摘要
将 Sephadex G-200与β-硫酸酯乙砜基苯胺(SESA)首先醚化制备对氨基苯砜乙基交联葡聚糖(ABSE-Sephadex G-200),然后经重氮化固定无花果蛋白酶。固定化酶的活力回收达69%。苯甲酰-DL-精氨酰-β-蓁胺(BANA)对该酶固定化过程中的活性变化有保护作用。天然酶与固定化酶都具有良好的耐热性,在59～60℃,80min,活性均无明显下降;在69～70℃,80min 固定化酶较天然酶更稳定。用 BANA 为底物,在半胱氨酸存在下,测定了两种形式酶的动力学性质。在 pH7.7的磷酸盐缓冲系统中,37℃天然无花果蛋白酶的 Km=0.32m-mol/L;在间歇振摇下固定化酶的表观 Km′=1.02mmol/L。最适 pH 无明显改变,均为7.7。
Sephadex G-200 and (P.β-Sulphatoethylsulphonyl)anine (SESA) were first etherified to prepare p-aminobenzene sulphone ethyl(ABSE)-Sephadex G-200 and then,ficin was fixed on ABSE-Sephadex G-200 by means of diazo-reaction.The activity recovery of the immobilized en- zyme approximatly was 69%.Benzoyl-DL-arginine-β-maphthylamide hydrochloride (BANA) was used to protect the enzyme activity during the immobilization process.The native and immobilized enzymes had higher thermal stability.The activity did not decreased in 59～60℃ for 80min.The thermal stability of the immobilized enzyme was higher than that of the native ficin in 69～70℃ for 80 min.When BANA was used as substrate,the kinetiks properties of the two types of en- zymes were investigated in the presence of cystein.In the system of phosphate-buffer pH 7.7, 37℃,Michaelis constant (Km)of the native ficin was 0.32 mmol/L,apparent Km' of the immo- bilized enzyme was 1.02 mmol/L while being shaken at intervals.The optimal pH of the two en- zymes did not changed,which was pH 7.7.
出处
《潍坊医学院学报》
1993年第1期1-5,共5页
Acta Academiae Medicinae Weifang
关键词
蛋白酶
固相酶类
动力学
肽
ficin
immobilization
kinetic properties
