Functional consequences of mutations in the smooth muscle myosin heavy chain at sites implicated in familial hypertrophic cardiomyopathy.
Familial hypertrophic cardiomyopathy (FHC) is frequently associated with mutations in the beta-cardiac myosin heavy chain. Many of the implicated residues are located in highly conserved regions of the myosin II class, suggesting that these mutations may impair the basic functions of the molecular motor. To test this hypothesis, we have prepared recombinant smooth muscle heavy meromyosin with mutations at sites homologous to those associated with FHC by using a baculovirus/insect cell expression system. Several of the heavy meromyosin mutants, in particular R403Q, showed an increase in actin filament velocity in a motility assay and an enhanced actin-activated ATPase activity. Single molecule mechanics, using a laser trap, gave unitary displacements and forces for the mutants that were similar to wild type, but the attachment times to actin following a unitary displacement were markedly reduced. These results suggest that the increases in activity are due to a change in kinetics and not due to a change in the intrinsic mechanical properties of the motor. In contrast to earlier reports, we find that mutations in residues implicated in FHC affect motor function by enhancing myosin activity rather than by a loss of function.
Yamashita
H
H
Department of Cardiovascular Medicine, Graduate School of Medicine, University of Tokyo, Tokyo 113-8655, Japan.
Tyska
M J
MJ
Warshaw
D M
DM
Lowey
S
S
Trybus
K M
KM
D000199:Actins / Q000378:metabolism; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D002312:Cardiomyopathy, Hypertrophic / Q000235:genetics* / Q000378:metabolism*; D002645:Chickens; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D005895:Gizzard, Avian; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D009130:Muscle, Smooth / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009206:Myocardium / Q000378:metabolism; D018995:Myosin Heavy Chains / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D009218:Myosins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017354:Point Mutation; D011487:Protein Conformation; D017433:Protein Structure, Secondary
D000199:Actins; D018995:Myosin Heavy Chains; D009218:Myosins