Purification and enzymatic properties of Auricularia auricula tyrosinase
-
摘要: 本文分离纯化了黑木耳酪氨酸酶,并对酶学性质进行了研究。采用经硫酸铵分级沉淀、Sephadex G-100和DEAE-Sepharose-FF柱层析对粗酶液进行分离纯化,得到电泳纯的黑木耳酪氨酸酶,比活力提高了21.43倍,酶活回收率为27.41%。该酶的酶学性质研究表明:蛋白亚基分子量为12.62ku;最适pH7.0,在中性和碱性条件下稳定;最适温度为40℃,50℃以下温度条件较为稳定;以酪氨酸为底物,米氏常数K m为5.88mmol/L,V max为64.10μmol/min。实验结果表明黑木耳酪氨酸酶具有其它酪氨酸酶相似的酶学特性。Abstract: Auricularia auricula (A.auricula) tyrosinase was purified and its enzymatic properties were investigated.By (NH 4) 2 SO 4 fractional precipitation, Sephadex G-100 and DEAE-Sepharose-FF column separation, electrophoretic pure A.auricula tyrosinase was obtained and the specific activity increased 21.43 fold, the enzyme recovery was 27.41%.The experimental results of enzymatic properties showed the molecular weight of protein subunit was 12.62ku.Optimal pH was 7.0 and tyrosinase was stable under neutral and alkaline conditions.Optimal temperature was 40℃ and tyrosinase exhibited a good heat stability when temperature below 50℃.When tyrosine was used as substrate, the K m was 5.88mmol /L and the V max was 64.10μmol /min.The experimental results indicated the enzymatic properties of A.auricula tyrosinase were very similar to those of other tyrosinase.
-
Keywords:
- Auricularia auricula /
- tyrosinase /
- purification /
- enzymatic properties
-
[1] Lezzi C, Bleve G, Spagnolo S, et al.Production of recombinant Agaricus bisporus tyrosinase in Saccharomyces cerevisiae cells[J].Journal Industrial Microbiology and Biotechnology, 2012, 39:1875-1880.
[2] Gasparetti C, Faccio G, Arvas M, et al.Discovery of a new tyrosinase-like enzyme family lacking a C-terminally processed domain:production and characterization of an Aspergillus oryzae catechol oxidase[J].Applied Microbiology and Biotechnology, 2010, 86:213-226.
[3] Falguera V, Pagan J, Ibarz A.A kinetic model describing melanin formation by means of mushroom tyrosinase[J].Food Research International, 2010, 43 (1) :66-69.
[4] Watensson G, Hagberg S, Anderson E, et al.Parkinson’s disease in diphenyl exposed workers-a causal association[J].Parkinsonism and Related Disorders, 2006, 12:29-34.
[5] Chandel M, Azmi W.Optimization of process parameters for the production of tyrosine phenol lyase by Citrobacter freundii MTCC 2424[J].Bioresource Technology, 2009, 100:1840-1846.
[6] Zou Y, Yang Y, Zeng B, et al.Comparison of Physicochemical Properties and Antioxidant Activities of Melanins from FruitBodies and Fermentation Broths of Auricularia auricular[J].International Journal of Food Properties, 2013, 16:803-813.
[7] 王学奎.植物生理生化实验原理和技术[M].北京:高等教育出版社, 2006:190-192. [8] Dalfard AB, Khajeh K, Soudi MR, et al.Isolation and biochemical characterization of laccase and tyrosinase activities in a novel melanogenic soil bacterium[J].Enzyme and Microbial Technology, 2006, 39:1409-1416.
[9] Selinheimo E, Nieidhin D, Steffensen C, et al.Comparison of the characteristics of fungal and plant tyrosinases[J].Journal of Biotechnology, 2007, 130:471-480.
[10] 王君玲, 口如琴, 成汇, 等.马铃薯酪氨酸酶的部分纯化及理化性质的研究[J].山西大学学报, 1995, 18 (2) :184-189. [11] Wang X, Chai B, Liao Y, et al.Molecular and biochemical characterization of a distinct tyrosinase involved in melanin production from Aeromonas media[J].Applied Microbiology and Biotechnology, 2009, l82:261-269.
[12] 樊廷俊, 汪小锋.中国对虾 (Penaeus chinensis) 酚氧化酶的分离纯化及其部分生物化学性质[J].生物化学与生物物理学报, 2002, 34 (5) :589-594. [13] Muoz-Muoz J L, Garcia-Molina F, Berna J, et al.Kinetic characterisation of o-aminophenols and aromatic o-diamines as suicide substrates of tyrosinase[J].Biochimica et Biophysica Acta, 2012, 1824:647-655.
[14] Cabrera-Valladares N, Martinez A, Pinero S, et al.Expression of the melA gene from Rhizobium etli CFN42 in Escherichia coli and characterization of the encoded tyrosinase[J].Enzyme and Microbial Technology, 2006, 38:772-779.
[15] 张晓晴, 蔡宇杰, 廖祥儒, 等.香樟 (Cinnamomum camphora) 果实酪氨酸酶的分离纯化及特性分析[J].安徽农业科学, 2008, 36 (15) :6167-6170.
计量
- 文章访问数: 160
- HTML全文浏览量: 16
- PDF下载量: 481
下载:
