Kinetic and spectroscopic evidence for three actomyosin:ADP states in smooth muscle.
Smooth muscle myosin II undergoes an additional movement of the regulatory domain with ADP release that is not seen with fast skeletal muscle myosin II. In this study, we have examined the interactions of smooth muscle myosin subfragment 1 with ADP to see if this additional movement corresponds to an identifiable state change. These studies indicate that for this myosin:ADP, both the catalytic site and the actin-binding site can each assume one of two conformations. Relatively loose coupling between these two binding sites leads to three discrete actin-associated ADP states. Following an initial, weakly bound state, binding of myosin:ADP to actin shifts the equilibrium toward a mixture of two states that each bind actin strongly but differ in the conformation of their catalytic sites. By contrast, fast myosins, including Dictyostelium myosin II, have reciprocal coupling between the actin- and ADP-binding sites, so that either actin or nucleotide, but not both, can be tightly bound. This uncoupling, which generates a second strongly bound actomyosin ADP state in smooth muscle, would prolong the fraction of the ATPase cycle time that this actomyosin spends in a force-generating conformation and may be central to explaining the physiologic differences between this and other myosins.
Rosenfeld
S S
SS
Department of Neurology, University of Alabama at Birmingham, 35294, USA. s_rosenfeld@email.neuro.uab.edu
Xing
J
J
Whitaker
M
M
Cheung
H C
HC
Brown
F
F
Wells
A
A
Milligan
R A
RA
Sweeney
H L
HL
D000199:Actins / Q000378:metabolism; D000205:Actomyosin / Q000737:chemistry* / Q000378:metabolism; D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D020134:Catalytic Domain; D002645:Chickens; D004023:Dictyostelium / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D008854:Microscopy, Electron; D008956:Models, Chemical; D018656:Muscle Fibers, Fast-Twitch / Q000737:chemistry; D018657:Muscle Fibers, Slow-Twitch / Q000737:chemistry; D009130:Muscle, Smooth / Q000737:chemistry*; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011817:Rabbits; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence
D000199:Actins; D011994:Recombinant Proteins; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D000205:Actomyosin