Purification of the surface cAMP receptor in Dictyostelium

Journal Article · Mon Jan 05 04:00:00 EST 1987 · J. Biol. Chem.; (United States)

OSTI ID:6860023

Klein, P; Knox, B; Borleis, J; Devreotes, P

We have previously identified and demonstrated reversible ligand-induced modification of the major cell surface cAMP receptor in Dictyostelium discoideum. The receptor, or a subunit of it, has been purified to homogeneity by hydroxylapatite chromatography followed by two-dimensional preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The purification was monitored by following /sup 32/Pi incorporated by photoaffinity labeling with 8-azido-(/sup 32/P)cAMP or by in vivo labeling with /sup 32/Pi. Two interconvertible forms of the receptor, designated R (Mr 40,000) and D (Mr 43,000), co-purified. Two-dimensional peptide maps of independently purified and /sup 125/I-iodinated R and D forms of the receptor were nearly identical but did have several distinct peptides. The estimated 6000-fold purification required is consistent with the number of cell surface binding sites assuming there are not multiple binding sites/polypeptide. In the accompanying article we report the generation of a monospecific polyclonal antiserum which has helped to further elucidate the physical properties and developmental regulation of the cAMP receptor.

• Research Organization: Johns Hopkins Univ. School of Medicine, Baltimore, MD
• OSTI ID: 6860023
• Journal Information: J. Biol. Chem.; (United States), Journal Name: J. Biol. Chem.; (United States) Vol. 1; ISSN JBCHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMP
AZIDES
CHEMISTRY
CHROMATOGRAPHY
CHYMOTRYPSIN
ENZYMES
FUNGI
HYDROLASES
LIGANDS
MEMBRANE PROTEINS
MYXOMYCETES
NITROGEN COMPOUNDS
NUCLEOTIDES
ORGANIC COMPOUNDS
PEPTIDE HYDROLASES
PHOTOCHEMISTRY
PLANTS
PROTEINS
PURIFICATION
RECEPTORS
SEPARATION PROCESSES
SERINE PROTEINASES