Roth L G, Berns D S, Chen C H
Wadsworth Center, New York State Department of Health and Department of Biomedical Sciences, The University at Albany, State University of New York, Albany, NY 12201-0509, USA.
Biophys Chem. 1996 Jun 11;60(3):89-97. doi: 10.1016/0301-4622(96)00013-0.
Differential scanning calorimetry, circular dichroism, and visible absorption spectrophotometry were employed to elucidate the structural stability of thermophilic phycocyanin derived from Cyanidium caldarium, a eucaryotic organism which contains a nucleus, grown in acidic conditions (pH 3.4) at 54 degrees C. The obtained results were compared with those previously reported for thermophilic phycocyanin derived from Synechococcus lividus, a procaryote containing no organized nucleus, grown in alkaline conditions (pH 8.5) at 52 degrees C. The temperature of thermal unfolding (t(d)) was found to be comparable between C. caldarium (73 degrees C) and S. lividus (74 degrees C) phycocyanins. The apparent free energy of unfolding (DeltaG([urea]=0)) at zero denaturant (urea) concentration was also comparable: 9.1 and 8.7 kcal/mole for unfolding the chromophore part of the protein, and 5.0 and 4.3 kcal/mole for unfolding the apoprotein part of the protein, respectively. These values of t(d) and DeltaG([urea]=0) were significantly higher than those previously reported for mesophilic Phormidium luridum phycocyanin (grown at 25 degrees C). These findings revealed that relatively higher values of t(d) and DeltaG([urea]=0) were characteristics of thermophilic proteins. In contrast, the enthalpies of completed unfolding (DeltaH(d)) and the half-completed unfolding (DeltaH(d)) 1 2 for C. caldarium phycocyanin were much lower than those for S. lividus protein (89 versus 180 kcal/mole and 62 versus 115 kcal/mole, respectively). Factors contributing to a lower DeltaH(d) in C. caldarium protein and the role of charged groups in enhancing the stability of thermophilic proteins were discussed.
采用差示扫描量热法、圆二色光谱法和可见吸收分光光度法，来阐明源自嗜热蓝藻的嗜热藻蓝蛋白的结构稳定性。嗜热蓝藻是一种真核生物，含有细胞核，在54摄氏度、酸性条件（pH 3.4）下生长。将所得结果与之前报道的源自蓝纤维藻的嗜热藻蓝蛋白的结果进行比较，蓝纤维藻是一种不含细胞核的原核生物，在52摄氏度、碱性条件（pH 8.5）下生长。发现嗜热蓝藻（73摄氏度）和蓝纤维藻（74摄氏度）藻蓝蛋白的热解链温度（t(d)）相当。在零变性剂（尿素）浓度下的表观解链自由能（DeltaG([尿素]=0)）也相当：蛋白质发色团部分解链时分别为9.1和8.7千卡/摩尔，蛋白质脱辅基蛋白部分解链时分别为5.0和4.3千卡/摩尔。这些t(d)和DeltaG([尿素]=0)值显著高于之前报道的嗜温 luridum鞘丝藻藻蓝蛋白（在25摄氏度下生长）的值。这些发现表明，相对较高的t(d)和DeltaG([尿素]=0)值是嗜热蛋白质的特征。相比之下，嗜热蓝藻藻蓝蛋白完全解链的焓（DeltaH(d)）和半解链的焓（DeltaH(d)）1 2远低于蓝纤维藻蛋白（分别为89对180千卡/摩尔和62对115千卡/摩尔）。讨论了导致嗜热蓝藻蛋白DeltaH(d)较低的因素以及带电基团在增强嗜热蛋白质稳定性中的作用。
