Carter P, Bedouelle H, Winter G
Proc Natl Acad Sci U S A. 1986 Mar;83(5):1189-92. doi: 10.1073/pnas.83.5.1189.
The tyrosyl-tRNA synthetase (EC 6.1.1.1) from Bacillus stearothermophilus is a dimer of two identical subunits. The dimer shows "half-of-the-sites" reactivity in that only one molecule of tyrosyladenylate is formed and one molecule of tRNATyr binds per dimer. To identify whether the tRNATyr binds to a single subunit in the dimer, or to both subunits, heterodimers were constructed by mixing two variant dimers together in 8 M urea. As the unfolded protein is electrophoresed into a native polyacrylamide gel, it refolds and reassociates, and heterodimers can be purified from the parental dimers. Kinetic analysis of heterodimers formed between variant enzymes with defective tyrosine activation or tRNA aminoacylation shows that a molecule of tRNATyr interacts with the N-terminal region of one subunit and the C-terminal region of the other subunit in the dimer.
嗜热脂肪芽孢杆菌的酪氨酰 - tRNA合成酶（EC 6.1.1.1）是由两个相同亚基组成的二聚体。该二聚体表现出“半位点”反应性，即每个二聚体仅形成一分子酪氨酰腺苷酸，且仅结合一分子tRNATyr。为了确定tRNATyr是与二聚体中的单个亚基结合，还是与两个亚基都结合，通过在8 M尿素中将两种变体二聚体混合在一起构建异源二聚体。当未折叠的蛋白质电泳进入天然聚丙烯酰胺凝胶时，它会重新折叠并重新缔合，并且可以从亲本二聚体中纯化出异源二聚体。对具有酪氨酸活化缺陷或tRNA氨酰化缺陷的变体酶之间形成的异源二聚体的动力学分析表明，一分子tRNATyr与二聚体中一个亚基的N端区域和另一个亚基的C端区域相互作用。
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