1. 毕业设计（论文）的内容和要求

1．文献查阅 掌握文献查阅， 使用网络资源如中国期刊网、 维普数据库、 超星数字图书 馆、 elsevier 电子期刊、 springer link 全文电子期刊等检索资源， 查阅关于大丙酮酸氧化酶生物传感器方面的相关文献。

2． 文献阅读及综述 阅读与课题相关的外文与中文文献，了解国内外的研究动态， 撰写文献综 述。

3． 明确实验任务， 拟定实验方案 根据所查阅文献的内容， 确定实验内容及方案， 拟定科学可行的研究计划。

2. 参考文献

[1]Arai G , Noma T , Habu H , et al. Pyruvate sensor based on pyruvate oxidase immobilized in a poly(mercapto- p-benzoquinone) film[J]. Journal of Electroanalytical Chemistry, 1999, 464(2):143-148. [2]Bergmann W , Rudolph R , Spohn U . A bienzyme modified carbon paste electrode for amperometric detection of pyruvate[J]. Analytica Chimica Acta, 1999, 394(2):233-241. [3]Risse B , G#195;#188;nter Stempfer, Rudolph R , et al. Characterization of the stabilizing effect of point mutations of pyruvate oxidase from lactobacillus plantarum: Protection of the native state by modulating coenzyme binding and subunit interaction[J]. Protein Science, 1992, 1(12):1710-1718. [4]Gajovic N , Binyamin G , Warsinke A , et al. Operation of a Miniature Redox Hydrogel-Based Pyruvate Sensor in Undiluted Deoxygenated Calf Serum[J]. Analytical Chemistry, 2000, 72(13):2963-2968. [5]Gajovic N , Katja Haberm#252;ller, Warsinke A , et al. A Pyruvate Oxidase Electrode Based on an Electrochemically Deposited Redox Polymer[J]. Electroanalysis, 2015, 11(18):1377-1383. [6]Gavalas V G , Chaniotakis N A . Phosphate biosensor based on polyelectrolyte-stabilized pyruvate oxidase[J]. Analytica Chimica Acta, 2001, 427(2):271-277. [7]Sedewitz B , Schleifer K H , F G#246;tz. Physiological role of pyruvate oxidase in the aerobic metabolism of Lactobacillus plantarum.[J]. Journal of Bacteriology, 1984, 160(1):462-5. [8]Juan E C M , Hoque M M , Hossain M T , et al. The structures of pyruvate oxidase from Aerococcus viridans with cofactors and with a reaction intermediate reveal the flexibility of the active-site tunnel for catalysis[J]. Acta Crystallographica Section F Structural Biology and Crystallization Communications, 2007, 63(Pt 11):900-907. [9]Ikebukuro K , Wakamura H , Karube I , et al. Phosphate sensing system using pyruvate oxidase and chemiluminescence detection[J]. Biosensors Bioelectronics, 1996, 11(10):959-965. [10]Kulys J , Wang L , Daugvilaite N . Amperometric methylene green-mediated pyruvate electrode based on pyruvate oxidase entrapped in carbon paste[J]. Analytica Chimica Acta, 1992, 265(1):15-20. [11]Kwan R C H , Leung H F , Hon P Y T , et al. A screen-printed biosensor using pyruvate oxidase for rapid determination of phosphate in synthetic wastewater[J]. Applied Microbiology and Biotechnology, 2005, 66(4):377-383. [12]Mascini M , Mazzei F . Amperometric sensor for pyruvate with immobilized pyruvate oxidase[J]. Analytica Chimica Acta, 1987, 192(1):9-16. [13]Muller Y , Schulz G . Structure of the thiamine- and flavin-dependent enzyme pyruvate oxidase[J]. Science, 1993, 259(5097):965-967. [14]Muller Y A , G#252;nter Schumacher, Rudolph R , et al. The Refined Structures of a Stabilized Mutant and of Wild-type Pyruvate Oxidase from Lactobacillus plantarum[J]. Journal of Molecular Biology, 1994, 237(3):0-335. [15]Rahman M A , Park D S , Chang S C , et al. The biosensor based on the pyruvate oxidase modified conducting polymer for phosphate ions determinations[J]. Biosensors Bioelectronics, 2006, 21(7):1116-1124. [16]Risse B , G#195;#188;nter Stempfer, Rudolph R , et al. Stability and reconstitution of pyruvate oxidase from lactobacillus plantarum: Dissection of the stabilizing effects of coenzyme binding and subunit interaction[J]. Protein Science, 1992, 1(12):1699-1709.

3. 毕业设计（论文）进程安排

2018.12-2019.02 文献查阅， 完成开题报告， 熟悉实验仪器 2019.03-2019.04 异源表达不同微生物来源丙酮酸氧化酶并获得蛋白 2019.04-2019.05 进行酶学性质测定并对辅因子需求度进行浓度考察，计算酶活力并应用于电极表征。

2019.05-2019.06 补充验证实验， 整理实验数据，撰写毕业论文