Purification, Characterization And Modification Of Groups Of The Catalase From Garlic Bolt

Catalase, also known as CAT, is a terminal oxidase wide-spreading in animals, plants and the vast majority of aerobe body. Physiologically, it catalyzes the decomposition of hydrogen peroxide in the cell to prevent peroxide, and removes active oxygen in vivo. It makes up a defense system of reactive oxygen in vivo together with SOD, POD. This Catalase is, as a marker enzyme of peroxisomes, of great significance to the improvement of resistant capacity of plants. It has a very important value in food processings clinical medical, textile industry, printing, dyeing, environmental protection and other industries. Besides, CAT is rare in the market and contains a relatively small number of other proteins, so it is of great value to research CAT in that it is obtained simply and conveniently, takes a low cost, and is high active and stable.Garlic bolt, also known as young garlic shoot or green garlic and grown from garlic seedling after a certain period, is a kind of green vegetables with a wide variety of sources and common in every season. Garlic contains a wealth of vitamin C, protein, free amino acid, polysaccharide, garlicrn and other nutrients. Besides, it can increase appetite, antiseptic effect. The study found that the CAT content is higher than other plants, stable, not reported from Garlic bolt, so this experiment, taking fresh garlic as research material, separates and purifies CAT and examines its partial enzymatic characteristics and modification of groups with an aim to providing a reference for further development application of the enzyme.The results of our study were follows:1. Purification of CAT from Garlic boltElectrophoresis-purity Catalase from fresh garlic bolt was obtained through procedures of homo-genization, Buffer solution extraction, ammonium sulfate precipitation, DEAE-Sepharose ion exchange chromatography and Superdex-200gel filtration. The specific activity of CAT reaches25975.36U/mg, as well as40.05%in terms of the recovery rate, and125.97in terms of the purification fold.2. Enzymatic characteristics of CAT from Garlic boltThe results of the study show:the molecular and subunit molecular weight of the CAT was about234.93Kd and59.16Kd respectively, presumably the enzyme is made up of four identical subunit; The enzyme was comparatively stable in the range of25-45℃and pH5-10, although its optimum temperature and pH were45℃and7.2; Furthermore, its.Km value, under optimum conditions, was38.2mmol/L.3. The effect of some chemical reagent to CAT from Garlic boltThe activity of CAT was inhabited by methanol, ethanol, isopropanol and SDS, KSCN, Ag+, Cu2-, Mn2-, Co2+, Cd2+, Zn2+,Ca2+, Mg2+, Ba2+, yet enhanced to some extent by Li+, Pb2-and low concentration of K". Besides, urea, EDTA and ASA have little effect on the enzyme.4. Functional group of CAT from Garlic boltThe results of the study show:Tryptophan residue may be essential function group of activity center of CAT from Garlic bolt. Yet, Disulfide bond, serine residue, lysine residue, sulfydryl, arginine residue, tyrosine residue, histidine residue, methionine sulfide base have no effect on CAT activity and are not essential function group of activity center of the enzyme. Besides, iodoacetic acid can activate the enzyme activity.