1. 毕业设计（论文）的内容和要求

论文内容： 本实验采用基因工程手段，筛选出具有活性ω转氨酶，以大肠杆菌为宿主诱导表达产酶，测出其酶活，然后通过定点突变方法对其进行分子改造，筛选出活性更高的酶。

论文要求： 1.掌握文献查阅的一般方法，学会在中国期刊网、web of science科学引文索引、springer link电子期刊、elsevier sdos电子期刊等检索资源上查阅关于生物催化槲皮素研究的相关文献，并对大肠杆菌表达系统有全面了解。

（2）文献阅读及综述：阅读与课题相关的中英文文献，了解国内外的研究动态，撰写文献综述。

2. 参考文献

[1] REHN G, ADLERCREUTZ P, GREY C. Supported liquid membrane as a novel tool for driving the equilibrium of ω-transaminase catalyzed asymmetric synthesis [J]. Journal of Biotechnology, 2014, 179(1): 50-5. [2] MATHEW S, NADARAJAN S P, CHUNG T, et al. Biochemical characterization of thermostable ω-transaminase from Sphaerobacter thermophilus and its application for producing aromatic β- and γ-amino acids [J]. Enzyme Microbial Technology, 2016, 87-88(52. [3] SAVILE C K, JANEY J M, MUNDORFF E C, et al. Biocatalytic asymmetric synthesis of chiral amines from ketones applied to sitagliptin manufacture [J]. Science, 2010, 329(5989): 305-9. [4] FESKO K, STEINER K, BREINBAUER R, et al. Investigation of one-enzyme systems in the ω-transaminase-catalyzed synthesis of chiral amines [J]. Journal of Molecular Catalysis B Enzymatic, 2013, 96(4): 103-10. [5] BEA H, SEO Y, YUN H. Production of Enantiomerically Pure Unnatural Amino acids Using Omega-transaminase [J]. Analytical Bioanalytical Chemistry, 2011, 407(9): 2455-62. [6] HOU A, DENG Z, MA H, et al. Substrate screening of amino transaminase for the synthesis of a sitagliptin intermediate [J]. Tetrahedron, 2016, 72(31): 4660-4. [7] XIA W, SUN Y, MIN C, et al. [Asymmetric synthesis of aromatic L-amino acids catalyzed by transaminase] [J]. Chinese Journal of Biotechnology, 2012, 28(11): 1346. [8] PARK E S, DONG J Y, SHIN J S. ω-Transaminase-catalyzed asymmetric synthesis of unnatural amino acids using isopropylamine as an amino donor [J]. Organic Biomolecular Chemistry, 2013, 11(40): 6929-33. [9] Enantioselective Enzymatic Desymmetrizations in Organic Synthesis (p ) [J]. Cheminform, 2010, 36(18): [10] KOSZELEWSKI D, LAVANDERA I, CLAY D, et al. Asymmetric Synthesis of Optically Pure Pharmacologically Relevant Amines Employing ω‐Transaminases [J]. Advanced Synthesis Catalysis, 2008, 350(17): 2761-6. [11] CERIOLI L, PLANCHESTAINER M, CASSIDY J, et al. Characterization of a novel amine transaminase from Halomonas elongata [J]. Journal of Molecular Catalysis B Enzymatic, 2015, 120(10): 141-50. [12] MATHEW S, DEEPANKUMAR K, SHIN G, et al. Identification of novel thermostable ω-transaminase and its application for enzymatic synthesis of chiral amines at high temperature [J]. #54620;#44397;#49373;#47932;#44277;#54617;#54924; #54617;#49696;#45824;#54924;, 2016, 6(73): [13] JIANG J, CHEN X, FENG J, et al. Substrate profile of an ω-transaminase from Burkholderia vietnamiensis and its potential for the production of optically pure amines and unnatural amino acids [J]. Journal of Molecular Catalysis B Enzymatic, 2014, 100(32-9. [14] KAULMANN U, SMITHIES K, SMITH M E B, et al. Substrate spectrum of ω-transaminase from Chromobacterium violaceum DSM30191 and its potential for biocatalysis [J]. Enzyme Microbial Technology, 2007, 41(5): 628-37. [15] ZHU W, LI Y, JIA H, et al. Expression, purification and characterization of a thermostable leucine dehydrogenase from the halophilic thermophile Laceyella sacchari [J]. Biotechnology Letters, 2016, 38(5): 855-61. [16] RICHTER N, SIMON R C, LECHNER H, et al. #207;#8240;-Transaminases for the amination of functionalised cyclic ketones [J]. Organic Biomolecular Chemistry, 2015, 13(33): 8843-51. [17] SHIN G, MATHEW S, YUN H. Kinetic resolution of amines by ( R )-selective omega-transaminase from Mycobacterium vanbaalenii [J]. Journal of Industrial Engineering Chemistry, 2015, 23(128-33.

3. 毕业设计（论文）进程安排

2018.1.11-2018.2.24 查阅文献，熟悉课题有关背景，合理设计。

实验方案。

2018.2.25-2018.3.25 将ω转氨酶导入大肠杆菌中，采用iptg 2018.3.26-2018.5.26 利用相关软件比对转氨酶序列，找出适当的突变位点，采用定点突变技术对该酶进行分子改造筛选突变体。