Related Experiment Videos
Effect of posttranslational modifications to myelin basic protein on its ability to aggregate acidic lipid vesicles
J M Boggs1, P M Yip, G Rangaraj
1Research Institute, Hospital for Sick Children, Toronto, Ontario, Canada. jmboggs@sickkids.on.ca
Abstract:
When isolated from central nervous system myelin, myelin basic protein (MBP) exhibits charge microheterogeneity due to posttranslational deamidation, phosphorylation, and deimination of arginine to citrulline. These modifications are known to decrease the ability of MBP to aggregate acidic lipid vesicles and thus could regulate the ability of MBP to mediate adhesion between the intracellular surfaces of myelin. The effects of salt (KCl) concentration and the protein to lipid ratio on the ability of charge isomers of MBP to aggregate large unilamellar vesicles (LUVs) were investigated. Increased salt concentration from 10 to 100 mM caused increasing aggregation of LUVs by low concentrations of all charge isomers but did not eliminate the differences in their abilities to aggregate. All isomers were bound equally up to about 100 mM K+ but were dissociated at higher K+ concentrations. The degree of dissociation increased with increasing net negative charge of the isomer. At high concentrations all charge isomers except the form in which six arginine residues are converted to citrulline (C8) aggregated LUVs of phosphatidylcholine/phosphatidylserine (PC/PS) 8:2 (mol/mol) similarly and salt increased the aggregation to the same degree for all. There was less difference in the ability of the charge isomers, including C8, to aggregate LUVs with a lipid composition resembling that of the cytoplasmic leaflet of myelin (Cyt-LUVs) than for PC/PS LUVs. Furthermore, high salt concentrations (400 mM) did not dissociate any of the charge isomers from the Cyt-LUVs. These results suggest that the reason for inhibition of aggregating ability by charge modification is not increased charge repulsion of the protein but rather its reduced multivalency of net positive charge. They indicate further that the lipid composition of the cytoplasmic leaflet is ideally suited to permit MBP-mediated adhesion and that charge modifications of MBP would probably not affect adhesion of the intracellular surfaces of compact myelin where MBP concentration is high. However, charge modifications might affect adhesion in cytoplasm-containing regions of myelin such as the paranodal loops, where MBP concentration is low and where K+ concentration may vary in the range of 60-75 mM.
Insights
Myelin basic protein (MBP) charge modifications affect its ability to aggregate lipid vesicles. These changes may influence myelin adhesion in specific regions, particularly where MBP concentration is low.
Area of Science:
- Neuroscience
- Biochemistry
- Cell Biology
Background:
- Myelin basic protein (MBP) is crucial for myelin structure.
- Posttranslational modifications like deamidation and phosphorylation cause charge heterogeneity in MBP.
- These modifications can alter MBP's lipid-binding and aggregation capabilities.
Purpose of the Study:
- To investigate how salt concentration and protein-to-lipid ratio affect the aggregation of lipid vesicles by different MBP charge isomers.
- To understand the impact of MBP charge modifications on its ability to mediate adhesion between intracellular myelin surfaces.
Main Methods:
- Utilized large unilamellar vesicles (LUVs) with varying lipid compositions (PC/PS and myelin-like cytoplasmic leaflet).
- Assessed the aggregation of LUVs by different MBP charge isomers across a range of salt (KCl) concentrations.
- Examined the binding and dissociation of MBP isomers from LUVs at different ionic strengths.
Main Results:
- Increased salt concentration (10-100 mM KCl) enhanced LUV aggregation by MBP isomers, but differences in aggregation ability persisted.
- MBP isomers showed equal binding up to 100 mM K+, with dissociation at higher concentrations, increasing with negative charge.
- At high salt, most MBP isomers aggregated PC/PS LUVs similarly, but differences were less pronounced with myelin-like cytoplasmic leaflet LUVs (Cyt-LUVs).
- High salt (400 mM) did not dissociate any MBP isomers from Cyt-LUVs.
Conclusions:
- Inhibition of MBP aggregation by charge modification is due to reduced positive charge multivalency, not charge repulsion.
- The cytoplasmic leaflet lipid composition favors MBP-mediated adhesion.
- MBP charge modifications likely do not impact adhesion in compact myelin but may affect adhesion in cytoplasm-rich regions like paranodal loops.