Differences in structural dynamics of muscle and yeast actin accompany differences in functional interactions with myosin.
We have used spectroscopic probes ErIA and IAEDANS attached to Cys374 to compare the structural dynamics of yeast actin filaments with that of muscle actin, to understand the structural basis of the less productive interaction of yeast actin with myosin. Time-resolved phosphorescence anisotropy (TPA) of ErIA and steady-state fluorescence of IAEDANS were measured. TPA indicated more rapid rotational motion and more restricted angular amplitude in yeast actin. The fluorescence spectrum was less intense and more red-shifted in yeast actin, suggesting more exposure of the probe to solvent. These results indicate that the two actins differ substantially in the conformational dynamics of the C-terminal region. Binding of myosin S1 induced significantly different spectroscopic changes in TPA and fluorescence of muscle and yeast actin. As a result, the spectroscopic differences between the two actins were decreased by the addition of S1. These results suggest that yeast actin is less effective at activating myosin because of larger changes required in the structure of actin upon strong myosin binding. These results provide insight into the relationship between actomyosin dynamics and function, and they provide a useful framework for structure-function analysis of mutant yeast actin.
Prochniewicz
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Department of Biochemistry, University of Minnesota Medical School, Minneapolis, Minnesota 55455, USA.
Thomas
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D000199:Actins / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D004735:Energy Transfer; D004789:Enzyme Activation; D004923:Erythrosine / Q000031:analogs & derivatives / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D008274:Magnesium / Q000378:metabolism; D018482:Muscle, Skeletal / Q000378:metabolism*; D009218:Myosins / Q000378:metabolism*; D009282:Naphthalenesulfonates / Q000378:metabolism; D011817:Rabbits; D013329:Structure-Activity Relationship; D015003:Yeasts
D000199:Actins; D005456:Fluorescent Dyes; D009282:Naphthalenesulfonates; C006421:1,5-I-AEDANS; C038909:iodoacetamidoerythrosin; D009218:Myosins; D008274:Magnesium; D004923:Erythrosine; D002118:Calcium