Topological analysis of the human beta 2-adrenergic receptor expressed in Escherichia coli.
We have investigated the topology of the human beta 2-adrenergic receptor expressed in Escherichia coli, using the genetic method described by Beckwith and coworkers. We found that fusions with alkaline phosphatase beyond a certain point on the human beta 2-adrenergic receptor sequence were assembled into the bacterial membrane with the same topology as the human beta 2-adrenergic receptor in the mammalian membrane. The pattern that might have been expected on the basis of the topology of the human beta 2-adrenergic receptor in mammalian membranes was not reflected in the levels of alkaline phosphatase activity of the fusions occurring between the N-terminal region and positions close to the second external domain. Our data suggest that the correct positioning of the N terminus of the receptor depends on the presence of its C-terminal portions.
Lacatena
R M
RM
EniChem, Rome, Italy.
Cellini
A
A
Scavizzi
F
F
Tocchini-Valentini
G P
GP
D000469:Alkaline Phosphatase / Q000096:biosynthesis / Q000302:isolation & purification; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli; D015151:Immunoblotting; D007545:Isoproterenol / Q000494:pharmacology; D008322:Mammals; D008715:Methionine / Q000378:metabolism; D008961:Models, Structural; D008969:Molecular Sequence Data; D010957:Plasmids; D011487:Protein Conformation; D018342:Receptors, Adrenergic, beta-1 / Q000096:biosynthesis / Q000737:chemistry* / Q000302:isolation & purification; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D013462:Sulfur Radioisotopes
D018342:Receptors, Adrenergic, beta-1; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D013462:Sulfur Radioisotopes; D008715:Methionine; D000469:Alkaline Phosphatase; D007545:Isoproterenol