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Photochemical properties and sensor applications of modified yellow fluorescent protein (YFP) covalently attached to the surfaces of etched optical fibers (EOFs)

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Abstract

Fluorescent proteins have the inherent ability to act as sensing components which function both in vitro and inside living cells. We describe here a novel study on a covalent site-specific bonding of fluorescent proteins to form self-assembled monolayers (SAMs) on the surface of etched optical fibers (EOFs). Deposition of fluorescent proteins on EOFs gives the opportunity to increase the interaction of guided light with deposited molecules relative to plane glass surfaces. The EOF modification is carried out by surface activation using 3-aminopropylthrimethoxysilane (APTMS) and bifunctional crosslinker sulfosuccinimidyl 4-[N-maleimidomethyl]cyclohexane-1-carboxylate (sulfo-SMCC) which exposes sulfhydryl-reactive maleimide groups followed by covalent site-specific coupling of modified yellow fluorescent protein (YFP). Steady-state and fluorescence lifetime measurements confirm the formation of SAM. The sensor applications of YPF SAMs on EOF are demonstrated by the gradual increase of emission intensity upon addition of Ca2+ ions in the concentration range from a few tens of micromolars up to a few tens of millimolars. The studies on the effect of pH, divalent cations, denaturing agents, and proteases reveal the stability of YFP on EOFs at normal physiological conditions. However, treatments with 0.5% SDS at pH 8.5 and protease trypsin are found to denaturate or cleave the YFP from fiber surfaces.

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References

  1. Lippincott-Schwartz J, Patterson GH (2003) Development and use of fluorescent protein markers in living cells. Science 300(5616):87–91

    Article  CAS  Google Scholar 

  2. Shaner NC, Steinbach PA, Tsien RY (2005) A guide to choosing fluorescent proteins. Nat Methods 2(12):905–909

    Article  CAS  Google Scholar 

  3. Mishin AS, Subach FV, Yampolsky IV, King W, Lukyanov KA, Verkhusha VV (2008) The first mutant of the Aequorea victoria green fluorescent protein that forms a red chromophore. Biochemistry 47(16):4666–4673

    Article  CAS  Google Scholar 

  4. Jayaraman S, Haggie P, Wachter RM, Remington SJ, Verkman AS (2000) Mechanism and cellular applications of a green fluorescent protein-based halide sensor. J Biol Chem 275:6047–6050

    Article  CAS  Google Scholar 

  5. Griesbeck O, Baird GS, Campbell RE, Zacharias DA, Tsien RY (2001) Reducing the environmental sensitivity of yellow fluorescent protein. J Biol Chem 276(31):29188–29194

    Article  CAS  Google Scholar 

  6. Nakai J, Ohkura M, Imoto K (2001) A high signal-to-noise Ca2+ probe composed of a single green fluorescent protein. Nat Biotechnol 19(2):137–141

    Article  CAS  Google Scholar 

  7. Hanson GT, McAnaney TB, Park ES, Rendell MEP, Yarbrough DK, Chu S, Xi L, Boxer SG, Montrose MH, Remington SJ (2002) Green fluorescent protein variants as ratiometric dual emission pH sensors. 1. Structural characterization and preliminary application. Biochemistry 41(52):15477–15488

    Article  CAS  Google Scholar 

  8. Hanson GT, Aggeler R, Oglesbee D, Cannon M, Capaldi RA, Tsien RY, Remington SJ (2004) Investigating mitochondrial redox potential with redox-sensitive green fluorescent protein indicators. J Biol Chem 279(13):13044–13053

    Article  CAS  Google Scholar 

  9. Siemering KR, Golbik R, Sever R, Haseloff J (1996) Mutations that suppress the thermosensitivity of green fluorescent protein. Curr Biol 6(12):1653–1663

    Article  CAS  Google Scholar 

  10. Chudakov DM, Lukyanov S, Lukyanov KA (2005) Fluorescent proteins as a toolkit for in vivo imaging. Trends Biotechnol 23:605–613

    Article  CAS  Google Scholar 

  11. Abraham BG, Tkachenko NV, Santala V, Lemmetyinen H, Karp M (2011) Bidirectional fluorescence resonance energy transfer (FRET) in mutated and chemically modified yellow fluorescent protein (YFP). Bioconjug Chem 22:227–234

    Article  CAS  Google Scholar 

  12. Zhang J, Campbell RE, Ting AY, Tsien RY (2002) Creating new fluorescent probes for cell biology. Nat Rev Mol Cell Biol 3(12):906–918

    Article  CAS  Google Scholar 

  13. Deuschle K, Okumoto S, Fehr M, Looger LL, Kozhukh L, Frommer WB (2005) Construction and optimization of a family of genetically encoded metabolite sensors by semirational protein engineering. Protein Sci 14:2304–2314

    Article  CAS  Google Scholar 

  14. Dittmer PJ, Miranda JG, Gorski JA, Palmer AE (2009) Genetically encoded sensors to elucidate spatial distribution of cellular zinc. J Biol Chem 284(24):16289–16297

    Article  CAS  Google Scholar 

  15. Tanaka Y, Tsuruda Y, Nishi M, Kamiya N, Goto M (2007) Exploring enzymatic catalysis at a solid surface: a case study with transglutaminase-mediated protein immobilization. Org Biomol Chem 5:1764–1770

    Article  CAS  Google Scholar 

  16. Kwon Y, Coleman MA, Camarero JA (2006) Selective immobilization of proteins onto solid supports through split-intein-mediated protein trans-splicing. Angew Chem Int Edit 45:1726–1729

    Article  CAS  Google Scholar 

  17. Yin J, Liu F, Li X, Walsh CT (2004) Labeling proteins with small molecules by site-specific posttranslational modification. J Am Chem Soc 126:7754–7755

    Article  CAS  Google Scholar 

  18. Kufer SK, Dietz H, Albrecht C, Blank K, Kardinal A, Rief M, Gaub HE (2005) Covalent immobilization of recombinant fusion proteins with hAGT for single molecule force spectroscopy. Eur Biophys J 35:72–78

    Article  CAS  Google Scholar 

  19. Wong LS, Khan F, Micklefield J (2009) Selective covalent protein immobilization: strategies and applications. Chem Rev 109:4025–4053

    Article  CAS  Google Scholar 

  20. Gauchet C, Labadie GR, Poulter CD (2006) Regio- and chemoselective covalent immobilization of proteins through unnatural amino acids. J Am Chem Soc 128:9274–9275

    Article  CAS  Google Scholar 

  21. Lapiene V, Kukolka F, Kiko K, Arndt A, Niemeyer CM (2010) Conjugation of fluorescent proteins with DNA oligonucleotides. Bioconjugate Chem 21:921–927

    Article  CAS  Google Scholar 

  22. Lin P, Ueng S, Tseng M, Ko J, Huang K, Yu S, Adak AK, Chen Y, Lin C (2006) Site-specific protein modification through CuI-catalyzed 1,2,3-triazole formation and its implementation in protein microarray fabrication. Angew Chem Int Edit 45:4286–4290

    Article  CAS  Google Scholar 

  23. Wu P, Hogrebe P, Grainger DW (2006) DNA and protein microarray printing on silicon nitride waveguide surfaces. Biosens Bioelectron 21:1252

    Article  CAS  Google Scholar 

  24. Jung Y, Jeong ML, Jung H et al (2007) Self-directed and self-oriented immobilization of antibody by protein G-DNA conjugate. Anal Chem 79:6534

    Article  CAS  Google Scholar 

  25. Liu Z, Tabakman SM, Chen Z et al (2009) Preparation of carbon nanotube bioconjugates for biomedical applications. Nat Protoc 4:1372

    Article  CAS  Google Scholar 

  26. Vo-Dinh T, Kasili P, Wabuyele M (2006) Nanoprobes and nanobiosensors for monitoring and imaging individual living cells. Nanomed: NBM 2(1):22–30

    Article  CAS  Google Scholar 

  27. Veselov A, Thür C, Efimov A, Guina M, Lemmetyinen H, Tkachenko N (2010) Acidity sensor based on porphyrin self-assembled monolayers covalently attached to the surfaces of tapered fibres. Meas Sci Technol 21:115205–115215

    Article  Google Scholar 

  28. Rusu M, Kivistö S, Gawith CBE, Okhotnikov OG (2005) Red-green-blue (RGB) light generator using tapered fiber pumped with a frequency-doubled Yb-fiber laser. Opt Express 13(21):8547–8554

    Article  CAS  Google Scholar 

  29. Corres JM, Matias IR, Bravo J, Arregui FJ (2008) Tapered optical fiber biosensor for the detection of anti-gliadin antibodies. Sens Actuators B 135:166–171

    Article  Google Scholar 

  30. Hoffmann P, Dutoit B, Salathé R (1995) Comparison of mechanically drawn and protection layer chemically etched optical fiber tips. Ultramicroscopy 61:165–170

    Article  CAS  Google Scholar 

  31. Haddock HS, Shankar PM, Mutharasan R (2003) Fabrication of biconical tapered optical fibers using hydrofluoric acid. Mater Sci Eng B 97:87–93

    Article  Google Scholar 

  32. Khalil S, Bansal L, El-Sherif M (2004) Intrinsic fiber optic chemical sensor for the detection of dimethyl methylphosphonate. Opt Eng 43(11):2683–2688

    Article  CAS  Google Scholar 

  33. Santala V, Lamminmäki U (2004) Production of a biotinylated single-chain antibody fragment in the cytoplasm of Escherichia coli. J Immunol Methods 284(1–2):165–175

    Article  CAS  Google Scholar 

  34. Suzuki M, Ito Y, Savage HE, Husimi Y, Douglas KT (2003) Intramolecular fluorescent resonance energy transfer (FRET) by BODIPY chemical modification of cysteine-engineered mutants of green fluorescent protein. Chem Lett 32:306–307

    Article  CAS  Google Scholar 

  35. Yang F, Moss LG, Phillips GN (1996) The molecular structure of green fluorescent protein. Nat Biotechnol 14(10):1246–1251

    Article  CAS  Google Scholar 

  36. Ormö M, Cubitt AB, Kallio K, Gross LA, Tsien RY, Remington SJ (1996) Crystal structure of the Aequorea victoria green fluorescent protein. Science 273:1392–1395

    Article  Google Scholar 

  37. Ishii M, Kunimura J, Jeng H, Penna T, Cholewa O (2007) Evaluation of the pH- and thermal stability of the recombinant green fluorescent protein (GFP) in the presence of sodium chloride. Appl Biochem Biotechnol 137–140(1):555–571

    Article  Google Scholar 

  38. Campbell TN, Choy FYM (2001) The effect of pH on green fluorescent protein: a brief review. Mol Biol Today 2:1–4

    CAS  Google Scholar 

  39. Alkaabi KM, Yafea A, Ashraf SS (2005) Effect of pH on thermal- and chemical-induced denaturation of GFP. Appl Biochem Biotechnol 126:149–156

    Article  CAS  Google Scholar 

  40. Saeed IA, Ashraf SS (2009) Denaturation studies reveal significant differences between GFP and blue fluorescent protein. Int J Biol Macromol 45(3):236–241

    Article  CAS  Google Scholar 

  41. Chiang C, Okou DT, Griffin TB, Verret CR, Williams MNV (2001) Green fluorescent protein rendered susceptible to proteolysis: positions for protease-sensitive insertions. Arch Biochem Biophys 394:229–235

    Article  CAS  Google Scholar 

  42. Bokman SH, Ward WW (1981) Renaturation of green-fluorescent protein. Biochem Biophys Res Commun 101:1372–1380

    Article  CAS  Google Scholar 

  43. Patterson GH, Knobel SM, Sharif WD, Kain SR, Piston DW (1997) Use of the green fluorescent protein and its mutants in quantitative fluorescence microscopy. Biophys J 73(5):2782–2790

    Article  CAS  Google Scholar 

  44. Elsliger M, Wachter RM, Hanson GT, Kallio K, Remington SJ (1999) Structural and spectral response of green fluorescent protein variants to changes in pH. Biochemistry 38(17):5296–5301

    Article  CAS  Google Scholar 

  45. Zou J, Hofer AM, Lurtz MM, Gadda G, Ellis AL, Chen N, Huang Y, Holder A, Ye Y, Louis CF, Welshhans K, Rehder V, Yang JJ (2007) Developing sensors for real-time measurement of high Ca2+ concentrations. Biochemistry (N Y) 46:12275–12288

    Article  CAS  Google Scholar 

  46. Schäferling M, Duerkop A (2008) In: Resch-Genger U (ed) Standardization and quality assurance in fluorescence measurements 1. Springer, Heidelberg, Series on Fluorescence

    Google Scholar 

  47. Malcik N, Ferrance JP, Landers JP, Caglar P (2005) The performance of a microchip-based fiber optic detection technique for the determination of Ca2+ ions in urine. Sens Act B 107:24–31

    Article  Google Scholar 

  48. Caglar P, Tuncel SA, Malcik N, Landers JP, Ferrance JP (2006) A microchip sensor for calcium determination. Anal Bioanal Chem 386:1303–1312

    Article  CAS  Google Scholar 

  49. Sloan WD, Uttamlal M (2001) A fibre-optic calcium ion sensor using a calcein derivative. Luminescence 16:179

    Article  CAS  Google Scholar 

  50. Miyawaki A, Llopis J, Heim R, McCaffery JM, Adams JA, Ikura M, Tsien RY (1997) Fluorescent indicators for Ca2+ based on green fluorescent proteins and calmodulin. Nature 388:882–887

    Article  CAS  Google Scholar 

  51. Miyawaki A, Griesbeck O, Heim R, Tsien RY (1999) Dynamic and quantitative Ca2+ measurements using improved cameleons. Proc Natl Acad Sci U S A 96(5):2135–2140

    Article  CAS  Google Scholar 

  52. Truong K, Sawano A, Miyawaki A, Ikura M (2007) Calcium indicators based on calmodulin-fluorescent protein fusions. Methods Mol Biol 352:71–82

    CAS  Google Scholar 

  53. Tsien RY (1998) The green fluorescent protein. Annu Rev Biochem 67:509–544

    Article  CAS  Google Scholar 

  54. Vo-Dinh T, Alarie J, Cullum BM, Griffin GD (2000) Antibody-based nanoprobe for measurement of a fluorescent analyte in a single cell. Nat Biotechnol 18(7):764–767

    Article  CAS  Google Scholar 

  55. Olsen JV, Ong S, Mann M (2004) Trypsin cleaves exclusively C-terminal to arginine and lysine residues. Mol Cell Proteomics 3(6):608–614

    Article  CAS  Google Scholar 

  56. Gasteiger E, Gattiker A, Hoogland C, Ivanyi I, Appel RD, Bairoch A (2003) ExPASy: the proteomics server for in-depth protein knowledge and analysis. Nucleic Acids Res 31:3784–3788

    Article  CAS  Google Scholar 

  57. Betzig E, Chichester RJ (1993) Single molecules observed by near-field scanning optical microscopy. Science 262:1422–1425

    Article  CAS  Google Scholar 

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Acknowledgments

This study was funded by the Academy of Finland within the frameworks of ActiveFiber, Äärimikro, and BUSU projects, The National Doctoral Programme in Nanoscience, and Biomedical Image Quantification/University Alliance Finland. The authors thank Dr. Alexander Efimov (Department of Chemistry and Bioengineering, Tampere University of Technology, Finland) for helpful discussions and Prof. R.Y. Tsien (HHMI-UCSD La Jolla, CA, USA) for providing the citrine gene.

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  1. Department of Chemistry and Bioengineering, Tampere University of Technology, P.O. Box 541, 33101, Tampere, Finland

    Alexey A. Veselov, Bobin George Abraham, Helge Lemmetyinen, Matti T. Karp & Nikolai V. Tkachenko

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  1. Alexey A. Veselov
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  2. Bobin George Abraham
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  3. Helge Lemmetyinen
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Correspondence to Alexey A. Veselov.

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Alexey A. Veselov and Bobin George Abraham contributed equally to this work.

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Veselov, A.A., Abraham, B.G., Lemmetyinen, H. et al. Photochemical properties and sensor applications of modified yellow fluorescent protein (YFP) covalently attached to the surfaces of etched optical fibers (EOFs). Anal Bioanal Chem 402, 1149–1158 (2012). https://doi.org/10.1007/s00216-011-5564-4

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