The formylpeptide chemoattractant receptor copurifies with a GTP-binding protein containing a distinct 40-kDa pertussis toxin substrate.
Detergent extraction of plasma membranes from differentiated HL60 cells, specifically labeled with the chemoattractant, formyl-Nle-Leu-Phe-Nle-[125I-Tyr] Lys, resulted in the solubilization of a receptor-radioligand complex. GTP-binding activity coeluted with the radioligand when the sodium cholate extract was purified by chromatography on wheat germ agglutinin-Sepharose 6MB. A molecular size of approximately 59 A was estimated for the lectin-Sepharose-purified receptor complex by gel filtration chromatography on Ultrogel AcA 34. The isolated complex eluted from the gel filtration column exhibited an enhanced rate of ligand dissociation in response to GTP gamma S. Approximately 0.65 mol of pertussis toxin substrate/mol of receptor was estimated following partial purification of the receptor-ligand complex by sequential chromatography on wheat germ agglutinin-Sepharose, DEAE-Fractogel, and Ultrogel AcA 34. The pertussis toxin substrate which copurified with the receptor was compared with two distinct G proteins, containing alpha-subunits of 40 and 41 kDa, previously purified from HL60 cell plasma membranes. Approximately 86% of the pertussis toxin substrate identified in the receptor preparation consisted of the 40-kDa polypeptide. Differences in the peptide maps indicate that the predominant G protein which coelutes with the receptor is distinct from the purified G protein with an alpha-subunit of 41 kDa but homologous to the purified G protein with an alpha-subunit of 40 kDa.
Polakis
P G
PG
Howard Hughes Medical Institute, Durham, North Carolina.
Uhing
R J
RJ
Snyderman
R
R
D000266:Adenylyl Imidodiphosphate / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000276:immunology / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000302:isolation & purification* / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008970:Molecular Weight; D009842:Oligopeptides / Q000378:metabolism* / Q000494:pharmacology; D037342:Pertussis Toxin; D044042:Receptors, Formyl Peptide; D011971:Receptors, Immunologic / Q000302:isolation & purification* / Q000378:metabolism; D013873:Thionucleotides / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000378:metabolism*
D009842:Oligopeptides; D044042:Receptors, Formyl Peptide; D011971:Receptors, Immunologic; D013873:Thionucleotides; D010566:Virulence Factors, Bordetella; D000266:Adenylyl Imidodiphosphate; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins