Pressure and Temperature Effects on Protein Solutions

用等温滴定量热法测定了溶菌酶在乙醇水溶液中的过量偏摩尔焓。这表明，酒精占用蛋白质表面的结合位点可能不是影响蛋白质性质的必要条件。溶菌酶和甘氨酸在水中溶解度的压力依赖性测定。前者溶解度随压力升高而增大，后者溶解度随温度升高而减小。这表明，从组成氨基酸的角度出发，压力是考虑蛋白质水合作用的关键参数之一。在-10-70℃范围内测定肌红蛋白（Mb）的高压FT-IR。第二种结构的压力依赖关系遵循双态模型。该结构在低温下可逆，但随着温度的升高，不可逆地转变为分子间无规卷曲的β片结构。通过构建细胞色素c血红素裂解酶与细胞色素c共表达的质粒，在大肠杆菌中成功地表达了酵母细胞色素c，该体系获得了较高的野生型细胞色素c产量。当血红素铁的关键的Met80配体被Ala取代并且不支持呼吸作用时，在大肠杆菌中可以表达，这表明Lys72不像在酵母中表达时那样是合成后三甲基化的。这一发现并不令人惊讶，因为大肠杆菌可能不具有催化这种修饰反应的转甲基酶。

Excess partial molar enthalpy of lysozyme in aqueous alcohol was measured using isothermal titration calorimetry. It was suggested that occupancy by the alcohol of a binding site on the surface of the protein may not be necessary for the alcohol to affect the properties of the protein. Pressure dependences of the solubiity for lysozyme and glycine in water measured. The formaer increased and the latter decreased with increasing pressure though both solubilities increased with increasing temperature. It suggests that the pressrue is one of key parameter to consider the protein hydration in the stand point of the constituent amino acids.High-pressure FT-IR of Myoglobin (Mb) was measured in the range -10-70ﾟC.Pressure dependence of the second structure of Mb follows two state model. The structure irreversibly changes to random coil forming intermolecular beta-sheet with increasing temperature though it becomes reversible at low temperature.It was, succeeded in expressing yeast cytochrome c in E.coli through construction of a plasmid for the co-expression of cytochrome c heme lyase and cytochrome c. The yield of wild-type cytochrome c from this system is relatively high. Variants in which the crucial Met80 ligand to the heme iron is replaced with Ala and that do no support respiration can be expressed in E.coli demonstrated that Lys72 is not trimethyllated postsynthetically as it is when expressed in yeast. This finding is no surprising because presumably E.coli does not posses the transmethylase enzyme that catalyzes this modification reaction.