Abstract
Cell surface expression of recombinant olfactory receptors (ORs) is a major limitation in characterizing their functional nature. We have shown that the recombinant expression of a human OR, OR 17-210, in the baculovirus/Sf9 insect cell system allows this protein to be expressed at the cell surface. We used Ca2+ imaging to demonstrate that recombinant OR 17-210 produces cellular activities upon odorant stimulation with ketones. Furthermore, this expression and functional system has been used to show that the preincubation of Human Odorant Binding Protein 2A decrease the calcium response of OR 17-210 following stimulation by acetophenone and beta ionone.
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References
Malnic B, Godfrey PA, Buck LB (2004) The human olfactory receptor gene family. Proc Natl Acad Sci U S A 101(8):2584–2589
Saito H et al (2009) Odor coding by a mammalian receptor repertoire. Sci Signal 2(60):ra9
Zhang X, Firestein S (2002) The olfactory receptor gene superfamily of the mouse. Nat Neurosci 5(2):124–133
Bush CF, Hall RA (2008) Olfactory receptor trafficking to the plasma membrane. Cell Mol Life Sci 65(15):2289–2295
McClintock TS, Sammeta N (2003) Trafficking prerogatives of olfactory receptors. Neuroreport 14(12):1547–1552
Malnic B (2007) Searching for the ligands of odorant receptors. Mol Neurobiol 35(2):175–181
Tcatchoff L et al (2006) A single lysyl residue defines the binding specificity of a human odorant-binding protein for aldehydes. FEBS Lett 580(8):2102–2108
Vaughn JL et al (1977) The establishment of two cell lines from the insect Spodoptera frugiperda (Lepidoptera; Noctuidae). In Vitro 13(4):213–217
Hu Y, Rajan L, Schilling WP (1994) Ca2+ signaling in Sf9 insect cells and the functional expression of a rat brain M5 muscarinic receptor. Am J Physiol 266(6 Pt 1):C1736–C1743
Raming K et al (1993) Cloning and expression of odorant receptors. Nature 361(6410):353–356
Riegel CI, Lanner-Herrera C, Slavicek JM (1994) Identification and characterization of the ecdysteroid UDP-glucosyltransferase gene of the Lymantria dispar multinucleocapsid nuclear polyhedrosis virus. J Gen Virol 75(Pt 4):829–838
Gaymard F et al (1996) The baculovirus/insect cell system as an alternative to Xenopus oocytes. First characterization of the AKT1 K + channel from Arabidopsis thaliana. J Biol Chem 271(37):22863–22870
Matarazzo V et al (2005) Functional characterization of two human olfactory receptors expressed in the baculovirus Sf9 insect cell system. Chem Senses 30(3):195–207
Poul MA et al (1995) Cassette baculovirus vectors for the production of chimeric, humanized, or human antibodies in insect cells. Eur J Immunol 25(7):2005–2009
Grynkiewicz G, Poenie M, Tsien RY (1985) A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem 260(6):3440–3450
Acknowledgments
The authors would like to thank Pr. Gérard DEVAUCHELLE (University of Montpellier, France), Dr Martine CERRUTI (CNRS, France) for virology experiments. We are also indebted to Dr. Thierry Thomas-Danguin and Dr Anne Tromlin (FLAVIC INRA, Dijon) for fruitful discussion on odorant mixtures and odor perception in humans. The authors are grateful to Dr Olivier Clot-Faybesse and Dr Brice Marcet for calcium imaging screening and Maryse Gremigni, Aurélie Seassau, and Nadia Elfangoud for experiments with hOBP-2. The work has been supported by a grant from a PNRA program of the French National Agency (ANR AROMALIM, coordinated by R. Gervais).
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Matarazzo, V., Ronin, C. (2013). Human Olfactory Receptors: Recombinant Expression in the Baculovirus/Sf9 Insect Cell System, Functional Characterization, and Odorant Identification. In: Crasto, C. (eds) Olfactory Receptors. Methods in Molecular Biology, vol 1003. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-377-0_8
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DOI: https://doi.org/10.1007/978-1-62703-377-0_8
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