白腐菌Phanerochaete sordida YK-624产锰过氧化物酶的生产及初步纯化

PRODUCTION AND PURIFICATION OF MANGANESE PEROXIDASE BY WHITE ROT FUNGI PHANEROCHAETE SORDIDA YK-624

利用白腐菌PhanerochaetesordidaYK 624(ATCC90872),采用Kirk液体培养基在富氧条件下进行振动培养,可得到锰过氧化物酶(MnP)活性较高和漆酶活性微量的酶液。该酶液通过超滤、透析、二乙氨基乙基琼脂糖凝胶(DEAE sepharoseCL 6B,pharmacia)离子交换柱层析的方法进行初步纯化后,得到比活性很高(1941.54IU/mg)的MnP酶液。漆酶活性和MnP活性比值的变化、SDS PAGE凝胶电泳试验和406nm附近的吸光值变化均表明,经离子交换柱层析后的YK1部分是主要的MnP,电泳试验同时表明纯化后的主要MnP的分子质量约为43kDa。经超滤、透析、离子交换柱层析纯化后得到的酶液YK1可用于进一步地研究及应用。

Crude enzymes liquor,which contained high manganese peroxidase(MnP) activity and trace laccase (activity),was produced by cultivating white rot fungi Phanerochaete sordida YK-624(ATCC 90872) under shaking and high O2 content conditions in Kirk liquor culture medium.Ultra-filtration,dialysis and DEAE-sepharose CL-6B((pharmacia)) anion-exchange chromatography were used to purify the enzymes solutions.Thus,enzymes solution of high specific MnP activity marked as YK1(1 941.54 IU/mg) was obtained.The ratio of MnP to laccase,the results of SDS-polyacrylamide gel electrophoresis and the absorption spectra around 406 nm proved that YK1,was the main part of MnP activity.Similarly,SDS-polyacrylamide gel electrophoresis showed molecular weight of YK1 to be about (43 kDa).Thus,YK1,obtained from ultrafiltration,dialysis and anion-exchange chromatography,could be chosen as the enzyme solution for more application research,such as biobleaching and improvement of fiber quality.