An actin-dependent conformational change in myosin.
Conformational changes within myosin lead to its movement relative to an actin filament. Several crystal structures exist for myosin bound to various nucleotides, but none with bound actin. Therefore, the effect of actin on the structure of myosin is poorly understood. Here we show that the swing of smooth muscle myosin lever arm requires both ADP and actin. This is the first direct observation that a conformation of myosin is dependent on actin. Conformational changes within myosin were monitored using fluorescence resonance energy transfer techniques. A cysteine-reactive probe is site-specifically labeled on a 'cysteine-light' myosin variant, in which the native reactive cysteines were removed and a cysteine engineered at a desired position. Using this construct, we show that the actin-dependent ADP swing causes an 18 A change in distance between a probe on the 25/50 kDa loop on the catalytic domain and a probe on the regulatory light chain, corresponding to a 23 degrees swing of the light-chain domain.
Xiao
Ming
M
Department of Physics, University of Illinois, 1110 West Green Street, Urbana, Illinois 61801, USA.
Reifenberger
Jeff G
JG
Wells
Amber L
AL
Baldacchino
Corry
C
Chen
Li-Qiong
LQ
Ge
Pinghua
P
Sweeney
H Lee
HL
Selvin
Paul R
PR
D000199:Actins / Q000737:chemistry* / Q000378:metabolism*; D019943:Amino Acid Substitution; D000818:Animals; D002645:Chickens; D003545:Cysteine; D031541:Fluorescence Resonance Energy Transfer; D005895:Gizzard, Avian / Q000378:metabolism; D007700:Kinetics; D009130:Muscle, Smooth / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D018994:Myosin Light Chains / Q000737:chemistry / Q000378:metabolism; D009218:Myosins / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation
D000199:Actins; D018994:Myosin Light Chains; D009218:Myosins; D003545:Cysteine