Straub Kristina, Merkl Rainer
Institute of Biophysics and Physical Biochemistry, University of Regensburg, Regensburg, Germany.
Methods Mol Biol. 2019;1851:171-182. doi: 10.1007/978-1-4939-8736-8_9.
Ancestral sequence reconstruction (ASR) is a powerful tool to infer primordial sequences from contemporary, i.e., extant ones. An essential element of ASR is the computation of a phylogenetic tree whose leaves are the chosen extant sequences. Most often, the reconstructed sequence related to the root of this tree is of greatest interest: It represents the common ancestor (CA) of the sequences under study. If this sequence encodes a protein, one can "resurrect" the CA by means of gene synthesis technology and study biochemical properties of this extinct predecessor with the help of wet-lab experiments.However, ASR deduces also sequences for all internal nodes of the tree, and the well-considered analysis of these "intermediates" can help to elucidate evolutionary processes. Moreover, one can identify key mutations that alter proteins or protein complexes and are responsible for the differing properties of extant proteins. As an illustrative example, we describe the protocol for the rapid identification of hotspots determining the binding of the two subunits within the heteromeric complex imidazole glycerol phosphate synthase.
祖先序列重建（ASR）是一种从当代（即现存）序列推断原始序列的强大工具。ASR的一个基本要素是计算一个系统发育树，其叶节点是所选的现存序列。最常被关注的是与该树的根相关的重建序列：它代表了所研究序列的共同祖先（CA）。如果该序列编码一种蛋白质，人们可以通过基因合成技术“复活”这个共同祖先，并借助湿实验室实验研究这种已灭绝的前身的生化特性。然而，ASR还能推断出树中所有内部节点的序列，对这些“中间序列”进行周全的分析有助于阐明进化过程。此外，人们可以识别那些改变蛋白质或蛋白质复合物并导致现存蛋白质具有不同特性的关键突变。作为一个示例，我们描述了快速识别决定异源复合物咪唑甘油磷酸合酶中两个亚基结合的热点的方案。
