Valente-Mesquita V L, Botelho M M, Ferreira S T
Departamento de Bioquímica Médica, Instituto de Ciências Biomédicas, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ 21941-590, Brazil.
Biophys J. 1998 Jul;75(1):471-6. doi: 10.1016/S0006-3495(98)77535-6.
Effects of hydrostatic pressure on dimeric beta-lactoglobulin A (beta-Lg) were investigated. Application of pressures of up to 3.5 kbar induced a significant red shift ( approximately 11 nm) and a 60% increase in intrinsic fluorescence emission of beta-Lg. These changes were very similar to those induced by guanidine hydrochloride, which caused subunit dissociation and unfolding of beta-Lg. A large hysteresis in the recovery of fluorescence parameters was observed upon decompression of beta-Lg. Pressure-induced dissociation and unfolding were not fully reversible, because of the formation of a nonnative intersubunit disulfide bond that hampered correct refolding of the dimer. Comparison between pressure dissociation/unfolding at 3 degrees C and 23 degrees C revealed a marked destabilization of beta-Lg at low temperature. The stability of beta-Lg toward pressure was significantly enhanced by 1 M NaCl, but not by glycerol (up to 20% v/v). These observations suggest that salt stabilization was not related to a general cosolvent effect, but may reflect charge screening. Interestingly, pressure-induced dissociation/unfolding was completely independent of beta-Lg concentration, in apparent violation of the law of mass action. Possible causes for this anomalous behavior are discussed.
研究了静水压力对二聚体β-乳球蛋白A（β-Lg）的影响。施加高达3.5千巴的压力会导致β-Lg出现显著的红移（约11纳米），其固有荧光发射增加60%。这些变化与盐酸胍诱导的变化非常相似，盐酸胍会导致β-Lg的亚基解离和展开。在β-Lg减压时，观察到荧光参数恢复过程中存在很大的滞后现象。压力诱导的解离和展开并非完全可逆，这是因为形成了非天然的亚基间二硫键，阻碍了二聚体的正确重折叠。在3℃和23℃下进行的压力解离/展开比较表明，β-Lg在低温下明显不稳定。1 M NaCl显著增强了β-Lg对压力的稳定性，但甘油（高达20% v/v）则没有这种作用。这些观察结果表明，盐的稳定作用并非与一般的共溶剂效应相关，而是可能反映了电荷屏蔽。有趣的是，压力诱导的解离/展开完全独立于β-Lg的浓度，这显然违反了质量作用定律。文中讨论了这种异常行为的可能原因。
