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Spin-label electron spin resonance study of bacteriophage M13 coat protein incorporation into mixed lipid bilayers
K P Datema1, C J Wolfs, D Marsh
1Department of Molecular Physics, Agricultural University, Wageningen, The Netherlands.
Abstract:
The major coat protein of bacteriophage M13 was incorporated in mixed dimyristoylphosphatidylcholine/dimyristoylphosphatidylglycerol (80/20 w/w) vesicles probed with different spin-labeled phospholipids, labeled on the C-14 atom of the sn-2 chain. The specificity for a series of phospholipids was determined from a motionally restricted component seen in the electron spin resonance (ESR) spectra of vesicles with the coat protein incorporated. At 30 degrees C and pH 8, the fraction of motionally restricted phosphatidic acid spin-label is 0.36, 0.52, and 0.72 for lipid/protein ratios of 18, 14, and 9 mol/mol, respectively. The ESR spectra, analyzed by digital subtraction, resulted in a phospholipid preference following the pattern cardiolipin = phosphatidic acid greater than stearic acid = phosphatidylserine = phosphatidylglycerol greater than phosphatidylcholine = phosphatidylethanolamine. The specificities found are related to the composition of the target Escherichia coli cytoplasmic membrane.
Insights
The M13 bacteriophage coat protein preferentially binds to specific phospholipids, such as cardiolipin and phosphatidic acid, in model cell membranes. This binding preference may reflect interactions within the Escherichia coli cytoplasmic membrane.
Area of Science:
- Biochemistry
- Structural Biology
- Membrane Biophysics
Background:
- Bacteriophage M13 major coat protein plays a role in viral assembly and membrane interaction.
- Understanding protein-lipid interactions is crucial for deciphering viral entry and membrane function.
- Bacterial cytoplasmic membranes have a complex lipid composition influencing protein behavior.
Purpose of the Study:
- To investigate the phospholipid specificity of the M13 major coat protein.
- To determine how lipid composition affects M13 coat protein incorporation into model membranes.
- To relate observed binding preferences to the native Escherichia coli membrane.
Main Methods:
- Incorporation of M13 major coat protein into mixed lipid vesicles (dimyristoylphosphatidylcholine/dimyristoylphosphatidylglycerol).
- Utilizing spin-labeled phospholipids (C-14 atom on sn-2 chain) to probe membrane environment.
- Analyzing electron spin resonance (ESR) spectra to quantify motionally restricted lipid components.
Main Results:
- M13 coat protein incorporation induced a motionally restricted component in ESR spectra, indicating specific lipid interactions.
- Phospholipid preference followed the order: cardiolipin = phosphatidic acid > stearic acid = phosphatidylserine = phosphatidylglycerol > phosphatidylcholine = phosphatidylethanolamine.
- Increased lipid/protein ratios (e.g., 9 mol/mol) resulted in a higher fraction of motionally restricted phosphatidic acid spin-label.
Conclusions:
- The M13 major coat protein exhibits a distinct preference for negatively charged phospholipids like cardiolipin and phosphatidic acid.
- These findings suggest specific interactions between the M13 coat protein and lipid components of the Escherichia coli cytoplasmic membrane.
- The observed phospholipid specificity provides insights into the molecular basis of M13-host interactions.