Characterization of mutant myosins of Dictyostelium discoideum equivalent to human familial hypertrophic cardiomyopathy mutants. Molecular force level of mutant myosins may have a prognostic implication.
Recent studies have revealed that familial hypertrophic cardiomyopathy (FHC) is caused by missence mutations in myosin heavy chain or other sarcomeric proteins. To investigate the functional impact of FHC mutations in myosin heavy chain, mutants of Dictyostelium discoideum myosin II equivalent to human FHC mutations were generated by site-directed mutagenesis, and their motor function was characterized at the molecular level. These mutants, i.e., R397Q, F506C, G575R, A699R, K703Q, and K703W are respectively equivalent to R403Q, F513C, G584R, G716R, R719Q, and R719W FHC mutants. We measured the force generated by these myosin mutants as well as the sliding velocity and the actin-activated ATPase activity. These measurements showed that the A699R, K703Q, and K703W myosins exhibited unexpectedly weak affinity with actin and the lowest level of force, though their ATPase activity remained rather high. F506C mutant which has been reported to have benign prognosis exhibited the least impairment of the motile and enzymatic activities. The motor functions of R397Q and G575R myosins were classified as intermediate. These results suggest that the force level of mutant myosin molecule may be one of the key factors for pathogenesis which affect the prognosis of human FHC.
Fujita
H
H
Second Department of Internal Medicine, University of Tokyo, Hongo, Japan. hfujita@medbank.or.jp
Sugiura
S
S
Momomura
S
S
Omata
M
M
Sugi
H
H
Sutoh
K
K
D000199:Actins / Q000378:metabolism / Q000502:physiology; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002312:Cardiomyopathy, Hypertrophic / Q000235:genetics*; D004023:Dictyostelium / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D006801:Humans; D008969:Molecular Sequence Data; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D009218:Myosins / Q000235:genetics* / Q000302:isolation & purification / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D011995:Recombination, Genetic; D014170:Transformation, Genetic
D000199:Actins; D011994:Recombinant Proteins; D000251:Adenosine Triphosphatases; D009218:Myosins