Ferguson K M, Higashijima T, Smigel M D, Gilman A G
J Biol Chem. 1986 Jun 5;261(16):7393-9.
Purified guanine nucleotide-binding regulatory proteins, as either the oligomers or the isolated nucleotide-binding alpha subunits, display anomalous kinetics of nucleotide binding. This is due to the presence of tightly bound GDP in these preparations. The dissociation of bound GDP is the rate-limiting step for nucleotide binding. GDP can be removed by chromatography in the presence of 1 M (NH4)2SO4 and 20% glycerol, which yields preparations of G proteins that contain less than 0.1 mol of GDP/mol of guanosine 5'-(gamma-thio)triphosphate (GTP gamma S)-binding site. When the GDP is removed, the binding of GTP gamma S displays kinetics consistent with a bimolecular reaction.
纯化的鸟嘌呤核苷酸结合调节蛋白，无论是寡聚体还是分离出的核苷酸结合α亚基，都表现出核苷酸结合的异常动力学。这是由于这些制剂中存在紧密结合的GDP。结合的GDP的解离是核苷酸结合的限速步骤。在1 M硫酸铵和20%甘油存在下，通过色谱法可以去除GDP，从而得到每摩尔鸟苷5'-（γ-硫代）三磷酸（GTPγS）结合位点所含GDP少于0.1摩尔的G蛋白制剂。当去除GDP后，GTPγS的结合表现出与双分子反应一致的动力学。
