Synthesis, biosynthesis, and characterization of transmembrane domains of a G protein-coupled receptor.
Peptide fragments have been widely used in biophysical studies on specific regions of integral membrane proteins. Because of their inherent insoluble nature and tendency to aggregate the preparation of such model peptides is challenging. We have developed synthetic and biosynthetic approaches to prepare peptides containing single and multiple domains of a G protein-coupled receptor. Both the synthetic and biosynthetic products can be isolated by reversed-phase high-performance liquid chromatography to near homogeneity. The biosynthetic product, a fusion protein, is processed by CNBr cleavage to yield the target peptide in various isotopic forms. The final peptides are studied by circular dichroism spectroscopy to determine their secondary structure under a variety of conditions.
Naider
Fred
F
Department of Chemistry, College of Staten Island, NY, USA.
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D002942:Circular Dichroism; D003488:Cyanogen Bromide; D015195:Drug Design; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D008958:Models, Molecular; D008967:Molecular Biology / Q000379:methods; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000096:biosynthesis / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D043562:Receptors, G-Protein-Coupled / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics; D051964:Receptors, Mating Factor / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics
D010446:Peptide Fragments; D043562:Receptors, G-Protein-Coupled; D051964:Receptors, Mating Factor; D011993:Recombinant Fusion Proteins; C498031:STE2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D003488:Cyanogen Bromide