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Literature summary for 7.6.2.2 extracted from
- Thiol-reactive drug substrates of human P-glycoprotein label the same sites to activate ATPase activity in membranes or dodecyl maltoside detergent micelles (2017), Biochem. Biophys. Res. Commun., 488, 573-577 .
Activating Compound
| Activating Compound | Comment | Organism | Structure |
|---|---|---|---|
| additional information | drug-stimulated ATPase activity | Homo sapiens | |
| tariquidar | the substrate inhibits P-gp ATPase activity in membranes, but stimulates ATPase activity when purified P-gp is assayed in the presence of n-dodecyl-beta-D-maltoside/sheep brain lipids. Tariquidar activates the enzyme in presence of lipids, overview | Homo sapiens |  |
| verapamil | verapamil activates the enzyme in presence of lipids, overview. When wild-type human P-gp is expressed in Sf9 insect cells and the membranes are assayed for activity in the absence or presence of saturating concentrations of verapamil, verapamil activates ATPase activity by 4.7fold | Homo sapiens | |
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| recombinant expression of His-tagged mutant enzymes in HEK 293 cells, recombinant expression of wild-type enzyme in Spodoptera frugiperda Sf9 cells | Homo sapiens |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| A80C/R741C | site-directed mutagenesis, cysteines A80C/R741C are close enough to spontaneously form a disulfide bond in the mature protein at the cell surface and trap Pgp in an inactive conformation. Tariquidar is the only P-gp drug substrate or modulator that blocks A80C/R741C crosslinking | Homo sapiens |
| F343C | site-directed mutagenesis, mutant F343C shows highly activated ATPase activity (about 7fold higher than untreated) after treatment with MTS-rhodamine when labeling is performed in the presence or absence of n-dodecyl-beta-D-maltoside | Homo sapiens |
| I306C | site-directed mutagenesis, mutant I306C (TM5) shows highly activated ATPase activity, about 10fold greater than the untreated mutant after treatment with MTS-verapamil when labeling is performed in the presence or absence of n-dodecyl-beta-D-maltoside | Homo sapiens |
| L1260A | site-directed mutagenesis, rescue of the L1260A processing mutant by detergents | Homo sapiens |
| additional information | introduction of L1260A processing mutation in wild-type P-gp containing a C-terminal A52 epitope tag. Cysteine mutations A80C and R741C are introduced into the A52-tagged human Cys-less Pgp cDNA. In transmembrane segments 4-6 or 10-12, single cysteine mutations are introduced at positions K213-A233 (TM4), T294-Y316 (TM5), Q330-P350 (TM6), W855-E875 (TM10), H936-F957 (TM11), or V974-F994 (TM12) | Homo sapiens |
Inhibitors
| Inhibitors | Comment | Organism | Structure |
|---|---|---|---|
| EDTA | - |
Homo sapiens | |
| SDS | - |
Homo sapiens | |
| tariquidar | the substrate inhibits P-gp ATPase activity in membranes, but stimulates ATPase activity when purified P-gp is assayed in the in the presence of n-dodecyl-beta-D-maltoside/sheep brain lipids. When wild-type human P-gp is expressed in Sf9 insect cells and the membranes are assayed for activity in the absence or presence of saturating concentrations of tariquidar, tariquidar reduces basal ATPase activity by about 60% | Homo sapiens | |
Localization
| Localization | Comment | Organism | GeneOntology No. | Textmining |
|---|---|---|---|---|
| membrane | a transmembrane enzyme with 12 transmembrane segments | Homo sapiens | 16020 | - |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| ATP + H2O + cyclosporine A[side 1] | Homo sapiens | - |
ADP + phosphate + cyclosporine A [side 2] | - |
? | |
| ATP + H2O + MTS-rhodamine [side 1] | Homo sapiens | the binding site in transmembrane segment 5 is F343C for rhodamine | ADP + phosphate + MTS-rhodamine [side 2] | - |
? | |
| ATP + H2O + tariquidar [side 1] | Homo sapiens | - |
ADP + phosphate + tariquidar [side 2] | - |
? | |
| ATP + H2O + verapamil [side 1] | Homo sapiens | the binding site in transmembrane segment 5 is I306C for verapamil | ADP + phosphate + verapamil [side 2] | - |
? | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Homo sapiens | P08183 | - |
- |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| ATP + H2O + cyclosporine A[side 1] | - |
Homo sapiens | ADP + phosphate + cyclosporine A [side 2] | - |
? | |
| ATP + H2O + MTS-rhodamine [side 1] | the binding site in transmembrane segment 5 is F343C for rhodamine | Homo sapiens | ADP + phosphate + MTS-rhodamine [side 2] | - |
? | |
| ATP + H2O + tariquidar [side 1] | - |
Homo sapiens | ADP + phosphate + tariquidar [side 2] | - |
? | |
| ATP + H2O + verapamil [side 1] | the binding site in transmembrane segment 5 is I306C for verapamil | Homo sapiens | ADP + phosphate + verapamil [side 2] | - |
? | |
| additional information | n-dodecyl-beta-D-maltoside is no substrate for the enzyme, and the presence of n-dodecyl-beta-D-maltoside does not alter the locations of drug-binding sites, e.g. for tariquidar. The enzyme shows drug-stimulated ATPase activity | Homo sapiens | ? | - |
- |
|
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| ABCB1 | - |
Homo sapiens |
| P-glycoprotein | - |
Homo sapiens |
| P-gp | - |
Homo sapiens |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
- |
20 | assay at | Homo sapiens |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 7.4 | - |
assay at | Homo sapiens |
General Information
| General Information | Comment | Organism |
|---|---|---|
| malfunction | rescue of the L1260A processing mutant by detergents, such as Triton X-100 or NP-35, but not of n-dodecyl-beta-D-maltoside. Only mutant I306C (TM5) shows highly activated ATPase activity (about 10fold greater than the untreated mutant) after treatment with MTS-verapamil while only mutant F343C shows highly activated ATPase activity (about 7fold higher than untreated) after treatment with MTS-rhodamine when labeling is performed in the presence or absence of n-dodecyl-beta-D-maltoside. All other mutants show less than a twofold activation after treatment in the presence or absence of detergent | Homo sapiens |
| physiological function | human P-glycoprotein (P-gp, ABCB1) is a drug pump that catalyzes the ATP-dependent export of a wide variety of lipophilic compounds such as hydrophobic drugs, steroids, peptides, and detergents. Its physiological role is to protect the organism from cytotoxic compounds and plays an important role in the development of multidrug resistance | Homo sapiens |
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