Related Experiment Videos
35S-Atractyloside binding affinity to the inner mitochondrial membrane
P V. Vignais1, P M. Vignais, M G. Colomb
1Laboratoire de Biochimie, Centre d'Etudes Nucléaires et Faculté de Médicine, 38-, Grenoble, France
FEBS Letters
|July 3, 1970
Summary
Aging and ADP decrease high-affinity atractyloside binding to the inner mitochondrial membrane. This suggests changes in adenine nucleotide translocation, impacting mitochondrial function.
Area of Science:
- Mitochondrial Biology
- Biochemistry
- Cellular Respiration
Background:
- The inner mitochondrial membrane (IMM) is crucial for cellular energy production.
- Atractyloside is a known inhibitor that binds to the adenine nucleotide translocator (ANT).
- Understanding ANT binding dynamics is key to deciphering mitochondrial function and dysfunction.
Purpose of the Study:
- To investigate the binding characteristics of atractyloside to the IMM.
- To explore how factors like aging, pH, ADP, and uncouplers affect atractyloside binding.
- To elucidate the role of atractyloside binding in probing IMM conformation related to adenine nucleotide translocation.
Main Methods:
- Isolation of the inner mitochondrial membrane.
- Binding assays using atractyloside (specifically 35S-atractyloside).
- Measurement of binding affinity (Kd) and number of binding sites (Bmax) under various conditions.
Main Results:
- High-affinity atractyloside binding sites (0.1 nmole/mg protein, Kd ~0.014 μM) were identified on the IMM.
- Aging, acidification, and ADP significantly reduced high-affinity binding.
- ADP induced a two-step transition to lower affinity binding (Kd > 0.50 μM) and doubled binding sites.
- Uncouplers like FCCP did not affect binding affinity.
Conclusions:
- IMM conformation, specifically related to adenine nucleotide translocation, is sensitive to aging, pH, and ADP levels.
- ADP binding induces significant conformational changes in the ANT.
- 35S-atractyloside serves as a valuable probe for studying IMM conformation and ANT function.