COMPUTER MODELING OF AN ATP-BINDING PROTEIN

ATP 结合蛋白的计算机建模

• 批准号: 7723521
• 负责人: ANDREW POHORILLE
• 金额: $ 0.58万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7723521
• 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; Numbers; Pattern; Phenylalanine; Protein Databases; Protein Family; Proteins; Research; Research Personnel; Resolution; Resources; Side; Source; Specificity; Standards of Weights and Measures; Techniques; Tyrosine; United States National Institutes of Health; Water; X-Ray Crystallography; Zinc; alpha helix; analog; beta pleated sheet; interest; novel; pointed protein; polypeptide; pressure; protein structure; size; 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利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 水溶性蛋白质的起源似乎比膜结合蛋白的起源要难得多。大多数跨膜蛋白，即使是那些功能和结构复杂的蛋白，都是由少数蛋白质家族共享的结构元件组成的。此外，有简单的、自然的或合成的模型，由相同的元素组成，可以执行基本的膜相关功能。细胞质蛋白质的情况并非如此。在当代细胞中，按照原生生物标准，它们通常相当大，但与膜蛋白相比，它们的大小无法在不丧失活性的情况下显著缩小。 使用一种新的体外技术，Keefe和Szostak从6万亿个随机多肽中选择了ATP结合蛋白。他们发现了四个新的蛋白质家族，每个家族都包含氨基酸序列高度相似的蛋白质，这些蛋白质彼此之间或与当前蛋白质数据库中发现的任何东西都没有关联。发现ATP结合蛋白的频率似乎是 与发现ATP结合核酶的频率相似。 一个家族的蛋白质已经得到了相当详细的表征。最初选择的蛋白质含有80个氨基酸，但缺失研究表明，最小结合单位长度不到50个氨基酸，因此是已知的最小的ATP结合蛋白。这些蛋白质对三磷酸腺苷及其受体具有高度选择性 紧密的类似物，二磷酸腺苷(ADP)，因为它们既不与鸟苷三磷酸(GTP)结合，也不与环状AMP结合。然而，它们的序列不包含任何已发现的ATP结合基序。要发挥作用，它们需要锌离子，并含有四个保守的半胱氨酸残基。最近，该家族的一种蛋白质的高分辨率三维结构被用X射线结晶学解决。与所有生物、水溶性蛋白质一样，这种结构具有疏水核心，但呈现出新的折叠。它由一个三股的 反平行的β-折叠和两个不相邻的α-螺旋。ADP稳定在 通过堆积与苯丙氨酸和酪氨酸残基的相互作用以及通过氢键与蛋白质中的几个侧链形成结合口袋。结合的选择性似乎受腺嘌呤的N1、N3和N6与甲氨酸-45和甘氨酸-63之间的氢键的保证。锌离子在不与结合口袋相邻的区域被保守的半胱氨酸配位。 ATP结合蛋白是一个非常有趣的原生生物模型，因为它是第一个简单的、功能蛋白质的例子，而这种蛋白质并没有长期进化优化的主题。然而，它的折叠模式可能在进化上存在缺陷。例如，它可能没有通过突变获得新的特异性的能力。我们建议从这个角度来研究蛋白质，如果需要的话，重新设计它的序列，试图在不改变折叠的情况下消除这些缺陷。如果这项任务成功，它将导致一个看起来适合进化的新折叠的产生，从而提供了一个经验性的论点支持 关于酶起源的“进化意外”假说。如果我们发现序列不能被适当地重新设计，这将表明，即使该折叠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。尽管从一个单一的负面例子得出结论显然为时过早，但这一结果将暗示，关于蛋白质结构进化修剪的假说值得认真考虑。在任何一种情况下，我们都将了解如何为进化上可行的原生酶构建和识别良好的候选模型。