Szostak使用一种新的体外技术，从6万亿随机多肽中选择了ATP结合蛋白。他们发现了四个新的蛋白质家族，每个家族都包含具有高度相似氨基酸序列的蛋白质，这些蛋白质彼此无关，也与当前蛋白质数据库中发现的任何蛋白质无关。发现ATP结合蛋白的频率似乎 类似于发现ATP结合核酶的频率。 来自一个家族的蛋白质已经被相当详细地表征。最初选择的蛋白质含有80个氨基酸，但缺失研究显示，最小结合单位长度小于50个氨基酸，因此是已知的最小ATP结合蛋白。这些蛋白质对ATP及其受体具有高度选择性。 类似物，腺苷二磷酸（ADP），因为它们既不结合鸟苷三磷酸（GTP），也不结合环AMP。然而，它们的序列不包含任何已经鉴定的ATP结合基序。为了发挥作用，它们需要锌离子，并含有四个保守的半胱氨酸残基。最近，使用X射线晶体学解决了该家族蛋白质的高分辨率三维结构。与所有生物的水溶性蛋白质一样，这种结构具有疏水核心，但表现出新的折叠。它由一个三股 反平行β折叠和两个不相邻的α螺旋。ADP稳定在 通过与苯丙氨酸和酪氨酸残基的堆积相互作用以及通过与蛋白质中的几个侧链的氢键来形成结合口袋。结合的选择性似乎是由腺嘌呤和蛋氨酸-45和甘氨酸-63的N1、N3和N6之间的氢键保证的。锌离子由不邻近结合口袋的区域中的保守半胱氨酸配位。 ATP结合蛋白是一个非常有趣的原生物学模型，因为它是第一个简单的、功能性蛋白质的例子，而这种蛋白质没有经过长期的进化优化。然而，它的折叠模式可能是进化上的缺陷。例如，它可能没有能力通过突变获得新的特异性。我们建议从这个角度来研究蛋白质，如果必要的话，重新设计其序列，试图在不改变折叠的情况下消除缺陷。如果这项任务是成功的，它将导致创造一个新的折叠，似乎是适合于进化，从而提供了一个实证论据，支持 关于酶起源的“进化偶然”假说。如果我们发现序列不能被适当地重新设计，这将表明折叠，即使它存在于原始生物蛋白质中，也不太可能在随后的进化压力中幸存下来。虽然从一个单一的负面例子得出结论显然还为时过早，但这一结果表明，关于蛋白质结构进化修剪的假设值得认真考虑。在任何一种情况下，我们都将了解如何构建和识别进化上可行的原生生物酶的良好候选模型。