Bachmair A, Finley D, Varshavsky A
Science. 1986 Oct 10;234(4773):179-86. doi: 10.1126/science.3018930.
When a chimeric gene encoding a ubiquitin-beta-galactosidase fusion protein is expressed in the yeast Saccharomyces cerevisiae, ubiquitin is cleaved off the nascent fusion protein, yielding a deubiquitinated beta-galactosidase (beta gal). With one exception, this cleavage takes place regardless of the nature of the amino acid residue of beta gal at the ubiquitin-beta gal junction, thereby making it possible to expose different residues at the amino-termini of the otherwise identical beta gal proteins. The beta gal proteins thus designed have strikingly different half-lives in vivo, from more than 20 hours to less than 3 minutes, depending on the nature of the amino acid at the amino-terminus of beta gal. The set of individual amino acids can thus be ordered with respect to the half-lives that they confer on beta gal when present at its amino-terminus (the "N-end rule"). The currently known amino-terminal residues in long-lived, noncompartmentalized intracellular proteins from both prokaryotes and eukaryotes belong exclusively to the stabilizing class as predicted by the N-end rule. The function of the previously described posttranslational addition of single amino acids to protein amino-termini may also be accounted for by the N-end rule. Thus the recognition of an amino-terminal residue in a protein may mediate both the metabolic stability of the protein and the potential for regulation of its stability.
当编码泛素-β-半乳糖苷酶融合蛋白的嵌合基因在酿酒酵母中表达时，泛素会从新生的融合蛋白上被切割下来，产生去泛素化的β-半乳糖苷酶（β-gal）。除了一个例外情况，无论β-gal在泛素-β-gal连接点处的氨基酸残基性质如何，这种切割都会发生，从而使得在原本相同的β-gal蛋白的氨基末端能够暴露不同的残基。如此设计的β-gal蛋白在体内具有显著不同的半衰期，从超过20小时到少于3分钟不等，这取决于β-gal氨基末端的氨基酸性质。因此，就单个氨基酸在β-gal氨基末端（“N端规则”）存在时赋予β-gal的半衰期而言，可以对这些氨基酸进行排序。目前已知的来自原核生物和真核生物的长寿命、非区室化细胞内蛋白质中的氨基末端残基，完全属于N端规则预测的稳定类别。先前描述的对蛋白质氨基末端进行单个氨基酸的翻译后添加的功能，也可能由N端规则来解释。因此，对蛋白质中氨基末端残基的识别可能介导了蛋白质的代谢稳定性及其稳定性调节的潜力。
