Faculty of Science, Department of Biochemistry, Khon Kaen University, Khon Kaen, 40002, Thailand.
Protein J. 2012 Jan;31(1):43-50. doi: 10.1007/s10930-011-9372-7.
Crocodylus siamensis hemoglobin (cHb) was purified by gel filtration chromatography and visualized by SDS-PAGE. Effects of temperature and pH on secondary structure and conformation changes of cHb were studied using circular dichroism spectropolarimeter and fourier transform infrared spectrophotometer. The secondary structure of intact cHb was mainly α-helices. cHb was not heat stable when heated at 65 °C and cooled down to original temperature, indicating the irreversible unfolding process. The stability of cHb at different pH ranging from 2.5 to 10.5 was determined. The maximum value of the α-helix content was found at pH 3.5 and tended to decrease at strong acid and strong base. The antioxidant activities of heat treated cHb and cHb in solution with pH range 2.5 to 10.5 were tested by DPPH radical scavenging assay. cHb at pH 4.5, having highest β-turn structure, showed highest radical scavenging activity. In contrast to pH, heat had no effect on antioxidant activity of cHb.
暹罗鳄血红蛋白（cHb）通过凝胶过滤层析进行纯化，并通过 SDS-PAGE 进行可视化。使用圆二色光谱仪和傅里叶变换红外分光光度计研究了温度和 pH 对 cHb 二级结构和构象变化的影响。完整的 cHb 的二级结构主要为α-螺旋。当在 65°C 下加热并冷却至原始温度时，cHb 不稳定，表明发生了不可逆的展开过程。测定了 cHb 在 pH 2.5 至 10.5 之间的不同 pH 值下的稳定性。在 pH 3.5 时发现α-螺旋含量的最大值，并在强酸和强碱下趋于降低。通过 DPPH 自由基清除试验测试了热处理的 cHb 和在 pH 2.5 至 10.5 范围内的溶液中的 cHb 的抗氧化活性。具有最高β-转角结构的 pH 4.5 的 cHb 显示出最高的自由基清除活性。与 pH 不同，热对 cHb 的抗氧化活性没有影响。
