Chi Ya-Hui, Kumar Thallampuranam Krishnaswamy S, Kathir Karuppanan Muthusamy, Lin Dong-Hai, Zhu Guang, Chiu Ing-Ming, Yu Chin
Department of Chemistry, National Tsing Hua University, Hsinchu 30013, Taiwan.
Biochemistry. 2002 Dec 24;41(51):15350-9. doi: 10.1021/bi026218a.
The conformational stability of the human acidic fibroblast growth factor (hFGF-1) is investigated using amide proton exchange and temperature-dependent chemical shifts, monitored by two-dimensional NMR spectroscopy. The change in free energy of unfolding (DeltaG(u)) of hFGF-1 is estimated to be 5.00 +/- 0.09 kcal.mol(-)(1). Amide proton-exchange rates of 74 residues (in hFGF-1) have been unambiguously measured, and the exchange process occurs predominately according to the conditions of the EX2 limit. The exchange rates of the fast-exchanging amide protons exposed to the solvent have been measured using the clean SEA-HSQC technique. The amide proton protection factor and temperature coefficient estimates show reasonably good correlation. Residues in beta-strands II and VI appear to constitute the stability core of the protein. Among the 12 beta-strands constituting the beta-barrel architecture of hFGF-1, beta-strand XI, located in the heparin binding domain, exhibits the lowest average protection factor value. Amide protons involved in the putative folding nucleation site in hFGF-1, identified by quench-flow NMR studies, do not represent the slow-exchanging core. Residues in portions of hFGF-1 experiencing high conformational flexibility mostly correspond to those involved in receptor recognition and binding.
利用酰胺质子交换和温度依赖化学位移，通过二维核磁共振波谱进行监测，研究了人酸性成纤维细胞生长因子（hFGF-1）的构象稳定性。hFGF-1去折叠的自由能变化（ΔG(u)）估计为5.00±0.09 kcal·mol⁻¹。已明确测量了hFGF-1中74个残基的酰胺质子交换速率，且交换过程主要根据EX2极限条件发生。使用清洁的SEA-HSQC技术测量了暴露于溶剂中的快速交换酰胺质子的交换速率。酰胺质子保护因子和温度系数估计显示出合理的良好相关性。β-链II和VI中的残基似乎构成了蛋白质的稳定性核心。在构成hFGF-1β-桶结构的12条β-链中，位于肝素结合域的β-链XI表现出最低的平均保护因子值。通过淬灭流动核磁共振研究确定的hFGF-1中假定折叠成核位点所涉及的酰胺质子并不代表缓慢交换核心。hFGF-1中经历高构象灵活性的部分残基大多对应于参与受体识别和结合的残基。
