Department Biochemistry of Microorganisms, Zabolotny Institute of Microbiology and Virology, National Academy of Sciences of Ukraine, 154 Zabolotny St, Kyiv, 03143, Ukraine.
Appl Biochem Biotechnol. 2018 Mar;184(3):953-969. doi: 10.1007/s12010-017-2593-2. Epub 2017 Sep 18.
Naringinase which was extracted from the fermented broth of Cryptococcus albidus was purified about 42-folds with yield 0.7% by sulfate fractionation and chromatography on Toyopearl HW-60, Fractogel DEAE-650-s, and Sepharose 6B columns. Molecular weight of protein determined by gel filtration and SDS-PAGE was 50 kDa. Naringinase of C. albidus includes high content of the dicarbonic and hydrophobic amino acids. Enzyme contains also carbohydrate component, represented by mannose, galactose, rhamnose, ribose, arabinose, xylose, and glucose. The enzyme was optimally active at pH 5.0 and 60 °C. Naringinase was found to exhibit specificity towards p-nitrophenyl-α-L-rhamnose, p-nitrophenyl-β-D-glucose, naringin, and neohesperidin. Its K towards naringin was 0.77 mM and the V was 36 U/mg. Naringinase was inhibited by high concentrations of reaction product-L-rhamnose. Enzyme revealed stability to 20% ethanol and 500 mM glucose in the reaction mixture that makes it possible to forecast its practical use in the food industry in the production of juices and wines.
柚皮苷酶是从白腐菌发酵液中提取的，经硫酸铵分级沉淀和 Toyopearl HW-60、Fractogel DEAE-650-s 和 Sepharose 6B 柱层析等方法纯化，比活提高 42 倍，回收率为 0.7%。凝胶过滤和 SDS-PAGE 测定酶的分子量为 50 kDa。白腐菌柚皮苷酶含有丰富的二羧酸和疏水性氨基酸。该酶还含有碳水化合物成分，如甘露糖、半乳糖、鼠李糖、核糖、阿拉伯糖、木糖和葡萄糖。该酶在 pH 5.0 和 60°C 时具有最佳活性。柚皮苷酶对 p-硝基苯-α-L-鼠李糖、p-硝基苯-β-D-葡萄糖、柚皮苷和新橙皮苷具有特异性。它对柚皮苷的 K 为 0.77 mM，V 为 36 U/mg。高浓度的反应产物-L-鼠李糖对柚皮苷酶有抑制作用。该酶在反应混合物中对 20%乙醇和 500 mM 葡萄糖具有稳定性，这使得它有可能在食品工业中用于生产果汁和葡萄酒。
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