茶花粉超氧化物歧化酶的性质研究
何晓红,王雪松,陈智博,于笑坤,张博珣,刘兰英 * (吉林大学生命科学学院,吉林 长春 130023)

摘 要:从茶花粉中提纯的超氧化物歧化酶(SOD)为Cu,Zn-SOD,分子量为69.5 kD,亚基分子量为34.7kD,是 由相同亚基组成的二聚体。等电聚焦电泳测得等电点为 pH4.1,二甲氨基丹磺酰氯法确定其 N- 末端氨基酸为 Gly, 圆二色谱法测得α螺旋含量为21.8 %。性质研究表明该蛋白在pH5~10范围内基本稳定,在5~35°C内保温30min 酶活性基本保持不变;化学试剂KCN、H2O2、EDTA、巯基乙醇、SDS 对酶活性完全抑止,4mol/L 的尿素不 影响酶活性。茶花粉SOD 用EDTA 去除金属离子后重新加入Cu2+、Zn2+ 恢复至天然活性,而加入Mn2+、Mg2+ 只 能部分恢复活性。

关键词:茶花粉;超氧化物歧化酶;性质

Characterization of Superoxide Dismutase from Camellia Pollen

HE Xiao-hong,WANG Xue-song,CHEN Zhi-bo,YU Xiao-kun,ZHANG Bo-xun,LIU Lan-ying* (College of Life Science, Jilin University, Changchun 130023, China)

Abstract:Super oxi dedismutase from camellia pollen was purified to homogeneity and assayed to be Cu,Zn-SOD according to its specific sensitivities to hydrogen peroxide, cyanogens potassium and chloroform-alcohol. The molecular weights of the SOD and its subunit were 69.5kD and 34.7kD, respectively. Isoelectric point analysis indicated that it was an acidic protein whose isoelectric point was pH4.1. The N-terminal amino acid of the enzyme was identified to be Gly by DNS- Cl method. Its content ofα-Helix was also assayed to be approximately 21.8% according to the result of circular dichroism (CD) spectra. The effects of pH, temperature, chemical reagents and metal ions on SOD were studied respectively in its molecular stability experiments. Results showed that the optimum temperature and pH of the enzyme°Cweraend265.5 respectively. It lost its activity entirely

when the pH was higher than 10, and its activity was also highly inhibited when2OK2,CNS,DSH and β- mercaptoethanol existed, but 4mol/L carbamide had no effect on it. The activity of the SOD whose metal ions were driven out by EDTA recovered 2+ 2+ 2+ 2+ totally after combining with Cu, Zn but only partly recovered after combing with,MnMg .

Key words:camellia pollen;superoxide dismutase;characterization 

中图分类号:Q554 文献标识码:A 文章编号:1002-6630(2006)04-0085-04

茶花粉具有清热、解毒、抗衰老、脱脂减肥、 治疗糖尿病、高血压等功效。茶花粉中氨基酸含量为 花粉之首,微量元素锰、钴、铭等以及烟酸含量均 比普通花粉高,故为保健、医疗及癌症的首选花粉 [ 1 ] 。本文针对茶花粉的抗衰老特性从新鲜茶花粉中分 离提纯了 S O D ,对其所属类型进行了鉴定以及分子稳 定性研究,同时对其结构进行了表征。这些研究结果对于揭示花粉抗衰老作用的机理在分子水平上提供了理 论依据,同时也为 S O D 的结构与功能研究增添了新的 成员。