Saturation transfer difference nuclear magnetic resonance spectroscopy as a method for screening proteins for anesthetic binding.
The effects of anesthetics on cellular function may result from direct interactions between anesthetic molecules and proteins. These interactions have a low affinity and are difficult to characterize. To identify proteins that bind anesthetics, we used nuclear magnetic resonance saturation transfer difference (STD) spectroscopy. The method is based on the nuclear Overhauser effect between bound anesthetic protons and all protein protons. To establish STD as a method for testing anesthetic binding to proteins, we conducted measurements on a series of protein/anesthetic solutions studied before by other methods. STD was able to identify that volatile anesthetics bind to bovine serum albumin, oleic acid reduces halothane binding to bovine serum albumin, and halothane binds to apomyoglobin but not lysozyme. Using STD, we found that halothane binding to calmodulin is Ca2+ -dependent, which demonstrates anesthetic specificity for a protein conformation. Thus, STD is a powerful tool for investigating anesthetic-protein interactions because of its abilities to detect weak binding, to screen a single protein for binding of multiple anesthetics simultaneously, and to detect a change in anesthetic binding caused by conformational changes or competition with other ligands.
Streiff
John H
JH
Department of Anesthesiology, Mayo College of Medicine, 200 First Street SW, Rochester, MN 55905, USA.
Juranic
Nenad O
NO
Macura
Slobodan I
SI
Warner
David O
DO
Jones
Keith A
KA
Perkins
William J
WJ
D000777:Anesthetics / Q000737:chemistry / Q000378:metabolism*; D001059:Apoproteins / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D006221:Halothane / Q000737:chemistry / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D009211:Myoglobin / Q000378:metabolism; D011485:Protein Binding; D012710:Serum Albumin, Bovine / Q000378:metabolism*
D000777:Anesthetics; D001059:Apoproteins; D002147:Calmodulin; D009211:Myoglobin; C010857:apomyoglobin; D012710:Serum Albumin, Bovine; D006221:Halothane