Inhibition of P-glycoprotein ATPase activity by beryllium fluoride.
ATPase activity of P-glycoprotein (multidrug-resistance protein) was found to be potently inhibited by beryllium fluoride (BeFx) in combination with MgATP, MgADP, or corresponding Mg-8-azido-nucleotides. Inhibition was due to trapping of nucleoside diphosphate at catalytic sites. Full inhibition was achieved on trapping of 1 mol of nucleotide per mol of Pgp. Reactivation was slow (t(1/2) = 32 min at 37 degrees C), and release of trapped nucleotide correlated with recovery of ATPase. Trapping of 8-azido-ADP followed by UV irradiation yielded permanent inactivation and specific labeling of Pgp in plasma membranes. Both N- and C-terminal nucleotide binding sites were labeled. These findings give strong confirmation of the concepts that in intact Pgp both nucleotide sites are active in MgATP hydrolysis, and that they interact strongly. The characteristics of inhibition by BeFx were similar in general to those seen with vanadate. However, PPi gave strong protection against BeFx inhibition, and in this respect, inhibition by BeFx was clearly different from vanadate inhibition.
Sankaran
B
B
Department of Biochemistry and Biophysics, University of Rochester Medical Center, New York 14642, USA.
Bhagat
S
S
Senior
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AE
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors*; D000244:Adenosine Diphosphate / Q000031:analogs & derivatives / Q000032:analysis / Q000378:metabolism / Q000494:pharmacology; D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000345:Affinity Labels; D000818:Animals; D001386:Azides / Q000378:metabolism / Q000494:pharmacology; D001608:Beryllium / Q000494:pharmacology*; D001667:Binding, Competitive; D016466:CHO Cells; D002413:Cations, Divalent; D006224:Cricetinae; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004793:Enzyme Reactivators; D005459:Fluorides / Q000494:pharmacology*; D007700:Kinetics; D010710:Phosphates / Q000378:metabolism / Q000494:pharmacology; D012969:Sodium Fluoride / Q000494:pharmacology; D014466:Ultraviolet Rays; D014700:Verapamil / Q000494:pharmacology
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000345:Affinity Labels; D001386:Azides; D002413:Cations, Divalent; D004791:Enzyme Inhibitors; D004793:Enzyme Reactivators; D010710:Phosphates; C035982:beryllium fluoride; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D012969:Sodium Fluoride; D014700:Verapamil; D000251:Adenosine Triphosphatases; C021622:8-azidoadenosine diphosphate; D001608:Beryllium; D005459:Fluorides