[Study of cation binding to myosin subfragment I using the fluorescent probe Eu3+].
It was shown that binding of the cation Eu3+ to myosin subfragment I (SI) results in fluorescence with a maximum at 595 nm which is increased as EuCl3 concentration rises. The ATPase activity of SI is simultaneously enhanced. An addition of bivalent cations (Ca2+ and Mg2+) causes quenching of fluorescence of the bound Eu3+ by 8-12%, which corresponds to Eu3+-ATPase inhibition by Mg2+. At low (down to 0.1 mM) concentrations of Eu3+ no fluorescence quenching by Mg2+ or Ca2+ takes place; under these conditions Mg2+ activate ATPase of SI in the presence of Eu3+. Eu3+ activate SI ATPase in the presence of low (down to 0.1 mM) concentrations of Ca2+, but exerts an inhibition action at high concentrations of Ca2+. NaCl does not affect the fluorescence intensity of bound Eu3+ but considerably inhibits the ATPase activity of SI in the presence of EuCl3. An existence of a bivalent cation binding site in the vicinity if the SI active center is postulated. Eu3+ whose ionic radius is close to that of Ca2+ interacts with protein surface and occupies this site as well, thus determining the activation of SI ATPase and can be replaced from it by Ca2+ and Mg2+, but not by Na+. Hence bivalent and monovalent cations are bound at different sites. The data obtained provide another proof in favour of a hypothesis suggesting that the regulation of myosin ATPase activity by bivalent and monovalent cations can be mediated by binding of these cations to the protein.
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology; D005063:Europium; D005456:Fluorescent Dyes; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D009132:Muscles / Q000201:enzymology; D015879:Myosin Subfragments; D009218:Myosins / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D011485:Protein Binding; D011817:Rabbits; D013050:Spectrometry, Fluorescence
D005456:Fluorescent Dyes; D015879:Myosin Subfragments; D010446:Peptide Fragments; D005063:Europium; D009218:Myosins; D008274:Magnesium; D002118:Calcium
Izuchenie sviazyvaniia kationov s subfragmentom I miozina pri pomoshchi fluorestsentnogo zonda Eu3+.