摘要
用硫酸铵沉淀、离子交换层析和亲和层析等方法,从鸡的胰和十二指肠中提纯了胰淀粉酶(P-Amyl)和十二指肠淀粉酶(D-Amyl).提纯倍数分别为67.5倍和78.8倍.二者提取液等电聚焦电泳鉴定均出现1条酶带,等电点分别为4.5和4.6.聚丙烯酰胺梯度凝胶电泳测定,二者的分子量均为53500.以可溶性淀粉为底物,测得P-Amyl和D-Amyl的Km值分别为6.20×10^-5和1.16×10^-5mol/L.P-Amyl对热敏感,D-Amyl则较稳定.EDTA强烈地抑制P-Amyl的活性,而对D-Amyl的抑制作用弱.P-Amyl和D-Amyl的最适pH分别为7.0和6.6.对P-Amyl和D-Amyl,Ca^(2+)、Mg^(2+)和Co^(2+)均有不同程度的激活作用,Cu^(2+)和Mn^(2+)有抑制作用;Zn^(2+)对P-Amyl有抑制作用;低浓度Fe^(3+)对P-Amyl有激活作用,高浓度Fe^(2+)对P-Amyl和D-Amyl均有抑制作用.
Amylases were purified from chicken pancreas and duodenum with ammonium sulfate precipitation, ion - exchange chromatography on DEAE - cellulose and affinity chro-matography on ConA-sepharose 4B. As a result of determination by using gradient electrophore-sis on slab gel of polyacrylamide (4 - 40%), both the pancreatic amylase (P-Amyl) and doudenal amylase (D-Amyl) have a moleular weight of 53500. Michelis constants of P-Amyl and D-Amyl are 6. 20×10-5 mol/L and 1. 16×10-5 mol/L, respectively. P-Amyl is ther-molabile and lost its all activities as being heated at 56℃ for five minutes. On the contrary, D-
Amyl has a stronger heatresistance and lost only 63.6% of activity. P-Amyl is inhibited strongly by EDTA, whereas D -Amyi is affected weakly. Optimal pH for P-Amyl and D -Amyiare 7. 0 and 6. 6 respectively. Co2+ , Ca2+, Mg2+ activate both of P-Amyl and D-Amyl to a different extent. Cu2+ and Mn2+ are effective in inhibiting both of them. Fe3+ at lower concentration activites P-Amyl but can inhibit the activaties of P-Amyl and D-Amyl at higher concentration.
关键词
鸡
胰
十二指肠
淀粉酶
提纯
chicken
pancreas j doudenum
amylase
isoenzyme
purification
property
