Resch Marcus, Striegl Harald, Henssler Eva Maria, Sevvana Madhumati, Egerer-Sieber Claudia, Schiltz Emile, Hillen Wolfgang, Muller Yves A
Lehrstuhl für Biotechnik, Department of Biology, Friedrich-Alexander University Erlangen-Nuremberg, Henkestrasse 91 and Staudtstrasse 5, D-91052 Erlangen, Germany.
Nucleic Acids Res. 2008 Aug;36(13):4390-401. doi: 10.1093/nar/gkn400. Epub 2008 Jun 28.
Today's proteome is the result of innumerous gene duplication, mutagenesis, drift and selection processes. Whereas random mutagenesis introduces predominantly only gradual changes in protein function, a case can be made that an abrupt switch in function caused by single amino acid substitutions will not only considerably further evolution but might constitute a prerequisite for the appearance of novel functionalities for which no promiscuous protein intermediates can be envisaged. Recently, tetracycline repressor (TetR) variants were identified in which binding of tetracycline triggers the repressor to associate with and not to dissociate from the operator DNA as in wild-type TetR. We investigated the origin of this activity reversal by limited proteolysis, CD spectroscopy and X-ray crystallography. We show that the TetR mutant Leu17Gly switches its function via a disorder-order mechanism that differs completely from the allosteric mechanism of wild-type TetR. Our study emphasizes how single point mutations can engender unexpected leaps in protein function thus enabling the appearance of new functionalities in proteins without the need for promiscuous intermediates.
如今的蛋白质组是无数基因复制、诱变、漂变和选择过程的结果。随机诱变主要只会在蛋白质功能上引入渐进性变化，然而，可以提出这样一种观点，即由单个氨基酸取代导致的功能突然转变不仅会极大地推动进一步进化，而且可能是出现新功能的先决条件，而对于这些新功能，无法设想存在混杂的蛋白质中间体。最近，人们鉴定出了四环素阻遏物（TetR）变体，其中四环素的结合会触发阻遏物与操纵子DNA结合，而不像野生型TetR那样从操纵子DNA上解离。我们通过有限蛋白酶解、圆二色光谱和X射线晶体学研究了这种活性逆转的起源。我们表明，TetR突变体Leu17Gly通过一种无序-有序机制切换其功能，该机制与野生型TetR的变构机制完全不同。我们的研究强调了单点突变如何能够在蛋白质功能上产生意想不到的飞跃，从而使蛋白质在无需混杂中间体的情况下出现新功能。
