Abstract
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.
Publication types
-
Research Support, Non-U.S. Gov't
MeSH terms
-
Alkaline Phosphatase / biosynthesis
-
Alkaline Phosphatase / isolation & purification
-
Animals
-
Cell Membrane / metabolism
-
Cell Membrane / ultrastructure
-
Cloning, Molecular
-
Electrophoresis, Polyacrylamide Gel
-
Enzyme-Linked Immunosorbent Assay
-
Escherichia coli
-
Immunoblotting
-
Isoproterenol / pharmacology
-
Mammals
-
Methionine / metabolism
-
Models, Structural
-
Molecular Sequence Data
-
Plasmids
-
Protein Conformation*
-
Receptors, Adrenergic, beta-1 / biosynthesis
-
Receptors, Adrenergic, beta-1 / chemistry*
-
Receptors, Adrenergic, beta-1 / isolation & purification
-
Recombinant Fusion Proteins / biosynthesis
-
Recombinant Fusion Proteins / chemistry
-
Recombinant Fusion Proteins / isolation & purification
-
Recombinant Proteins / biosynthesis
-
Recombinant Proteins / chemistry
-
Recombinant Proteins / isolation & purification
-
Sulfur Radioisotopes
Substances
-
Receptors, Adrenergic, beta-1
-
Recombinant Fusion Proteins
-
Recombinant Proteins
-
Sulfur Radioisotopes
-
Methionine
-
Alkaline Phosphatase
-
Isoproterenol