Purification, cloning, and tissue distribution of a 23-kDa rat protein isolated by morphine affinity chromatography

David K. Grandy, Eric Hanneman, James Bunzow, Marjorie Shih, Curtis A. Machida, Jean M. Bidlack, Olivier Civelli

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Abstract

A 23-kDa (p23k) rat brain protein was stereospecifically eluted from a 14β-bromoacetamidomorphine affinity column, purified to apparent homogeneity by reverse phase HPLC, and partially sequenced. Three degenerate oligodeoxynucleotide probes were synthesized based on this partial amino acid sequence. A rat brain cDNA library was screened using these probes, and a full-length cDNA was isolated. The deduced protein, 187 amino acids long, is rich in glutamic and aspartic acid residues, endowing p23k with a net negative charge at neutral pH. The protein lacks a signal sequence as well as any transmembrane domains. Based on predictions of secondary structure, p23k is a globular protein composed of 30% α-helices and 18% β-pleated sheets. Northern blot analysis revealed p23k transcripts in rat brain, liver, and the mouse x rat neuroblastoma-glioma NG108-14 cell line. Although not an opioid receptor itself, this protein may be associated with such a receptor or be related to a protein that has been shown to be cross-linked to the opioid peptide β-endorphin.

Original languageEnglish (US)
Pages (from-to)1370-1376
Number of pages7
JournalMolecular Endocrinology
Volume4
Issue number9
DOIs
StatePublished - Sep 1990

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ASJC Scopus subject areas

  • Molecular Biology
  • Endocrinology

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