Energy-dependent binding of dansylgalactoside to the lac carrier protein: direct binding measurements
| Autor: | H R Kaback, Rudolf Weil, Shimon Schuldiner |
|---|---|
| Rok vydání: | 1976 |
| Předmět: |
Carbonyl Cyanide m-Chlorophenyl Hydrazone
Biological Transport Active Carbonyl cyanide m-chlorophenyl hydrazone Membrane Potentials Cell membrane chemistry.chemical_compound Escherichia coli medicine Glycosides Binding site Dansyl Compounds Membrane potential Binding Sites Multidisciplinary Cell-Free System Chemistry Vesicle Cell Membrane Galactosides Fluorescence Binding constant medicine.anatomical_structure Biochemistry Membrane protein Lactates Biophysics Methylphenazonium Methosulfate Carrier Proteins 4-Chloromercuribenzenesulfonate Research Article |
| Zdroj: | Proceedings of the National Academy of Sciences. 73:109-112 |
| ISSN: | 1091-6490 0027-8424 |
| DOI: | 10.1073/pnas.73.1.109 |
| Popis: | High specific activity 6'-N-[3H]dansyl)aminohexyl 1-thio-beta-D-galactopyranoside (Dns6-Gal) has been synthesized, and its binding to Escherichia coli membrane vesicles measured directly by flow dialysis. With ML 308-225 vesicles containing the lac carrier protein, specific binding is not detected in the absence of D-lactate or reduced phenazine methosulfate. In the presence of these electron donors, binding is observed, and the binding constant and number of binding sites are approximately 4 muM and 1.5 nmol/mg of membrane protein, respectively. These values are in excellent agreement with those obtained by fluorescence titration. p-Chloromercuribenzenesulfonate, which directly inactivates the lac carrier protein, and carbonylcyanide m-chlorophenylhydrazone, which collapses the membrane potential, cause release of bound Dns6-Gal. Moreover, significant binding is not observed with membrane vesicles that are devoid of the lac carrier protein. The results provide qualitative and quantitative confirmation of previous studies which indicate that changes in dansylgalactoside fluorescence observed on "energization" of membrane vesicles reflect binding of the probe to the lac carrier protein. |
| Databáze: | OpenAIRE |
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