大熊猫乳酸脱氢酶同工酶M_4的纯化与某些性质的研究

利用8-(6-氨已基)-氨基-5’-AMP Sepharose亲和层析和DEAE-Sephadex A50离子交换层析纯化了大熊猫LDH-M_4。纯化的大熊猫LDH-M_4呈针状晶体,比活为412单位/毫克。聚丙烯酰胺凝胶电泳鉴定为一条区带。SDS凝胶电泳测得其亚基分子量为35,900;等电聚焦电泳测得其等电点为8.05。经氨基酸组成分析,得出每个大熊猫LDH-M亚基含有5个Cys,26个Lys和10个Arg。其N-末端氨基酸残基可能为封闭的,C末端氨基酸残基经测定为Phe。大熊猫LDH-M_4的TPCK-胰蛋白酶水解物在纤维素膜指纹图谱上呈现35个肽斑,与已知序列的猪LDH-M_4的指纹图谱相比较,多数肽斑位置相同,约有10个肽斑在两者指纹图谱上有差异。

PURIFICATION AND PARTIAL CHARACTERIZATION OF LDH-M_4 ISOZYME FROM GIANT PANDA

The LDH-M4 isozyme from giant panda muscle was purified to homogeneity by employing 8-(6-aminohexyl)-amino-5-AMP-Sepharose affinity chromatography followed by DEAE-Sephadex ion exchange chromatography.The enzyme was crystal lized and its specific activity was found to be 412 unit/mg.The subunit weight is 35,900 daltons as determined by SDS-PAGE.The pI is 8.05 as determined by IEF.Among 331 amino acid residues per subunit,there are 26 lysine,10 arginine and 5 half-cystine residues.The amino-terminus seems to be blocked.The carboxyl-terminus is phenylalanine.The tryptic peptide map of LDH-M4 from pig and giant panda are compared.Among their 35 and 34 peptide spots found,most of them have identical positions while ten of them seem to be different in the two maps.