摘要
目的考察不同环境因素对过氧化氢酶(CAT)活性和构象的影响,为蛋白质制剂过程提供依据。方法改变温度、pH、变性剂浓度,用紫外分光光度法考察CAT溶液活性变化,并用荧光光谱法研究其构象变化。结果CAT溶液25℃p、H 7时,活性及荧光强度都达到最大;温度升高,荧光强度逐渐下降;pH降低或升高,荧光最大发射波长均红移;低浓度变性剂对CAT活性中心有稳定作用;浓度增加,荧光谱带红移,活性降低甚至丧失;脲和盐酸胍对CAT荧光有动态猝灭作用。结论用荧光光谱法研究CAT溶液稳定性,方法简便、准确、可靠。
Purpose To investigate the effect of different environmental factors on catalase （CAT） activity and conformation and to provide a basis for preparation of protein. Methods Altering the factors like temperature, pH and concentration of denaturant （Urea and GuHCI）, figuring out the activity of CAT using fluorescence spectroscopy to investigate the conformation of CAT. Results At 25℃ and pH = 7, CAT solution was the most stable. As the temperature rising, the fluorescence intensity dropped, and as pH reducing or rising, the fluorescence spectrum of CAT solution showed red shift. The increasing concentration of denaturants changed the microenvironment of tyrosine and tryptophan inside the CAT molecule, and gradually exposed to the hydrophilic environment, causing the fluorescence spectrum band＇s red shift and the descent activity of CAT. It also shows that denaturants have a strong ability to quench the catalase fluorescence mainly through a dynamic quenching procedure. Conclusion The method of fluoreseenee spectroseopy to investigate the stability of CAT is simple, accurate and reliable.
出处
《中国生化药物杂志》
CAS
CSCD
北大核心
2009年第4期243-247,共5页
Chinese Journal of Biochemical Pharmaceutics
关键词
过氧化氢酶
稳定性
活性
构象
荧光光谱法
catalase
stability
activity
conformation
fluorescence spectroscopy
