title: A general chemical method to regulate protein stability in the mammalian central nervous system.
abstract: The ability to make specific perturbations to biological molecules in a cell or organism is a central experimental strategy in modern research biology. We have developed a general technique in which the stability of a specific protein is regulated by a cell-permeable small molecule. Mutants of the Escherichia coli dihydrofolate reductase (ecDHFR) were engineered to be degraded, and, when this destabilizing domain is fused to a protein of interest, its instability is conferred to the fused protein resulting in rapid degradation of the entire fusion protein. A small-molecule ligand trimethoprim (TMP) stabilizes the destabilizing domain in a rapid, reversible, and dose-dependent manner, and protein levels in the absence of TMP are barely detectable. The ability of TMP to cross the blood-brain barrier enables the tunable regulation of proteins expressed in the mammalian central nervous system.
authors.affiliation: Department of Chemical and Systems Biology, Stanford University, CA 94305, USA.
authors.forename: Mari
authors.initials: M
authors.lastname: Iwamoto
authors.forename: Tomas
authors.initials: T
authors.lastname: Björklund
authors.forename: Cecilia
authors.initials: C
authors.lastname: Lundberg
authors.forename: Deniz
authors.initials: D
authors.lastname: Kirik
authors.forename: Thomas J
authors.initials: TJ
authors.lastname: Wandless
mesh_terms: D000818:Animals; D001921:Brain / Q000378:metabolism*; D002460:Cell Line; D004926:Escherichia coli / Q000201:enzymology; D005493:Folic Acid Antagonists / Q000737:chemistry* / Q000494:pharmacology; D008024:Ligands; D051379:Mice; D015202:Protein Engineering; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D022021:Tacrolimus Binding Proteins / Q000737:chemistry / Q000378:metabolism; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014295:Trimethoprim / Q000737:chemistry* / Q000494:pharmacology
chemical_list: D005493:Folic Acid Antagonists; D008024:Ligands; D011993:Recombinant Fusion Proteins; D014295:Trimethoprim; D013762:Tetrahydrofolate Dehydrogenase; D022021:Tacrolimus Binding Proteins