Expression of functional melanocortin 1 receptors in insect cells

Helgi Schiöth, Arja Kuusinen, Ruta Muceniece, Michael Szardenings, Kari Keinänen, Jarl Wikberg

Research output: Contribution to journalArticleScientificpeer-review

26 Citations (Scopus)

Abstract

We expressed epitope-tagged human melanocortin 1 receptor (MC1R) in insect cells using two different recombinant baculovirus constructs; one of which encoded MC1R with an N-terminal Flag epitope and a C-terminal polyHis tag, while the other encoded the MC1R with a C-terminal Myc tag. The constructs were used to infect Sf9 insect cells. For both constructs, immunoblotting with tag-specific antibodies demonstrated the presence of the receptor in the infected cells. The infected Sf9 cells were characterized by radioligand binding using [125I][Nle4, D-Phe7]α-MSH. Both saturation and competition analysis, using α-, β-, and γ1-MSH on the tagged MC1R expressed in the insect cells, gave binding constants and potency orders that were undistinguishable from those obtained on MC1R expressed in COS cells. The expression level obtained (in the order of pmoles of binding sites per mg of protein) will now facilitate attempts to purify the receptor. This is the first report that demonstrates a functional expression of recombinant melanocortin receptor in nonmammalian cells.
Original languageEnglish
Pages (from-to)807-814
JournalBiochemical and Biophysical Research Communications
Volume221
Issue number3
DOIs
Publication statusPublished - 1996
MoE publication typeA1 Journal article-refereed

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