平菇漆酶对不同重金属胁迫的响应

Change in Laccase Activities of Pleurotus ostreatus in Response to Heavy Metals in Liquid Culture

平菇是一种重要的白腐真菌,可以合成包括漆酶在内的多种木质素纤维素降解酶.以平菇(ACCC 52857)为材料,通过在培养基中分别添加不同浓度的重金属离子(Cu2+、Pb2+、Cd2+),考察菌体生长及培养基中漆酶活性的变化情况,并利用非变性聚丙烯酰胺电泳检测不同重金属离子对平菇漆酶同工酶(Lacc2和Lacc10)活性的影响.结果表明:在培养基中添加Cu2+、Pb2+、Cd2+对平菇漆酶活性均有不同程度的增强作用.培养至第9天时,对照组漆酶活性达到最大值,为(565.6±5.2)nkat/m L;而添加重金属离子的各组漆酶活性仍在上升,Cu2+、Pb2+、Cd2+分别以终浓度为3 000、2 000、150μmol/L时对漆酶活性的增强作用最大,培养至第20天、第15天、第17天时,漆酶活性达到最大值,分别为(8 612.1±840.0)、(2 204.0±347.0)、(2 928.0±217.7)nkat/m L.非变性聚丙烯酰胺凝胶电泳结果显示,3种重金属离子对液体培养条件下Lacc2和Lacc10这2种漆酶同工酶活性的影响不同.培养基中较高c(Cu2+)可以显著增加Lacc10活性;随着c(Pb2+)的升高,Lacc2活性升高;随着c(Cd2+)不断增加,Lacc2活性略有降低,但Lacc10活性逐渐增加.研究结果有助于进一步解释漆酶的生理功能和漆酶基因的表达调控机制.

Pleurotus ostreatus is a white-rot fungus secreting different enzymes involved in lignin and cellulose degradation. Laccase and isoenzymes activities of P. ostreatus（ ACCC 52857） were tested in the presence of different concentrations of Cu2 ＋,Pb2 ＋or Cd2 ＋under shaking conditions. The mycelia growth and laccase activities were assayed,and laccase isoenzymes were analyzed using non-denaturing polyacrylamide gel electrophoresis. The results showed that the laccase activity increased with treatments of Cu2 ＋,Pb2 ＋or Cd2 ＋in the medium compared with the control group. The highest laccase activity（ 565. 6 ± 5. 2） nkat/m L was obtained on the 9thday of cultivation in the control group,while laccase activity continued to increase in the presences of Cu2 ＋,Pb2 ＋or Cd2 ＋. The highest laccase activities-（ 8612. 1 ± 840. 0）,（ 2204. 0 ± 347. 0） and（ 2928. 0 ± 217. 7） nkat/m L-were obtained on the 20 th,15thand 17 thday of cultivation by adding 3000 μmol/L Cu2 ＋,2000 μmol/L Pb2 ＋or 150 μmol/L Cd2 ＋to the medium,respectively. The results of non-denaturing polyacrylamide gel electrophoresis indicated that the two most abundant isoenzymes（ Lacc2 and Lacc10） in liquid culture showed different expression patterns in response to the concentration variations of Cu2 ＋,Pb2 ＋and Cd2 ＋. Lacc10 activity was increased greatly with high concentration of Cu2 ＋in the medium,while Lacc2 activity was increased by Pb2 ＋. In the presence of Cd2 ＋,Lacc2 activity was slightly suppressed,while Lacc10 showed a reverse trend. The results may shed light on the biological functions and the regulation mechanisms of laccase in P. ostreatus.