Burbaum J J, Starzyk R M, Schimmel P
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Proteins. 1990;7(2):99-111. doi: 10.1002/prot.340070202.
The locations of functionally important sequences and general structural motifs have been assigned to Ile-tRNA synthetase. However, a function has not been established for some segments of the protein (e.g., CP1). The method of structural modeling described here cannot establish the details of a 3 A crystal structure, and, in contrast to a crystal structure, the precision of the model varies according to the extent of a sequence similarity or the functional importance of a region. In Ile-tRNA synthetase, the signature sequence and the flanking regions are likely to be similar in structure to the proteins on which the model is based. For other regions, it may be possible to build a three-dimensional model by connecting well defined regions and refining the positions of the connecting elements by energy minimization. Structural modelling of this kind must be done cautiously, because the order and orientation of the elements of a structural motif can change in subtle ways. In the case of Tyr-tRNA synthetase, the beta-strand nearest the N-terminus is the outermost strand of the nucleotide binding fold; in Met-tRNA synthetase, the same strand is innermost. Furthermore, the orientation of this strand may be antiparallel (Tyr-tRNA synthetase) or parallel (Met-tRNA synthetase). Because multiple structures that differ in their orientations of structural elements are possible, the structural analogies between proteins should not be naively extrapolated without independent experimental support. As described above, some regions of proteins tolerate internal deletions and insertions. This provides further experimental support for the practice of allowing for gaps in computer-generated sequence alignments. Nevertheless, because some regions are more tolerant of insertions and deletions than others, the structural and functional significance of a region of broken alignment must be assessed carefully. All gaps in sequence alignments cannot be treated equally, and each must be evaluated within its own context. In the synthetases of known structure, structural analogy can be used to identify important functional elements. For example, the amino acid binding site of Met-tRNA synthetase might be formed, at least in part, by a peptide that encompasses Ala50; this amino acid aligns with Gly94 of the Ile-tRNA synthetase. This is an example in which results on a protein of unknown structure (Ile-tRNA synthetases) can lead to identification of a potential substrate binding site in a protein of known structure (Met-tRNA synthetase).
功能重要序列和一般结构基序的位置已被确定在异亮氨酸 - tRNA合成酶上。然而，该蛋白质的某些片段（例如CP1）的功能尚未明确。这里描述的结构建模方法无法确定3埃晶体结构的细节，并且与晶体结构不同，模型的精度会根据序列相似性的程度或区域的功能重要性而有所不同。在异亮氨酸 - tRNA合成酶中，特征序列及其侧翼区域在结构上可能与构建模型所依据的蛋白质相似。对于其他区域，可以通过连接定义明确的区域并通过能量最小化来优化连接元件的位置，从而构建三维模型。这种结构建模必须谨慎进行，因为结构基序的元件顺序和方向可能会以微妙的方式发生变化。例如，在酪氨酸 - tRNA合成酶中，最靠近N端的β链是核苷酸结合折叠的最外层链；在甲硫氨酸 - tRNA合成酶中，同一条链是最内层的。此外，这条链的方向可能是反平行的（酪氨酸 - tRNA合成酶）或平行的（甲硫氨酸 - tRNA合成酶）。由于蛋白质结构元件方向不同的多种结构都是可能的，因此在没有独立实验支持的情况下，不应盲目推断蛋白质之间的结构相似性。如上所述，蛋白质的某些区域能够耐受内部缺失和插入。这为在计算机生成的序列比对中允许出现空位的做法提供了进一步的实验支持。然而，由于某些区域比其他区域更能耐受插入和缺失，因此必须仔细评估比对中断裂区域的结构和功能意义。序列比对中的所有空位不能同等对待，每个空位都必须在其自身的背景下进行评估。在已知结构的合成酶中，结构相似性可用于识别重要的功能元件。例如，甲硫氨酸 - tRNA合成酶的氨基酸结合位点可能至少部分由包含丙氨酸50的肽段形成；该氨基酸与异亮氨酸 - tRNA合成酶的甘氨酸94对齐。这是一个例子，即对未知结构蛋白质（异亮氨酸 - tRNA合成酶）的研究结果能够导致识别已知结构蛋白质（甲硫氨酸 - tRNA合成酶）中潜在的底物结合位点。
