Inhibition of SERCA Ca2+ pumps by 2-aminoethoxydiphenyl borate (2-APB). 2-APB reduces both Ca2+ binding and phosphoryl transfer from ATP, by interfering with the pathway leading to the Ca2+-binding sites
| Autor: | Rita E. Godfrey, Laura L. Wootton, Jonathan G. Bilmen, Oliver S. Smart, Francesco Michelangeli |
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| Rok vydání: | 2002 |
| Předmět: |
Boron Compounds
SERCA Protein Conformation ATPase Calcium-Transporting ATPases Biochemistry Fluorescence Sarcoplasmic Reticulum Calcium-Transporting ATPases Adenosine Triphosphate Animals Nucleotide Binding site Enzyme Inhibitors Phosphorylation Lipid bilayer chemistry.chemical_classification biology Endoplasmic reticulum Cell Membrane Tryptophan Transmembrane domain Kinetics chemistry Mutation Biophysics biology.protein Plasma membrane Ca2+ ATPase Calcium sense organs |
| Zdroj: | European journal of biochemistry. 269(15) |
| ISSN: | 0014-2956 |
| Popis: | 2-Aminoethoxydiphenyl Borate (2-APB) has been extensively used recently as a membrane permeable modulator of inositol-1,4,5-trisphosphate-sensitive Ca2+ channels and store-operated Ca2+ entry. Here, we report that 2-APB is also an inhibitor of sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) Ca2+ pumps, and additionally increases ion leakage across the phospholipid bilayer. Therefore, we advise caution in the interpretation of results when used in Ca2+ signalling experiments. The inhibition of 2-APB on the SERCA Ca2+ pumps is isoform-dependent, with SERCA 2B being more sensitive than SERCA 1A (IC50 values for inhibition being 325 and 725 micro m, respectively, measured at pH 7.2). The Ca2+-ATPase is also more potently inhibited at lower pH (IC50 = 70 micro m for SERCA1A at pH 6). 2-APB decreases the affinity for Ca2+ binding to the ATPase by more than 20-fold, and also inhibits phosphoryl transfer from ATP (by 35%), without inhibiting nucleotide binding. Activity studies performed using mutant Ca2+-ATPases show that Tyr837 is critical for the inhibition of activity by 2-APB. Molecular modeling studies of 2-APB binding to the Ca2+ ATPase identified two potential binding sites close to this residue, near or between transmembrane helices M3, M4, M5 and M7. The binding of 2-APB to these sites could influence the movement of the loop between M6 and M7 (L6-7), and reduce access of Ca2+ to their binding sites. |
| Databáze: | OpenAIRE |
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