Salt-triggered intermembrane exchange of phospholipids and hemifusion by myelin basic protein.
Intervesicle phospholipid exchange through molecular contacts induced by the C1 molecular species of myelin basic protein (MBP) are characterized by using methods that amplify the effect of MBP-membrane interaction. The effect of salt concentration (KCl) on the vesicle-vesicle interaction of anionic sonicated covesicles of 30% 1-palmitoyl-2-oleoylglycero-sn-3-phosphocholine and 70% 1,2-dimyristoylglycero-sn-3-phosphomethanol (POPC/DMPM) by MBP is dissected by a combination of protocols into individual steps: aggregation of vesicles, apposition and contact formation, and hemifusion. Scattering and resonance energy transfer measurements reveal that, in the absence of KCl, MBP promotes rapid aggregation of the vesicles without lipid mixing. At >40 mM KCl, the extent of aggregation is larger and time-dependent. Fluorescence dequenching due to dilution of labeled phospholipids indicates that on a somewhat slower time scale, hemifusion of vesicles is triggered by salt, with mixing of the outer monolayer lipids but without flip-flop of phospholipids and without mixing or leakage of the aqueous contents. The exchange and hemifusion are seen with anionic vesicles; the effect of the structure of phospholipid, composition of vesicles, and the protein/lipid ratio is primarily on the kinetics of these and other competing processes. Thus, at 0.022 mol % of MBP and less than 100 mM KCl, it is possible to uncouple three sequential steps: (1) aggregation of vesicles by MBP; (2) apposition of bilayers and selective lipid exchange through vesicle-vesicle contacts established by MBP, i.e., anionic and zwitterionic phospholipids exchange, but cationic probes are excluded; and (3) hemifusion and lipid mixing of contacting monolayers of vesicles.
Cajal
Y
Y
Department of Chemistry and Biochemistry, University of Delaware, Newark 19716, USA.
Boggs
J M
JM
Jain
M K
MK
D000838:Anions; D001665:Binding Sites / Q000187:drug effects; D002412:Cations; D004735:Energy Transfer; D005456:Fluorescent Dyes; D020404:Glycerophospholipids; D006801:Humans; D007223:Infant; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D008561:Membrane Fusion / Q000187:drug effects*; D004676:Myelin Basic Protein / Q000737:chemistry / Q000187:drug effects / Q000494:pharmacology*; D010316:Particle Size; D010712:Phosphatidic Acids; D010713:Phosphatidylcholines; D010743:Phospholipids / Q000737:chemistry* / Q000378:metabolism; D011189:Potassium Chloride / Q000494:pharmacology*; D012235:Rhodamines; D013050:Spectrometry, Fluorescence; D014867:Water / Q000378:metabolism
D000838:Anions; D002412:Cations; D005456:Fluorescent Dyes; D020404:Glycerophospholipids; D008051:Lipid Bilayers; D004676:Myelin Basic Protein; D010712:Phosphatidic Acids; D010713:Phosphatidylcholines; D010743:Phospholipids; D012235:Rhodamines; D014867:Water; C029313:dimyristoylmethylphosphatidic acid; D011189:Potassium Chloride; C044542:octadecyl Rhodamine B chloride; C028694:1-palmitoyl-2-oleoylphosphatidylcholine