Hopper Jonathan T S, Oldham Neil J
School of Chemistry, University of Nottingham, University Park, Nottingham, United Kingdom.
J Am Soc Mass Spectrom. 2009 Oct;20(10):1851-8. doi: 10.1016/j.jasms.2009.06.010. Epub 2009 Jul 1.
Ion mobility spectrometry, with subsequent mass spectrometric detection, has been employed to study the stability of compact protein conformations of FK-binding protein, hen egg-white lysozyme, and horse heart myoglobin in the presence and absence of bound ligands. Protein ions, generated by electrospray ionization from ammonium acetate buffer, were activated by collision with argon gas to induce unfolding of their compact structures. The collisional cross sections (Omega) of folded and unfolded conformations were measured in the T-Wave mobility cell of a Waters Synapt HDMS (Waters, Altrincham, UK) employing a calibration against literature values for a range of protein standards. In the absence of activation, collisional cross section measurements were found to be consistent with those predicted for folded protein structures. Under conditions of defined collisional activation energies partially unfolded conformations were produced. The degree of unfolding and dissociation induced by these defined collision energies are related to the stability of noncovalent intra- and intermolecular interactions within protein complexes. These findings highlight the additional conformational stability of protein ions in the gas phase resulting from ligand binding.
离子淌度光谱法结合后续的质谱检测，已被用于研究FK结合蛋白、鸡蛋清溶菌酶和马心肌红蛋白在存在和不存在结合配体的情况下紧密蛋白构象的稳定性。通过电喷雾电离从醋酸铵缓冲液中产生的蛋白质离子，与氩气碰撞而被激活，以诱导其紧密结构的展开。在沃特世Synapt HDMS（英国奥尔特林厄姆的沃特世公司）的T波淌度池中，针对一系列蛋白质标准品的文献值进行校准，测量折叠和未折叠构象的碰撞截面（Ω）。在没有激活的情况下，发现碰撞截面测量结果与预测的折叠蛋白结构一致。在确定的碰撞激活能量条件下，产生了部分未折叠的构象。这些确定的碰撞能量诱导的展开和解离程度与蛋白质复合物中非共价分子内和分子间相互作用的稳定性有关。这些发现突出了配体结合导致的气相中蛋白质离子额外的构象稳定性。
