Horvath L A, Sturtevant J M, Prestegard J H
Department of Chemistry, Yale University, New Haven, Connecticut 06511.
Protein Sci. 1994 Jan;3(1):103-8. doi: 10.1002/pro.5560030113.
The denaturation of Escherichia coli acyl carrier protein (ACP) in buffers containing both monovalent and divalent cations was followed by variable-temperature NMR and differential scanning calorimetry. Both high concentrations of monovalent salts (Na+) and moderate concentrations of divalent salts (Ca2+) raise the denaturation temperature, but calorimetry indicates that a significant increase in the enthalpy of denaturation is obtained only with the addition of a divalent salt. NMR experiments in both low ionic strength monovalent buffers and low ionic strength monovalent buffers containing calcium ions show exchange between native and denatured forms to be slow on the NMR time scale. However, in high ionic strength monovalent buffers, where the temperature of denaturation is elevated as it is in the presence of Ca2+, the transition is fast on the NMR time scale. These results suggest that monovalent and divalent cations may act to stabilize ACP in different ways. Monovalent ions may nonspecifically balance the intrinsic negative charge of this protein in a way that is similar for native, denatured, and intermediate forms. Divalent cations provide stability by binding to specific sites present only in the native state.
利用变温核磁共振（NMR）和差示扫描量热法，研究了大肠杆菌酰基载体蛋白（ACP）在同时含有单价阳离子和二价阳离子的缓冲液中的变性情况。高浓度的单价盐（Na+）和中等浓度的二价盐（Ca2+）都会提高变性温度，但量热法表明，只有加入二价盐才能使变性焓显著增加。在低离子强度的单价缓冲液和含有钙离子的低离子强度单价缓冲液中进行的NMR实验表明，在NMR时间尺度上，天然形式和变性形式之间的交换很慢。然而，在高离子强度的单价缓冲液中，变性温度如在Ca2+存在时一样升高，在NMR时间尺度上转变很快。这些结果表明，单价阳离子和二价阳离子可能以不同方式稳定ACP。单价离子可能以类似的方式非特异性地平衡该蛋白质的固有负电荷，无论是天然形式、变性形式还是中间形式。二价阳离子通过结合仅存在于天然状态的特定位点来提供稳定性。
