Monitoring the human beta1, beta2, beta3 adrenergic receptors expression and purification in Pichia pastoris using the fluorescence properties of the enhanced green fluorescent protein.
The three beta adrenergic receptor subtypes, beta1-, beta2- and beta3-, were expressed in the methylotrophic yeast Pichia pastoris. These receptors were N-terminally fused to the enhanced green fluorescent protein (EGFP) and the fluorescent properties of EGFP were used: (1) to select the recombinant strains, (2) to monitor the expression of the fluorescent receptors, and (3) to monitor the purification of the receptors by immobilized metal affinity chromatography. We demonstrate here that Pichia pastoris can be an alternative host to express and purify milligram amounts of human beta adrenergic receptors.
Talmont
Franck
F
Institut de Pharmacologie et de Biologie Structurale (UMR 5089), Centre National de la Recherche Scientifique-Université de Toulouse, 205 route de Narbonne, 31077 Toulouse Cedex 4, France. franck.talmont@ipbs.fr
D005453:Fluorescence; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D007700:Kinetics; D010843:Pichia / Q000378:metabolism*; D011943:Receptors, Adrenergic, beta / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D018342:Receptors, Adrenergic, beta-1 / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D018343:Receptors, Adrenergic, beta-2 / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D022702:Receptors, Adrenergic, beta-3 / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012835:Silver Nitrate; D012995:Solubility
D011943:Receptors, Adrenergic, beta; D018342:Receptors, Adrenergic, beta-1; D018343:Receptors, Adrenergic, beta-2; D022702:Receptors, Adrenergic, beta-3; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D012835:Silver Nitrate