1. 毕业设计（论文）的内容和要求

四氢嘧啶和5-羟化四氢嘧啶作为相容性物质，不仅可以调节渗透压，还可以稳定蛋白结构，在医药、生物制造和化工行业具有广阔的发展前景。

四氢嘧啶羟化酶（ectd）属于fe 2 #160;与2-酮戊二酸依赖型双加氧酶超家族，ectd在催化反应中体现作用底物的专一性，主要催化四氢嘧啶生成5-羟化四氢嘧啶，是微生物发酵生产5-羟化四氢嘧啶过程中的关键酶。

我们将就ectd的克隆表达与性质研究进行综合分析和研究。

2. 参考文献

[1]Reuter K,Pittelkow M,Bursy J,et al.Synthesis of 5-hy#8259;droxyectoine from ectoine:crystal structure of the non-heme iron(II)and 2-oxoglutarate-dependent dioxygenase EctD[J].PloS One,2010,5(5):e10647. [2]Bursy J,Pierik A J,Pica N,et al.Osmotically induced synthesis of the compatible solute hydroxyectoine is mediated by an evolutionarily conserved ectoine hydroxylase[J].J Biol Chem,2007,282(43):31147-31155. [3]Smiatek J,Harishchandra R K,Galla H J,et al.Low concentrated hydroxyectoine solutions in presence of DPPC lipid bilayers:a computer simulation study[J].Biophys Chem,2013,180-181:102-109. [4]]Bursy J,Kuhlmann A U,Pittelkow M,et al.Synthesis and uptake of the compatible solutes ectoine and 5-hydroxyectoine by StreptomycescoelicolorA3(2)in responsetosalt and heatstresses[J].ApplEnviron Microbiol,2008,74(23):7286-7296. [5]朱道辰,刘杏荣.相容性溶质四氢嘧啶及其羟基化衍生物的研究进展[J].中国生物工程杂志,2011,31(2):95-101 [6]Garcia-Estepa R,Argandona M,Reina-Bueno M,et al.The ectD gene,which is involved in the synthesis of the compatible solute hydroxyectoine,is essential for thermoprotection of the halophilic bacterium Chromohalobacter salexigens[J].J Bacteriol,2006,188(11):3774-3784. [7]Smiatek J,Harishchandra R K,Galla H J,et al.Low concen#8259;trated hydroxyectoine solutions in presence of DPPC lipid bilayers:a computer simulation study[J].Biophys Chem,2013,180-181:102-109. [8]Van-Thuoc D,Hashim S O,Hatti-Kaul R,et al.Ectoine-mediated protection of enzyme from the effect of pH and temperature stress:a study using Bacillus halodurans xylanase as a model[J].Appl Microbiol Biotechnol,2013,97 (14):6271-6278. [9]Kanapathipillai M,Lentzen G,Sierks M,et al.Ectoine and hydroxyectoine inhibitaggregation and neurotoxicity ofAlzheimer's beta-amyloid[J].FEBS Lett,2005,579(21):4775-4780. [10]Harishchandra R K,Sachan A K,Kerth A,et al.Compatible solutes:ectoine and hydroxyectoine improve functional nanostructures in artificial lung surfactants[J].Biochim Biophys Acta,2011,1808(12):2830-2840. [11]Schnoor M,Voss P,Cullen P,et al.Characterization of the synthetic compatible solute homoectoine as a potent PCR enhancer[J].Biophys Res Commun,2004,322(3):867-872. [12]Reuter K,Pittelkow M,Bursy J,et al.Synthesis of 5-hydroxyectoine from ectoine:crystal structure of the non-heme iron(II)and 2-oxoglutarate-dependent dioxygenase EctD[J].PloS One,2010,5(5):e10647. [13]Shibasaki T,Mori H,Chiba S,et al.Microbial proline 4-hydroxylase screening and gene cloning[J].Appl Environ Microbi#8259;ol,1999,65(9):4028-4031. [14]何健,黄星,顾立峰,等.盐单胞菌属BYS-1四氢嘧啶合成基因ectABC克隆及其盐激表达[J].微生物学报,2006,46(1):28-32. [15]Eilert E,Kranz A,Hollenberg C P,et al.Synthesis and release of the bacterial compatible solute 5-hydroxyectoine in Hansenula polymorpha[J].J Biotechnol,2013,167(2):85-93. [16]Rodriguez-Moya J,Argandona M,Iglesias-Guerra F,et al.Temperature-and salinity-decoupled overproduction ofhydroxyectoine by Chromohalobacter salexigens[J].Appl Environ Microbiol,2013,79(3):1018-1023. [17]李岩,董雷,王磊,等.极端耐盐放线菌白色普氏菌YIM90005T四氢嘧啶及5-羟基四氢嘧啶合成相关基因的克隆[J].微生物学报,2011,51(5):603-608. [18]Klein C,H#252;ttel W.A simple procedure for selective hydroxyl-ation of L-proline and L-pipecolic acid with recombinantly expressed proline hydroxylases[J].Adv Synthesis Catalysis,2011,353(8):1375-1383. [19]Shibasaki T,Mori H,Ozaki A.Enzymatic production of trans-4-hydroxy-L-proline by regio- and stereospecific hydroxyl-ation of L-proline[J].Biosci Biotechnol Biochem,2000,64(4):746-750. [20]Widderich N,Pittelkow M,Hoppner A,et al.Molecular dy#8259;namics simulations and structure-guided mutagenesis provide insight into the architecture of the catalytic core of the ec#8259;toine hydroxylase[J].J Mol Biol,2014,426(3):586-600.

3. 毕业设计（论文）进程安排

1月#8212;#8212;2月：查阅文献，设计实验 3月#8212;#8212;5月：进行实验 5月之后：进行论文写作