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"Reversed" alamethicin conductance in lipid bilayers
1Chemistry Department, Georgetown University, Washington DC 20057.
Biophysical Journal
|April 1, 1991
Summary
Alamethicin forms asymmetric ion channels in lipid bilayers. Below room temperature, these channels can reverse polarity and remain stable, allowing for detailed study of alamethicin aggregates.
Area of Science:
- Biophysics
- Membrane Biophysics
- Ion Channel Research
Background:
- Alamethicin is a peptide antibiotic known to form pores in lipid bilayers.
- Ion channel activity is typically voltage-dependent and sensitive to membrane polarity.
- Studying single ion channels provides insights into their structure and function.
Purpose of the Study:
- To investigate the behavior of alamethicin channels in lipid bilayers under reversed polarity conditions.
- To characterize the stability and conductance states of alamethicin channels after polarity reversal.
- To explore the utility of reversed alamethicin channels for studying isolated peptide aggregates.
Main Methods:
- Formation of a lipid bilayer from diphytanoyl phosphatidylcholine and cholesterol using patch clamp techniques.
- Incorporation of alamethicin (2 µg/ml) into one side of the lipid bilayer in a buffered salt solution (0.5 M KCl, 5 mM Hepes, pH 7.0).
- Measurement of current-voltage (I-V) curves and observation of single channel behavior at temperatures below room temperature.
Main Results:
- Alamethicin exhibited an asymmetric current-voltage curve, conducting ions primarily when the peptide-added side was positive.
- Below room temperature, single alamethicin channels sometimes survived polarity reversal, forming stable 'reversed' channels.
- These reversed channels were distinct from transient closing states and could be monitored for minutes, displaying over 20 discrete conductance states.
- No further incorporation of ion-conducting alamethicin aggregates occurred in membranes with reversed channels.
Conclusions:
- Reversed alamethicin channels provide a stable system for studying individual ion-conducting alamethicin aggregates.
- The observed conductance fluctuations in reversed channels offer a unique opportunity to analyze the dynamic behavior of alamethicin pores.
- This phenomenon facilitates the investigation of alamethicin's aggregation and ion transport mechanisms in a controlled manner.