摘要
将小麦麦胚和麸皮的浸取液，经加热和用ｐＨ沉淀，获得巯基蛋白酶抑制剂（简称ＣＰＩ）粗品；再经ＤＥＡＥ－Ｓｅｐｈａｒｏｓｅ，ＳｅｐｈａｄｅｘＧ－１００分子筛层析，在ＳＤＳ－ＰＡＧＥ和ＨＰＬＣ柱上得到均为单一蛋白带的小麦ＣＰＩ纯品．其纯化度为原料浸液的４２倍．由ＳＤＳ－ＰＡＧＥ测得ＣＰＩ分子为单一肽链组成，分子量为１３４００ｋｄ，ＳｅｐｈａｄｅｘＧ－１００柱测得ＣＰＩ分子量为１３０００ｋｄ，等电点为ｐＨ５．０，Ｎ末端氨基酸为Ａｌａ．纯化的ＣＰＩ在９０℃下处理４５ｍｉｎ或１００℃下处理１０ｍｉｎ后，仍保持有１００％的抑制活性；在ｐＨ３．０～１１．０的范围内，小麦ＣＰＩ分子非常稳定，抑制活性保持１００％，显示其分子有很高的酸碱耐受性．小麦ＣＰＩ与木瓜蛋白酶的结合是瞬时的．实验表明，该抑制剂对木瓜蛋白酶和无花果蛋白酶有很强的抑制作用，对菠萝蛋白酶有弱抑制作用，但对胰蛋白酶则无抑制作用，表明小麦ＣＰＩ是一种对巯基蛋白酶专一的抑制剂；对木瓜蛋白酶的抑制摩尔比为５．８∶１，属竞争性抑制类型，Ｋｉ值约为２．
Cysteine proteinase inhibitor(CPI) from wheat seed was purified by heating,adjusting pH,DEAE Sepharose chromatography and gel filtration on Sephadex G 100 column.The CPI was thus purified as high as about 42 fold.The purified CPI shows a single peak on HPLC and also a single band on SDS PAGE.The wheat CPI is a single polypetide with a molecular weight of 13400kd by SDS PAGE and 13000kd by gel filtration,with an isoelectric point at pH5.0,and its N terminal amino acid is Ala.The purified CPI retains its full activity between pH3.0～11.0 at 90℃ for 45min or at 100℃ for 10min.It exhibits strong inhibition too papain and ficin and slight inhibition to bromelian,but not to trypsin at all.There indicate that wheat CPI is a specific inhibitor to cysteine proteinase.The inhibition of papain by wheat CPI was competitive,with a K i value of 2.44×10 -8 mol/L and the mole ratio is 5.8∶1.The reaction of CPI to papain is instanteneous.
出处
《四川大学学报（自然科学版）》
CAS
CSCD
北大核心
1997年第2期235-240,共6页
Journal of Sichuan University(Natural Science Edition)
基金
国家自然科学基金
关键词
巯基蛋白酶
抑制剂
蛋白酶抑制剂
小麦
提纯
cysteine proteinase inhibitor, competitive inhibitor, protein, proteinase inhibitor
