P-31-CP-MAS NMR studies on TPP+ bound to the ion-coupled multidrug transport protein EmrE

The binding of tetraphenylphosphonium (TPP+) to EmrE, a membrane-bound, 110 residue Escherichia coli multidrug transport protein, has been observed by P-31 cross-polarisation-magic-angle spinning nuclear magnetic resonance sp ectroscopy (CP-MAS NMR). EmrE has been reconstituted into dimyristoyl phosp hatidylcholine bilayers. CP-MAS could selectively distinguish binding of TP P+ to EmrE in the fluid membrane. A population of bound ligand appears shif ted 4 ppm to lower frequency compared to free ligand in solution, which sug gests a rather direct and specific type of interaction of the ligand with t he protein. This is also supported by the observed restricted motion of the bound ligand, The observation of another weakly bound substrate population arises from ligand binding to negatively charged residues in the protein l oop regions. (C) 2000 Federation of European Biochemical Societies. Publish ed by Elsevier Science B,V. All rights reserved.