Analysis of ligand-receptor cross-linked fragments by mass spectrometry

Journal Article · Sat Jan 01 00:00:00 EST 2005 · Journal of Peptide Research, The

DOI:https://doi.org/10.1111/j.1399-3011.2005.00248.x· OSTI ID:1005206

Son, C. D. ^[1]; Sargsyan, H. ^[2]; Naider, F. ^[2]; Becker, J. M. ^[1]

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1. University of Tennessee, Knoxville (UTK)
2. City University of New York (CUNY)

G-protein coupled receptors (GPCRs) are a class of integral membrane receptor proteins that are characterized by a signature seven-transmembrane (7-TM) configuration. The a-factor receptor (Ste2p) from Saccharomyces cerevisiae is a GPCR that, upon binding of a peptide ligand, transduces a signal to initiate a cascade of events leading to the mating of haploid yeast cells. This study summarizes the application of affinity purification and of matrix-assisted laser-desorption ionization time-of-flight (MALDI-TOF) experiments using biotinylated photoactivatable a-factor analogs. Affinity purification and enrichment of biotinylated peptides by monomeric avidin beads resulted in mass spectrometric detection of specific signals corresponding to crosslinked fragments of Ste2p. Data obtained from cyanogen bromide (CNBr) fragments of receptor cross-linked to an a-factor analog with the photoaffinity group p-benzoyl-L-phenylalanine on position 1 were in agreement with the previous results reported by our laboratory suggesting the cross-linking between position 1 of a-factor and a region of Ste2p covering residues 251 294.

• Research Organization: Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
• Sponsoring Organization: USDOE Office of Science (SC)
• DOE Contract Number: DE-AC05-00OR22725
• OSTI ID: 1005206
• Journal Information: Journal of Peptide Research, The, Vol. 65, Issue 3; ISSN 1397--002X
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
99 GENERAL AND MISCELLANEOUS//MATHEMATICS, COMPUTING, AND INFORMATION SCIENCE
AFFINITY
AVIDIN
BROMIDES
CONFIGURATION
CROSS-LINKING
CYANOGEN
DETECTION
GTP-ASES
IONIZATION
MASS SPECTROSCOPY
MATING
MEMBRANES
PEPTIDES
PROTEINS
PURIFICATION
RESIDUES
SACCHAROMYCES CEREVISIAE
YEASTS