Format

Send to

Choose Destination
See comment in PubMed Commons below
Chem Senses. 2012 Nov;37(9):837-47. doi: 10.1093/chemse/bjs067. Epub 2012 Sep 5.

Functional reconstitution of olfactory receptor complex on baculovirus.

Author information

  • 1Department of Molecular Biology and Medicine, Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Tokyo 153-8904, Japan.

Abstract

Despite that recent progress in genomics has elucidated the genomic structure of the olfactory receptors (ORs), most of them are still orphan receptors. The low expression level of ORs in heterologous cells has hampered many attempts to establish cell biological OR assay systems. Recently, we demonstrated that certain G protein-coupled receptors, such as the leukotriene B4 receptor or the dopamine D1 receptor, were efficiently reconstituted on baculovirus budding from infected Sf9 cells. The budded virus (BV) was shown to be mostly free of exogenous proteins other than those related to viral infection, resulting in low-noise assay conditions. Taking advantage of these conditions, we attempted to reconstitute OR complexes on BV. Sf9 cells were coinfected with recombinant baculoviruses harboring the cDNAs encoding adenylyl cyclase, trimeric G-protein, and the receptor: mOR-EG or S6. The coexpression of these proteins was detected by western blot, and the agonist- or antagonist-dependent receptor response was confirmed using ligand-dependent cyclic AMP production. These results demonstrated the successful reconstitution of functional OR complex on BV. Additionally, the expression of OR8B3 on BV, one of human orphan ORs, was also confirmed. This BV expression system is expected to be a highly effective tool for screening unknown ligands for ORs.

[PubMed - indexed for MEDLINE]
Free full text
PubMed Commons home

PubMed Commons

0 comments
How to join PubMed Commons

    Supplemental Content

    Full text links

    Icon for HighWire
    Loading ...
    Write to the Help Desk