Non-equivalent cooperation between the two nucleotide-binding folds of P-glycoprotein.

Journal article

To identify the roles of the two nucleotide-binding folds (NBFs) in the function of human P-glycoprotein, a multidrug transporter, we mutated the key lysine residues to methionines and the cysteine residues to alanines in the Walker A (WA) motifs (the core consensus sequence) in the NBFs. We examined the effects of these mutations on N-ethylmaleimide (NEM) and ATP binding, as well as on the vanadate-induced nucleotide trapping with 8-azido-[alpha-32P]ATP. Mutation of the WA lysine or NEM binding cysteine in either of the NBFs blocked vanadate-induced nucleotide trapping of P-glycoprotein. These results suggest that if one NBF is non-functional, there is no ATP hydrolysis even if the other functional NBF contains a bound nucleotide, further indicating the strong cooperation between the two NBFs of P-glycoprotein. However, we found that the effect of NEM modification at one NBF on ATP binding at the other NBF was not equivalent, suggesting a non-equivalency of the role of the two NBFs in P-glycoprotein function.

Authors: Takada, Y; Yamada, K; Taguchi, Y; Kino, K; Matsuo, M

• Publication status: Published
• Journal: Biochimica et biophysica acta
• Volume: 1373
• Issue: 1
• Pages: 131-136
• Publication date: 1998-08-01
• DOI: 10.1016/s0005-2736(98)00099-6
• ISSN: 0006-3002
• Language: English
• Keywords: Vanadates; Cysteine; P-Glycoprotein; Humans; Protein Binding; Adenosine Triphosphate; Protein Folding; Ethylmaleimide