一种具有热滞活性的女贞叶质外体过氧化物酶

An Apoplastic Peroxidase with Thermal Hysteretic Activity Purified from Leaves of Ligustrum lucidum Ait.

采用浸出离心法从女贞叶中分离的质外体蛋白,约占叶片可溶蛋白的1.63%,其中的质外体过氧化物酶占叶过氧化物酶(POD)总酶活性的3.14%。以凝胶过滤和离子交换层析分离纯化得到一分子量为62 kDa的质外体过氧化物酶,其对H_2O_2的表观K_m值为1.3 mmol·L^(-1),表观V_(max)值为189.33 U·mg^(-1)(蛋白);对愈创木酚的表观K_m值为1.78 mmol·L^(-1),表观V_(max)值为215.26 U·mg^(-1)(蛋白)。在蛋白质浓度为10 mg·mL^(-1)时,其热滞活性为0.332℃。此种蛋白的酸性氨基酸含量和亲水性氨基酸含量较高,类似于沙冬青和黑麦的抗冻蛋白。

An apoplastic protein was isolated by soaking and centrifugation, its content was about 1.6% of the total leaf protein. The activity of the apoplastic peroxidase was about 3.14% of the total leaf peroxidase. An apoplastic peroxidase was purified by gel filtration and ion exchange chromatography. The molecular weight of the purified peroxidase was 62 kDa. Its apparent Km value of hydrogen peroxide and guaiacol was 1.3 mmol·L^-1 and 1.78 mmol·L^-1 respectively, the apparent Vmax was 189.33 U·mg^-1 （protein） and 215.26 U·mg^-1 （protein） respectively. Its thermal hysteretic activity was 0.332℃ at the protein concentration of 10 mg·mL^-1. The compositions of acidic amino acids and hydrophilic amino acids of the enzyme were similar to that of antifreeze proteins in Ammopiptanthus mongolicus and Secale cereale.