Interaction of LDS-751 with the drug-binding site of P-glycoprotein: a Trp fluorescence steady-state and lifetime study.
P-glycoprotein (ABCB1) is an ATP-driven efflux pump which binds drugs within a large flexible binding pocket. Intrinsic Trp fluorescence was used to probe the interactions of LDS-751 (2-[4-(4-[dimethylamino]phenyl)-1,3-butadienyl]-3-ethylbenzo-thiazolium perchlorate) with purified P-glycoprotein, using steady-state/lifetime measurements and collisional quenching. The fast decay component of P-glycoprotein intrinsic fluorescence (tau(1)=0.97 ns) was unaffected by LDS-751 binding, while the slow decay component (tau(2)=4.02 ns) was quenched by dynamic and static mechanisms. Both the wavelength-dependence of the decay kinetics, and the time-resolved emission spectra, suggested the existence of excited-state relaxation processes within the protein matrix on the nanosecond time-scale, which were altered by LDS-751 binding. The fast decay component, which is more solvent-exposed, can be attributed to cytosolic/extracellular Trp residues, while the slow decay component likely arises from more buried transmembrane Trp residues. Interaction of a drug with the binding pocket of P-glycoprotein thus affects its molecular structure and fast dynamics.
Lugo
Miguel R
MR
Instituto de Biología Experimental, Universidad Central de Venezuela, Caracas, DC 47114, Venezuela.
Sharom
Frances J
FJ
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D003600:Cytosol / Q000737:chemistry; D004347:Drug Interactions; D005110:Extracellular Space / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry* / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D051379:Mice; D009930:Organic Chemicals / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D013050:Spectrometry, Fluorescence; D013329:Structure-Activity Relationship; D014364:Tryptophan / Q000737:chemistry*
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D005456:Fluorescent Dyes; D008565:Membrane Proteins; D009930:Organic Chemicals; C057718:6-(Dimethylamino)-2-((1E,3E)-4-(4-(dimethylamino)phenyl)-1,3-butadien-1-yl)-1-ethylquinolinium perchlorate; D014364:Tryptophan