- 作者: 李耀坤 ; 張麗芬 ; 許渲姝 等
- 作者服務機構: 交通大學應用化學系
- 中文摘要: A sweet almond .beta.-glucosidase (EC 3.2.1.21) isozyme was purified from commercial crude product. The process of purification consisted of a Protein-Pak Q anion exchange chromatography following by a Superdex 75 HR gel filtration separation. The purified enzyme is a monomeric glycoprotein with molecular weight of 58kDa and pI=4.55 which is distinguished from reported isozymes. The enzyme has a pH optimum in the range of 5.2-5.6 when p-nitrophenyl-.beta.-D-glycopyranosides are used as substrate and is stable up to 50.degree.C at that pH range. The purified protein also exhibits profound .beta.-galactosidase and.alpha.-L-arabinosidase activity. The study of substrate specificity revealed that lacking of hydroxymethyl group at C-5 of glycosides resulted in higher affinity for substrate binding to enzyme, whereas the chemical step of hydrolysis(k/sub cat/) was prevented significantly. The pH activity profile displayed a bell-shaped curve for all measured p- nitrophenyl-.beta.-D -glycopyranosides with apparent pK/sub 1/ and pK/sub 2/ values of 4.4-4.7 and 6.2-6.4, respectively. This isozyme was strongly inhibited by.delta.-gluconolactone (K/sub i/=160.mu.M) and 4- phenylimidazole (K/sub i/=17.8.mu.M) reversibly at pH 6.2. Among the tested glycoses, the binding affinity of N-acetyl-.beta.-D-glucosamine to the enzyme (K/sub i/=52mM) was 6 times stronger than that of glucose and its epimers.
- 英文摘要: --
- 中文關鍵字: .Beta.-Galactosidase; .Beta.-D-Glycopyranoside; Isozyme; Substrate Specificity
- 英文關鍵字: .beta.-半乳糖;.beta.-D-喃葡糖;同功;基質特異性