Chen J M, Brandt-Rauf P W, Pincus M R
Dupont Agricultural Products, Stein-Haskell Research Center, Newark, DE 19714, USA.
J Biomol Struct Dyn. 1996 Jun;13(6):925-33. doi: 10.1080/07391102.1996.10508907.
rap-1A is a membrane-bound G-protein in the ras superfamily that, like the ras-p21 protein, is activated by binding GTP in place of GDP. When activated, however, this protein inhibits the action of ras-p21, which is to induce mitogenesis in cells A chimeric protein containing RAS-p21 residues 1-65 and rap-1A residues 66-184 becomes ras-p21-like in its activity. The critical changes in sequence that result in this transformation are G26N, 127H, E30D, K31E, and E45V. All of these substitutions occur in or around a critical effector domain of p21 that is involved in interacting with GTPase activating protein (GAP), raf-p74 protein and inositol-3-hydroxy kinase. Using molecular dynamics, we have computed the average low energy structures for each of the three proteins, ras-p21, rap-1A and mutant rap1A, called rap-M, that contains these critical amino acid substitutions. We find that rap-M more closely superimposes on ras-p21 (rms deviation 1.9 A) than on wild-type rap-1A (rms deviation 3.4 A). In particular, the amino terminal domains (residues 3-59) of both RAS-p21 and rap-M are superimposable while they deviate when the average structures of these two proteins are superimposed on that of wild-type rap-1A. We have identified Pro 34 as a critical residue which may determine if the protein transforms cells or inhibits cell transformation. In addition, we have found that ras-p21 and rap-M proteins are superimposable in the region 96-110 except at Asp 105. The 96-110 domain of ras-p21 has been found to be involved in the binding of this protein to the nuclear transcription protein, jun and its kinase, jun kinase, JNK. Both segments differ in structure from that of the rap-1A segment at Asp 108, implicating this residue as also being important in determining the activity of the protein. Overall, the oncogenic substitutions introduced into the rap-1A protein cause it to adopt a conformation that is very similar to that of ras-p21 rather than wild-type rap-1A.
Rap-1A是Ras超家族中的一种膜结合G蛋白，与Ras-p21蛋白一样，通过结合GTP取代GDP而被激活。然而，激活后，这种蛋白会抑制Ras-p21的作用，Ras-p21的作用是诱导细胞有丝分裂。一种包含RAS-p21第1至65位残基和rap-1A第66至184位残基的嵌合蛋白在活性上变得类似于Ras-p21。导致这种转变的关键序列变化是G26N、127H、E30D、K31E和E45V。所有这些取代都发生在p21的一个关键效应结构域内或其周围，该结构域参与与GTP酶激活蛋白（GAP）、raf-p74蛋白和肌醇-3-羟基激酶相互作用。利用分子动力学，我们计算了三种蛋白Ras-p21、rap-1A和含有这些关键氨基酸取代的突变型rap1A（称为rap-M）各自的平均低能结构。我们发现rap-M与Ras-p21的重叠程度更高（均方根偏差为1.9 Å），而与野生型rap-1A的重叠程度较低（均方根偏差为3.4 Å）。特别是，RAS-p21和rap-M的氨基末端结构域（第3至59位残基）是可重叠的，而当将这两种蛋白的平均结构与野生型rap-1A的结构重叠时，它们会出现偏差。我们已确定Pro 34是一个关键残基，它可能决定该蛋白是转化细胞还是抑制细胞转化。此外，我们发现Ras-p21和rap-M蛋白在96 - 110区域除了Asp 105处是可重叠的。已发现Ras-p21的96 - 110结构域参与该蛋白与核转录蛋白jun及其激酶jun激酶JNK的结合。这两个片段在Asp 108处的结构与rap-1A片段的结构不同，这表明该残基在决定蛋白活性方面也很重要。总体而言，引入rap-1A蛋白的致癌取代使其采用了一种与Ras-p21非常相似而非野生型rap-1A的构象。
