用于单分子动力学实验的微流控混合器

doi:10.3866/PKU.WHXB20110804

Acta Phys. -Chim. Sin. ›› 2011, Vol. 27 ›› Issue (08): 1990-1995.doi: 10.3866/PKU.WHXB20110804

• BIOPHYSICAL CHEMISTRY • Previous Articles Next Articles

A Microfluidic Mixer for Single-Molecule Kinetics Experiments

ZHI Ze-Yong^1,3, LIU Peng-Cheng^1,3, HUANG Yan-Yi^2,3, ZHAO Xin-Sheng^1,3

1. Beijing National Laboratory for Molecular Sciences, State Key Laboratory for Structural Chemistry of Unstable and Stable Species, Department of Chemical Biology, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, P. R. China;
2. College of Engineering, Peking University, Beijing 100871, P. R. China;
3. Biodynamic Optical Imaging Center, Peking University, Beijing 100871, P. R. China

Received:2011-04-27 Revised:2011-06-01 Published:2011-07-19
Contact: ZHAO Xin-Sheng E-mail:zhaoxs@pku.edu.cn
Supported by:
The project was supported by the National Natural Science Foundation of China (20733001, 20973015) and National Key Basic Research Program of China (973) (2006CB910300, 2010CB912302).

Abstract

Abstract:

We designed and built a microfluidic mixer based on the principle of hydrodynamic focusing governed by Navier-Stokes equation for single-molecule kinetics experiments. The mixer is a cast of poly(dimethylsiloxane) (PDMS) sealed with transparent fused-silica coverglass, which results in low fluorescence background and broad biological compatibility and this enables single-molecule fluorescence detection under nonequilibrium conditions. The pressure regulated sample delivery system is convenient for loading a sample and allows for precise and stable flow velocity control. The combination of microfluidic mixer and single-molecule fluorescence resonance energy transfer (smFRET) allows us to measure the time course of the distribution of the smFRET efficiency in protein folding. We used the fact that denatured protein collapses much faster than the mixing process to characterize the mixing time using donor and acceptor dyes labeled staphylococcal nuclease (SNase) as an smFRET efficiency indicator. By monitoring the smFRET efficiency of denatured SNase during the course of mixing, we determined that the mixing time was 150 ms under conditions suitable for single-molecule detection.

Key words: Microfluidic mixing, Single-molecule detection, Fluorescence resonance energy transfer, Protein folding, Staphylococcal nuclease

MSC2000:

O643

ZHI Ze-Yong, LIU Peng-Cheng, HUANG Yan-Yi, ZHAO Xin-Sheng. A Microfluidic Mixer for Single-Molecule Kinetics Experiments[J].Acta Phys. -Chim. Sin., 2011, 27(08): 1990-1995.

References

(1) Wolynes, P. G.; Onuchic, J. N.; Thirumalai, D. Science 1995, 267, 1619.
(2) Oliveberg, M.;Wolynes, P. G. Q. Rev. Biophys. 2005, 38, 245.
(3) Ferreon, A. C. M.; Deniz, A. A. BBA-Proteins Proteomics 2011, In Press.
(4) Haas, E. ChemPhysChem 2005, 6, 858.
(5) Bilsel, O.; Matthews, C. R. Curr. Opin. Struct. Biol. 2006, 16, 86.
(6) Ha, T.; Enderle, T.; Ogletree, D. F.; Chemla, D. S.; Selvin, P. R.; Weiss, S. Proc. Natl. Acad. Sci. U. S. A. 1996, 93, 6264.
(7) Weiss, S. Science 1999, 283, 1676.
(8) Deniz, A. A.; Mukhopadhyay, S.; Lemke, E. A. J. R. Soc. Interface 2008, 5, 15.
(9) Deniz, A. A.; Laurence, T. A.; Beligere, G. S.; Dahan, M.; Martin, A. B.; Chemla, D. S.; Dawson, P. E.; Schultz, P. G.; Weiss, S. Proc. Natl. Acad. Sci. U. S. A. 2000, 97, 5179.
(10) Schuler, B.; Lipman, E. A.; Eaton,W. A. Nature 2002, 419, 743.
(11) Hoffmann, A.; Kane, A.; Nettels, D.; Hertzog, D. E.; Baumgartel, P.; Lengefeld, J.; Reichardt, G.; Horsley, D. A.; Seckler, R.; Bakajin, O.; Schuler, B. Proc. Natl. Acad. Sci. U. S. A. 2007, 104, 105.
(12) Muller-Spath, S.; Soranno, A.; Hirschfeld, V.; Hofmann, H.; Ruegger, S.; Reymond, L.; Nettels, D.; Schuler, B. Proc. Natl. Acad. Sci. U. S. A. 2010, 107, 14609.
(13) Hertzog, D. E.; Michalet, X.; Jager, M.; Kong, X. X.; Santiago, J. G.;Weiss, S.; Bakajin, O. Anal. Chem. 2004, 76, 7169.
(14) Hertzog, D. E.; Ivorra, B.; Mohammadi, B.; Bakajin, O.; Santiago, J. G. Anal. Chem. 2006, 78, 4299.
(15) Park, H. Y.; Qiu, X. Y.; Rhoades, E.; Korlach, J.; Kwok, L.W.; Zipfel,W. R.;Webb,W.W.; Pollack, L. Anal. Chem. 2006, 78, 4465.
(16) Lapidus, L. J.; Yao, S. H.; McGarrity, K. S.; Hertzog, D. E.; Tubman, E.; Bakajin, O. Biophys. J. 2007, 93, 218.
(17) Park, H. Y.; Kim, S. A.; Korlach, J.; Rhoades, E.; Kwok, L.W.; Zipfell,W. R.;Waxham, M. N.;Webb,W.W.; Pollack, L. Proc. Natl. Acad. Sci. U. S. A. 2008, 105, 542.
(18) Guo, S.; Xue, M. Q.; Qian, M. X.; Cao, T. B.; Zhao, X. S. Acta Phys. -Chim. Sin. 2007, 23, 1827. [郭索, 薛面起, 钱民协, 曹廷炳, 赵新生. 物理化学学报, 2007, 23, 1827.]
(19) Knight, J. B.; Vishwanath, A.; Brody, J. P.; Austin, R. H. Phys. Rev. Lett. 1998, 80, 3863.
(20) Hamadani, K. M.;Weiss, S. Biophys. J. 2008, 95, 352.
(21) Pfeil, S. H.;Wickersham, C. E.; Hoffmann, A.; Lipman, E. A. Rev. Sci. Instrum. 2009, 80, 055105.
(22) Gambin, Y.; VanDelinder, V.; Ferreon, A. C. M.; Lemke, E. A.; Groisman, A.; Deniz, A. A. Nat. Methods 2011, 8, 239.
(23) Maki, K.; Cheng, H.; Dolgikh, D. A.; Roder, H. J. Mol. Biol. 2007, 368, 244.
(24) Lipman, E. A.; Schuler, B.; Bakajin, O.; Eaton,W. A. Science 2003, 301, 1233.
(25) Ye, K. Q.;Wang, J. F. J. Mol. Biol. 2001, 307, 309.
(26) Kim, S. J.; Blainey, P. C.; Schroeder, C. M.; Xie, X. S. Nat. Methods 2007, 4, 397.
(27) Krichevsky, O.; Bonnet, G. Rep. Prog. Phys. 2002, 65, 251.
(28) Chen, X. D.; Zhou, Y.; Qu, P.; Zhao, X. S. J. Am. Chem. Soc. 2008, 130, 16947.
(29) Sherman, E.; Haran, G. Proc. Natl. Acad. Sci. U. S. A. 2006, 103, 11539.
(30) Liu, P. C.; Meng, X. L.; Qu, P.; Zhao, X. S.;Wang, C. C. J. Phys. Chem. B 2009, 113, 12030.
(31) White, F. Viscous Fluid Flow, 2nd ed.; McGraw Hill: Boston, Massachusetts, 1991.
(32) Gosch, M.; Blom, H.; Holm, J.; Heino, T.; Rigler, R. Anal. Chem. 2000, 72, 3260.
(33) Kuricheti, K. K.; Buschmann, V.;Weston, K. D. Appl. Spectrosc. 2004, 58, 1180.
(34) Nie, S. M.; Chiu, D. T.; Zare, R. N. Anal. Chem. 1995, 67, 2849.
(35) Gell, C.; Brockwell, D.; Smith, A. Handbook of Single Molecule Fluorescence Spectroscopy; Oxford University: Oxford, 2006.
(36) Nettels, D.; Gopich, I. V.; Hoffmann, A.; Schuler, B. Proc. Natl. Acad. Sci. U. S. A. 2007, 104, 2655.

[1]	Aoneng Cao. "Confined Lowest Energy Structure Fragments (CLESFs)" Hypothesis for Protein Structure and the "Stone Age" of Protein Prebiotic Evolution [J]. Acta Physico-Chimica Sinica, 2020, 36(1): 1907002-0.
[2]	LI Yong, ZHOU Wei, DAI Zhi-Jun, CHEN Yuan, WANG Zhi-Ming, YUAN Zhe-Ming. Predicting the Protein Folding Rate Based on Sequence Feature Screening and Support Vector Regression [J]. Acta Phys. -Chim. Sin., 2014, 30(6): 1091-1098.
[3]	XU Ze-Qing, GAO Bao-Jiao, HOU Xiao-Dong. Twofold Influence of Nitro Substituent on Aromatic Ring for Photoluminescence Properties of Benzoic Acid-Functionalized Polystyrene and Eu(Ⅲ) Complexes [J]. Acta Phys. -Chim. Sin., 2014, 30(4): 745-752.
[4]	ZHANG Zhu-Qing. Folding Mechanism of De novo Designed Proteins [J]. Acta Phys. -Chim. Sin., 2012, 28(10): 2381-2389.
[5]	HUANG Yong-Qi, LIU Zhi-Rong. Intrinsically Disordered Proteins: the New [J]. Acta Phys. -Chim. Sin., 2010, 26(08): 2061-2072.
[6]	GAO Meng, YAO Xin-Qiu, SHE Zhen-Su, LIU Zhi-Rong, ZHU Huai-Qiu. Intermediate Structure and Slow HydrationWater Dynamics in Protein Folding Process [J]. Acta Phys. -Chim. Sin., 2010, 26(07): 1998-2006.
[7]	ZHOU Wen-Hua, YING Li-Ming. Single Molecule Fluorescence Resonance Energy Transfer and Ensemble Biophysical Characterization of a G-quadruplex Formed in the Promoter of Human Myocyte Enhancer Factor 2D [J]. Acta Phys. -Chim. Sin., 2010, 26(04): 1099-1106.
[8]	WANG Zheng, LAI Bing, CAO Jie, LI Zhu, QU Li-Li, CAO Ao-Neng, LAI Lu-Hua. Hierarchical Unfolding of Mj HSP16.5 [J]. Acta Phys. -Chim. Sin., 2008, 24(10): 1745-1750.
[9]	Liu Ting-Ting, Luo Guo-Bin, Ying Li-Ming, Zhao Xia-Sheng. Single-Molecule Detection Studies of Diffusion of Rhodamine-labeled Phospholipids [J]. Acta Phys. -Chim. Sin., 2000, 16(05): 393-397.

A Microfluidic Mixer for Single-Molecule Kinetics Experiments

PDF (PC)

Like

Knowledge

Abstract

Cite this article

share this article

References

Related Articles 9

Metrics

Comments

Recommended 0