Characteristics of the interaction of melittin with sarcoplasmic reticulum membranes.
Addition of an amphipathic bee venom peptide, melittin, to sarcoplasmic reticulum (SR) vesicles isolated from rabbit skeletal muscles resulted in a fast (<1 min) blue shift in the fluorescence maximum of the melittin--SR membrane complex. Over the following 45 min the position of the fluorescence maximum did not change, but the fluorescence intensity of the melittin--SR membrane complex decreased by approximately 35% with rate constant 0.14 min-1. Melittin rapidly quenched the isotropic signal in the EPR spectrum of spin-labeled stearic acid added to SR membranes. Further changes in the spectral parameters of the spin probe bound to SR membranes in the presence of melittin indicated an increase of the viscosity of the probe microenvironment (empiric parameter T/eta was decreased by approximately 35% with rate constant 0.11 min-1). The surface potential of SR membranes measured using a pH-sensitive dye, neutral red, decreased after melittin addition from -60 to -30 mV. It was demonstrated with the use of a cross-linking agent, cupric o-phenanthroline, that melittin induced slow aggregation of Ca-ATPase protein in SR membranes; the content of enzyme in the monomeric form decreased with rate constant 0.14 min-1. It is concluded that melittin binds rapidly to SR membranes, inducing slow changes in Ca-ATPase conformation and oligomeric state as well as structural transitions in the lipid bilayer of SR membranes.
Shorina
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EA
Department of Biochemistry, School of Biology, Lomonosov Moscow State University, Moscow, 119899, Russia. amr@atpase.bio.msu.su
Mast
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NV
Storey
K B
KB
Lopina
O D
OD
Rubtsov
A M
AM
D000818:Animals; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007425:Intracellular Membranes / Q000378:metabolism*; D007700:Kinetics; D008555:Melitten / Q000494:pharmacology*; D018482:Muscle, Skeletal / Q000187:drug effects; D051381:Rats; D012519:Sarcoplasmic Reticulum / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013997:Time Factors
D008555:Melitten; D000252:Calcium-Transporting ATPases