Characterization of specific protein association by 15N CPMG relaxation dispersion NMR:: The GB1A34F monomer-dimer equilibrium
Characterization of specific protein association by 15N CPMG relaxation dispersion NMR:: The GB1A34F monomer-dimer equilibrium
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DOI：
10.1021/jp076094h
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发表时间：
2008-05-15
影响因子：
3.3
通讯作者：
Gronenborn, Angela M.
中科院分区：
文献类型：
--
作者：
Jee, JunGoo;Ishima, Rieko;Gronenborn, Angela M.
Carr-Purcell-Meiom-Gill(CPMG)横向弛豫色散核磁共振实验是检测和表征构象交换的有力手段。这个实验报告了化学位移的交换，因此可以监测所有的化学交换现象，而不仅仅是分子内的构象交换。在这里，我们报道了对GB1点突变Ala-34-Phe(GB1(A34F))单体-二聚体平衡的CPMG横向弛豫色散研究。这种变体在高浓度(>；1 mM)时主要以并排二聚体的形式存在。我们证明了色散实验对于研究缔合平衡是特别有价值的，因为它对平衡中的少数群体非常敏感。通过2D N-15-H-1HSQC光谱监测了GB1(A34F)二聚体蛋白中的28个单独的酰胺位点，所有松弛得到的数据都与二聚体和单体物种之间的交换过程基本一致。
The Carr-Purcell-Meiboom-Gill (CPMG) transverse relaxation dispersion NMR experiment is a powerful means for detecting and characterizing conformational exchange. This experiment reports the exchange of chemical shifts and therefore can monitor all chemical exchange phenomena, not only intramolecular conformational exchange. Here, we report a CPMG transverse relaxation dispersion study for the monomer-dimer equilibrium of the GB1 point mutant, Ala-34-Phe (GB1(A34F)). This variant exists predominantly as a side-by-side dimer at high concentration (> 1 mM). We demonstrate that the dispersion experiment is exceptionally valuable for studying association equilibria since it is extremely sensitive to the minor population in the equilibrium. Twenty-eight individual amide sites in the GB1(A34F) dimer protein, were monitored via a 2D N-15-H-1 HSQC spectroscopy, and all relaxation-derived data are consistent with predominantly an exchange process between dimer and monomer species.