糖苷水解酶

nullβ－GALACTOSIDASEβ－GALACTOSIDASE β-Galactosidase(β－D-galactoside galactohydrolase, EC 3.2.1.23), referred to frequently as lactase (trivial name), has been one of the most carefully studied glycosidases because of its ready inducibility in microorganisms. The nature of the lac operon has been well elucidated and researchers have been successful in separating the area containing this operon from the chromosome. nullThe enzyme catalyzes the hydrolysis of β-D-galactosides and α - L - arabinosides. All lactases are β-galactosidases but some β-galactosidases, including those from plant cells and mammalian organs other than the intestine, have little or no activity on lactose because their function is to hydrolyze other galactosyl moieties, including glycolipids, glycoproteins, and mucopolysaccharides nullβ-Galactosidase hydrolyzes O-glycosyl bonds and exhibits strict specificity for the glycone part of the substrate nullThe enzyme shows wide tolerance for the structure of the aglycone which may be another sugar, an alkyl group or an aryl group. The nature of the aglycone, however, strongly influences the kinetic parameters IMPORTANCE TO FOOD QUALITY IMPORTANCE TO FOOD QUALITY Hydrolysis of lactose leads to glucose and galactose, which are much sweeter than lactose. Lactose has a limited solubility and tends to precipitate out in frozen dairy products, which gives a grainy texture to the product. Partial hydrolysis of the lactose prevents this. nullActivity of intestinal lactase in mammals decreases after weaning, and this leads to some degree of lactose intolerance in many population groups. Consequently, the enzyme is used in clinical settings to aid lactose digestion and in commercial operations to produce low-lactose diary products. It is important to appreciate that most adults are often allergic to milk since they do not have β-galactosidase and therefore cannot metabolize lactose.null Whey is the principal byproduct of cheese manufacture and contains > 70% lactose on a dry-weight basis. Hydrolysis of the lactose improves whey utilization in a variety of products and processes nullnull SOURCES of lactase PROPERTIES AS AN ENZYME PROPERTIES AS AN ENZYME The enzymatic properties ofβ-galactosidase vary with the source. Comparison of kcat/Km shows that the synthetic substrate ONPG (o-nitrophenyl β-D-galactopyranoside) is a better substrate than lactose in all cases. The fungal enzyme from Aspergillus niger is the most thermostable and is unaffected by cations or sulfhydryl modifiers such as p-chloromercuribenzoate. It is the most susceptible to product inhibition by galactose.null乳糖酶作用的机制 乳糖酶作用的机制 巯基作为广义的酸使得糖苷键的氧原于质子化，而咪唑基作为亲核试剂进攻糖基的C（1）形成一个含碳—氮键的共价中间物．在半乳糖基被切割下来之后，巯基阴离子作为广义碱从水分子抽取一个质子，从而形成OH-进攻C（1）。在反应的各个步骤． 异头碳的构型都没有变化，因而，由乳糖酶催化水解的产物仍然保持原来的β—构型． null乳糖酶活力乳糖酶活力 测定的方法有下列几种： （1）根据反应过程中溶液旋光值的变化计算反应速度．虽然乳糖水解成半乳糖和果糖引起的旋光率的改变是较小的(从＋52.5至+67.0)，但是采用旋光仪还是能够定量地测定这个变化 （2）最常用的方法采用葡萄糖氧化酶－过氧化氢酶－色素原测定反应中生成的葡萄糖的量．也可以计算反应的速度；null（3）根据反应过程中还原糖量的变化计算反应速度这个方法由于乳糖也是还原糖，因而反应体系具有高的空白值，给测定带来很大的困难 （4）采用一种生色的化合物作为底物，经乳糖酶催化水解后，就转变成一种有色产物，于是很容易采用分光光度法测定反应的速度． 目前，邻/对—硝基苯—β—D—半乳糖苷（PNPG/ONPG)）是乳糖酶最佳的生色底物．这不仅是因为用分光光度法能非常方便地测定酶催化反应的速度，而且还由于在底物浓度低于使酶饱和的水平时的反应速度相当于采用乳糖作为底物时的300倍．β-FRUCTOFURANOSIDASE β-FRUCTOFURANOSIDASE Because of inversion of rotation (+66.5 to –20.5~) of the solution during reaction, the enzyme became known as invertase. The rate of glucose production can be followed by the glucose oxidase-peroxidase-chromogen method. nullChemical Structure of Substrates 水苏四糖棉子糖nullβ-Fructofuranosidase is fund widely distributed in nature and is present in plants, animals, and microorganisms. nullThere are two types of enzymes, β-fructofuranosidase and α-glucosidase (α-D-glucoside glucohydrolase, EC 3.2.1.20), which can hydrolyze sucrose. The glycosidic bond in sucrose involves the reducing groups of both D-glucose and D-fructose. One type of enzyme hydrolyzes the glucosidic bond bet,a'een C(2） and O (β-fructofuranosidase) and the other hydrolyzes the bond between C(1) and O (α-glucosidase）nullThe following properties have been described for yeast β-fructofuranosidase. Km values for sucrose and raffinose are 0.016 M and 0.24 M. respectively. In order to see appreciable activity with raffinose, one must have a very high concentration of substrate. The pH optimum of the enzyme is in the range 4. 5 to 5.5. The enzyme is stable to around 35C. The enzyme is inhibited by iodine (to only 50%) and by heavy metal ions (Hg2＋. Ag1＋, Cu2＋). Iodoacetate（碘乙酸） and ferricyanide have very little effect on activity.