西兰花（甘蓝变种意大利芥蓝）过氧化物酶的纯化及部分特性分析
Purification and partial characterization of broccoli (Brassica oleracea Var. Italica) peroxidases.
作者信息
Thongsook Tipawan, Barrett Diane M
机构信息
Department of Food Science and Technology, University of California, One Shields Avenue, Davis, California 95616-8598, USA.
出版信息
J Agric Food Chem. 2005 Apr 20;53(8):3206-14. doi: 10.1021/jf048162s.
Three peroxidase (POD) isoenzymes were purified from a soluble extract of broccoli stems. The acidic and neutral PODs were purified to homogeneity by using ion exchange and hydrophobic chromatography. The basic POD was purified by cation exchange and gel filtration chromatography. The neutral and basic PODs had molecular masses of approximately 43 kDa, and the acidic POD had a molecular mass of 48 kDa by SDS-PAGE. pI was approximately 4, 5, and 8 for acidic, neutral, and basic PODs, respectively. Optimum activity using guaiacol as the H donor was obtained at pH approximately 6 for both neutral and basic PODs and at pH approximately 4 for acidic POD. All three of the purified isoenzymes are glycosylated. Reaction rates with various substrates including guaiacol, guaiacol/MBTH, DMAB/MBTH, and ferulic acid/MBTH were different among the isoenzymes. K(m) and amino acid composition were also determined.
从西兰花茎的可溶性提取物中纯化出三种过氧化物酶（POD）同工酶。酸性和中性POD通过离子交换和疏水色谱法纯化至同质。碱性POD通过阳离子交换和凝胶过滤色谱法纯化。通过SDS-PAGE分析，中性和碱性POD的分子量约为43 kDa，酸性POD的分子量为48 kDa。酸性、中性和碱性POD的pI分别约为4、5和8。以愈创木酚作为氢供体时，中性和碱性POD在pH约为6时获得最佳活性，酸性POD在pH约为4时获得最佳活性。所有三种纯化的同工酶都进行了糖基化。三种同工酶对包括愈创木酚、愈创木酚/亚甲基蓝噻唑啉酮（MBTH）、二甲基苯胺/MBTH和阿魏酸/MBTH在内的各种底物的反应速率不同。还测定了米氏常数（K(m)）和氨基酸组成。
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