Two-state equilibria of myosin subfragment 1 and its complexes with ADP and actin.
We previously reported that the nucleotide complex of myosin subfragment 1, S1.epsilon ADP, exists in two states on the basis of the temperature dependence of the fluorescence decay of bound 1,N6-ethenoadenosine diphosphate (epsilon ADP) [Aguirre, R., Lin. S.-H., Gonsoulin, F., Wang, C.-K., & Cheung, H.C. (1989) Biochemistry 28, 799-809]. We have extended the previous study of the equilibrium between the two states, S1L.ADP in equilibrium S1H.ADP, by using a fluorescently labeled myosin S1 (S1-AF). In S1 alkylated with IAF [5-(iodoacetamido)fluorescein], the decay of the label emission was biexponential both in the presence and absence of ADP and/or actin. In the presence of ADP, the two decay times were 4.30 (alpha 1 = 0.55) and 0.80 ns (alpha 2 = 0.45) at 12.4 degrees C, in a medium containing 60 mM KCl, 30 mM TES (pH 7.5), and 2 mM MgCl2. The steady-state fluorescence intensities of S1-AF, (S1-AF).ADP, acto.(S1-AF), and acto.(S1-AF).ADP were dependent on temperature over the range of 5-30 degrees C. By combining lifetime and steady-state intensity data, we obtained for the two-state transition (S1-AF)L.ADP in equilibrium (S1-AF)H.ADP the following parameters: delta H degrees = 16.1 kcal/mol (67.3 kJ/mol) and delta S degrees = 55.8 cal/(deg.mol) [233.5 J/(deg.mol)], in agreement with previous results obtained with epsilon ADP. The delta H degrees values for the two-state transition of S1-AF, acto.(S1-AF), and acto.(S1-AF).ADP are 13.0, 21.6, and 5.2 kcal/mol, respectively. The corresponding delta S degrees values are 46.9, 79.5, and 17.4 cal/(deg.mol).(ABSTRACT TRUNCATED AT 250 WORDS)
Lin
S H
SH
Graduate Program in Biophysical Sciences, University of Alabama, Birmingham 35294.
Cheung
H C
HC
D000199:Actins / Q000737:chemistry*; D000244:Adenosine Diphosphate / Q000737:chemistry*; D000818:Animals; D005454:Fluorescence Polarization; D009218:Myosins / Q000737:chemistry*; D011817:Rabbits; D013696:Temperature; D013816:Thermodynamics
D000199:Actins; D000244:Adenosine Diphosphate; D009218:Myosins