Triton X-114-aided purification of latent tyrosinase

被引:37
作者
NunezDelicado, E [1 ]
Bru, R [1 ]
SanchezFerrer, A [1 ]
GarciaCarmona, F [1 ]
机构
[1] UNIV MURCIA,FAC BIOL,DEPT BIOQUIM & BIOL MOLEC A,E-30071 MURCIA,SPAIN
来源
JOURNAL OF CHROMATOGRAPHY B-BIOMEDICAL APPLICATIONS | 1996年 / 680卷 / 1-2期
关键词
aqueous two-phase systems; tyrosinase; enzymes; triton X-114;
D O I
10.1016/0378-4347(96)00012-6
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Mushroom tyrosinase was partially purified using an aqueous two-phase system with Triton X-114. The purification achieved was 5.5-fold from a crude extract of mushroom pileus, with a high recovery of 84%. The phenols were reduced to 8% of the original content, avoiding pre- and post-purification tanning of the enzyme. The enzyme obtained was latent and was activated 3-fold by trypsin, 2.7-fold by changes in the pH and to different extents by cationic and anionic detergents, the latter being the more effective. There was also a synergistic effect between trypsin and detergent, at low detergent concentrations. When kinetically characterized, latent enzyme showed both monophenolase and diphenolase activities, the latter activity displaying an unexpected lag period before reaching the steady-state rate. This behaviour is characteristic of a hysteretic enzyme, acid has not been previously described for this enzyme. In addition, inhibition studies with substrate analogues were carried out, tropolone being found to be the most effective inhibitor.
引用
收藏
页码:105 / 112
页数:8
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