Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/95996
標題: Design the protein switches based on the kinetics of protein folding
以蛋白質動力學特性為基礎來設計一系列蛋白質開關
作者: Shen-Yi Zheng
鄭慎禕
關鍵字: 蛋白質摺疊動力學
Rapidly unfolding/folding (RUF)
螢光蛋白
金黃色葡萄球菌protein A
IgG binding domains
螢光共振能量轉移效率(FRET efficiency)
Kinetics of protein folding
Rapidly unfolding/folding (RUF)
fluorescent protein
staphylococcus aureus protein A
IgG binding domains
FRET efficiency
引用: 1. Ulmer, Kevin M. 'Protein engineering.' Science 219.4585 (1983): 666-671. 2. Brannigan, James A., and Anthony J. Wilkinson. 'Protein engineering 20 years on.' Nature Reviews Molecular Cell Biology 3.12 (2002): 964-970. 3. Bornscheuer, U. T., et al. 'Engineering the third wave of biocatalysis.' Nature 485.7397 (2012): 185-194. 4. Davids, Timo, et al. 'Strategies for the discovery and engineering of enzymes for biocatalysis.' Current opinion in chemical biology 17.2 (2013): 215-220. 5. Bloom, Jesse D., et al. 'Evolving strategies for enzyme engineering.' Current opinion in structural biology 15.4 (2005): 447-452. 6. Ho, Steffan N., et al. 'Site-directed mutagenesis by overlap extension using the polymerase chain reaction.' Gene 77.1 (1989): 51-59. 7. Shaner, Nathan C., Paul A. Steinbach, and Roger Y. Tsien. 'A guide to choosing fluorescent proteins.' Nature methods 2.12 (2005): 905-909. 8. Tsien, Roger Y. 'The green fluorescent protein.' Annual review of biochemistry 67.1 (1998): 509-544. 9. Zhou, Xin X., and Michael Z. Lin. 'Photoswitchable fluorescent proteins: ten years of colorful chemistry and exciting applications.' Current opinion in chemical biology 17.4 (2013): 682-690. 10. Tavare, J. M., L. M. Fletcher, and G. I. Welsh. 'Using green fluorescent protein to study intracellular signalling.' Journal of Endocrinology 170.2 (2001): 297-306. 11. Sapra, Puja, and Boris Shor. 'Monoclonal antibody-based therapies in cancer: advances and challenges.' Pharmacology & therapeutics 138.3 (2013): 452-469. 12. Bourne, David G., et al. 'Enzymatic pathway for the bacterial degradation of the cyanobacterial cyclic peptide toxin microcystin LR.' Applied and Environmental microbiology 62.11 (1996): 4086-4094. 13. Jäckel, Christian, Peter Kast, and Donald Hilvert. 'Protein design by directed evolution.' Annu. Rev. Biophys. 37 (2008): 153-173. 14. Nicholls, Ian A., et al. 'Theoretical and computational strategies for rational molecularly imprinted polymer design.' Biosensors and Bioelectronics 25.3 (2009): 543-552. 15. Kaur, Jasjeet, and Rohit Sharma. 'Directed evolution: an approach to engineer enzymes.' Critical reviews in biotechnology 26.3 (2006): 165-199. 16. Matthews, B. W., H. Nicholson, and W. J. Becktel. 'Enhanced protein thermostability from site-directed mutations that decrease the entropy of unfolding.' Proceedings of the National Academy of Sciences 84.19 (1987): 6663-6667. 17. Zhu, Guo Ping, et al. 'Increasing the thermostability of D-xylose isomerase by introduction of a proline into the turn of a random coil.' Protein engineering 12.8 (1999): 635-638. 18. Zheng, Lei, Ulrich Baumann, and Jean-Louis Reymond. 'An efficient one-step site-directed and site-saturation mutagenesis protocol.' Nucleic acids research 32.14 (2004): e115-e115. 19. Steiner, Kerstin, and Helmut Schwab. 'Recent advances in rational approaches for enzyme engineering.' Computational and structural biotechnology journal 2.3 (2012): 1-12. 20. Strohmeier, Gernot A., et al. 'Application of designed enzymes in organic synthesis.' Chemical reviews 111.7 (2011): 4141-4164. 21. Ha, Jeung‐Hoi, and Stewart N. Loh. 'Protein conformational switches: from nature to design.' Chemistry–A European Journal 18.26 (2012): 7984-7999. 22. Ritterson, Ryan S., et al. 'Design of a photoswitchable cadherin.' Journal of the American Chemical Society 135.34 (2013): 12516-12519. 23. Huang, Jin, and Shohei Koide. 'Rational conversion of affinity reagents into label-free sensors for peptide motifs by designed allostery.' ACS chemical biology 5.3 (2010): 273-277. 24. Ha, Jeung-Hoi, et al. 'Engineering domain-swapped binding interfaces by mutually exclusive folding.' Journal of molecular biology 416.4 (2012): 495-502. 25. Ha, Jeung-Hoi, et al. 'Engineered Domain Swapping as an On/Off Switch for Protein Function.' Chemistry & biology 22.10 (2015): 1384-1393. 26. Harper, Shannon M., Lori C. Neil, and Kevin H. Gardner. 'Structural basis of a phototropin light switch.' Science 301.5639 (2003): 1541-1544. 27. Berezhkovskii, Alexander, Gerhard Hummer, and Attila Szabo. 'Reactive flux and folding pathways in network models of coarse-grained protein dynamics.'The Journal of chemical physics 130.20 (2009): 205102. 28. Van Der Meer, B. Wieb, George Coker, and S-Y. Simon Chen. Resonance energy transfer: theory and data. Wiley-VCH, 1994. 29. Merkx, Maarten, et al. 'Rational design of FRET sensor proteins based on mutually exclusive domain interactions.' Biochemical Society Transactions 41.5 (2013): 1201-1205. 30. Nguyen, Annalee W., and Patrick S. Daugherty. 'Evolutionary optimization of fluorescent proteins for intracellular FRET.' Nature biotechnology 23.3 (2005): 355-360. 31. Talaga, David S., et al. 'Dynamics and folding of single two-stranded coiled-coil peptides studied by fluorescent energy transfer confocal microscopy.' Proceedings of the National Academy of Sciences 97.24 (2000): 13021-13026. 32. Oikawa, Hiroyuki, et al. 'Complexity of the Folding Transition of the B Domain of Protein A Revealed by the High-Speed Tracking of Single-Molecule Fluorescence Time Series.' The Journal of Physical Chemistry B 119.20 (2015): 6081-6091. 33. Chung, Chan-I., et al. 'Open Flower Fluoroimmunoassay: A general method to make fluorescent protein-based immunosensor probes.' Analytical chemistry 87.6 (2015): 3513-3519. 34. Schulenburg, C., et al. 'A FRET-based biosensor for the detection of neutrophil elastase.' Analyst 141.5 (2016): 1645-1648. 35. Giepmans, Ben NG, et al. 'The fluorescent toolbox for assessing protein location and function.' science 312.5771 (2006): 217-224. 36. Medintz, Igor L., et al. 'Self-assembled nanoscale biosensors based on quantum dot FRET donors.' Nature materials 2.9 (2003): 630-638. 37. Gould, Travis J., et al. 'Nanoscale imaging of molecular positions and anisotropies.' Nature methods 5.12 (2008): 1027-1030. 38. Vu, Dung M., et al. 'Probing the folding and unfolding dynamics of secondary and tertiary structures in a three-helix bundle protein.' Biochemistry 43.12 (2004): 3582-3589. 39. Arora, Pooja, Terrence G. Oas, and Jeffrey K. Myers. 'Fast and faster: A designed variant of the B‐domain of protein A folds in 3 μsec.' Protein Science 13.4 (2004): 847-853. 40. Arora, Pooja, Gordon G. Hammes, and Terrence G. Oas. 'Folding mechanism of a multiple independently-folding domain protein: double B domain of protein A.' Biochemistry 45.40 (2006): 12312-12324. 41. Capp, Jo A., et al. 'The statistical conformation of a highly flexible protein: small-angle X-ray scattering of S. aureus protein A.' Structure 22.8 (2014): 1184-1195. 42. Deis, Lindsay N., et al. 'Multiscale conformational heterogeneity in staphylococcal protein a: possible determinant of functional plasticity.' Structure 22.10 (2014): 1467-1477. 43. Ansbacher, Tamar, et al. 'Calculation of transition dipole moment in fluorescent proteins—towards efficient energy transfer.' Physical Chemistry Chemical Physics 14.12 (2012): 4109-4117. 44. Lindenburg, Laurens H., et al. 'Quantifying stickiness: thermodynamic characterization of intramolecular domain interactions to guide the design of Forster resonance energy transfer sensors.' Biochemistry 53.40 (2014): 6370-6381. 45. Dickson, Robert M., et al. 'On/off blinking and switching behaviour of single molecules of green fluorescent protein.' Nature 388.6640 (1997): 355-358. 46. Sato, Satoshi, et al. 'Testing protein-folding simulations by experiment: B domain of protein A.' Proceedings of the National Academy of Sciences of the United States of America 101.18 (2004): 6952-6956. 47. Pham, Kara, K. S. LaForge, and M. J. Kreek. 'Sticky-end PCR: new method for subcloning.' Biotechniques 25 (1998): 206-208. 48. Patterson, George H., et al. 'Use of the green fluorescent protein and its mutants in quantitative fluorescence microscopy.' Biophysical journal 73.5 (1997): 2782. 49. Vogel, Steven S., Christopher Thaler, and Srinagesh V. Koushik. 'Fanciful fret.' Sci. STKE 2006.331 (2006): re2-re2. 50. Schuler, Benjamin, and William A. Eaton. 'Protein folding studied by single-molecule FRET.' Current opinion in structural biology 18.1 (2008): 16-26. 51. Ha, Taekjip. 'Single-molecule methods leap ahead.' Nature methods 11.10 (2014): 1015. 52. Huang, Jie-rong, et al. 'Stable intermediate states and high energy barriers in the unfolding of GFP.' Journal of molecular biology 370.2 (2007): 356-371. 53. Yoo, Tae Yeon, et al. 'Small-angle X-ray scattering and single-molecule FRET spectroscopy produce highly divergent views of the low-denaturant unfolded state.' Journal of molecular biology 418.3 (2012): 226-236. 54. Grünberg, Raik, et al. 'Engineering of weak helper interactions for high-efficiency FRET probes.' Nature methods 10.10 (2013): 1021-1027.
摘要: 以蛋白質工程學的原理為基礎來設計功能性蛋白質已被廣泛應用在許多研究領域之中,而在許多蛋白質設計的策略裏蛋白質開關(protein-switch) 於近幾年更有許多成功的實際應用。此種蛋白質開關的設計原理主要為利用蛋白質分子因「熱力學穩定度」的變化而能在兩種構型之間來回切換,用以執行不同的功能。有趣的是,有別於熱力學穩定度的應用,過去並無文獻使用蛋白質「摺疊動力學」來設計蛋白質開關,而本研究即在探討將折疊動動力學性質應用在蛋白質設計的可能性。金黃色葡萄球菌(Staphylococcus aureus)表面蛋白質¬¬protein A有五個具rapidly unfolding / folding (RUF) 特性的IgG binding domains,因為具有過渡態(transition state)自由能偏低特色,可以從摺疊態(folded state)自行經歷過渡態到不具有構型的解摺疊態(unfolded state),再回到摺疊態,本實驗即使用上述的其中兩個domain,做為以蛋白質摺疊動力學為基礎設計蛋白質開關的模型。於本研究中,我們成功的將螢光蛋白CyPet和YPet構築在選定的domain兩端,搭配螢光共振能量轉移(Förster resonance energy transfer,FRET)的技術進行蛋白質折疊動力學的定量觀測。比較一系列設計的融合蛋白之能量轉移效率(FRET efficiency),我們發現FRET efficiency會隨CyPet以及YPet之間domain的折疊動力學性質而有明顯的變化。另外,我們也觀察到不同的domain串連個數也會對FRET efficiency有明顯的改變。此研究的結果或可作為將來蛋白質功能設計上的新策略以及依據。
Protein engineering is widely used in various research fields and industrial applications. Among different engineering strategies, protein switch gets more and more attention in recent years. The basic principle of designing protein-switch is to alter the conformation of proteins according to the difference of 'thermodynamic stability'. Protein switch is a powerful tool used in designing biosensor, biomaterial or controllable-enzyme. Interestingly, folding kinetics is another characteristic of protein conformation, based on our literature search there is no application of this characteristic on designing protein-switch. Therefore we seek to demonstrate the possibility of applying protein-folding kinetics on the design of protein-switch. Protein A is a surface protein of staphylococcus aureus, which has five IgG binding domains on its N-termius. These domains were selected as our test model because of their special property with rapidly unfolding and folding (RUF) kinetics. In order to test this RUF kinetics effect on the protein engineering, the fluorescent proteins CyPet and YPet were fused on the two sides of designated domains. We then observed the kinetic effect of these domains by Förster Resonance Energy Transfer (FRET). Our results indicate that RUF kinetics effect can still be observed by fusion with flanked proteins. Meanwhile, the FRET efficiency of designed constructs was changed according to the difference of RUF characteristic. This identification can potentially be used as a new tool in the strategies of designing protein function.
URI: http://hdl.handle.net/11455/95996
文章公開時間: 10000-01-01
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