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Identification of functional differences between recombinant human α and β cardiac myosin motors.

Deacon JC, Bloemink MJ, Rezavandi H, Geeves MA, Leinwand LA - Cell. Mol. Life Sci. (2012)

Bottom Line: For these parameters, α-subfragment 1 (S1) is far more similar to adult fast skeletal muscle myosin isoforms than to the slow β isoform despite 91% sequence identity between the motor domains of α- and β-myosin.Among the features that differentiate α- from β-S1: the ATP hydrolysis step of α-S1 is ~ten-fold faster than β-S1, α-S1 exhibits ~five-fold weaker actin affinity than β-S1, and actin·α-S1 exhibits rapid ADP release, which is >ten-fold faster than ADP release for β-S1.Overall, the cycle times are ten-fold faster for α-S1 but the portion of time each myosin spends tightly bound to actin (the duty ratio) is similar.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular, Cellular and Developmental Biology and Biofrontiers Institute, University of Colorado, MCDB, Boulder, CO 80309, USA.

ABSTRACT
The myosin isoform composition of the heart is dynamic in health and disease and has been shown to affect contractile velocity and force generation. While different mammalian species express different proportions of α and β myosin heavy chain, healthy human heart ventricles express these isoforms in a ratio of about 1:9 (α:β) while failing human ventricles express no detectable α-myosin. We report here fast-kinetic analysis of recombinant human α and β myosin heavy chain motor domains. This represents the first such analysis of any human muscle myosin motor and the first of α-myosin from any species. Our findings reveal substantial isoform differences in individual kinetic parameters, overall contractile character, and predicted cycle times. For these parameters, α-subfragment 1 (S1) is far more similar to adult fast skeletal muscle myosin isoforms than to the slow β isoform despite 91% sequence identity between the motor domains of α- and β-myosin. Among the features that differentiate α- from β-S1: the ATP hydrolysis step of α-S1 is ~ten-fold faster than β-S1, α-S1 exhibits ~five-fold weaker actin affinity than β-S1, and actin·α-S1 exhibits rapid ADP release, which is >ten-fold faster than ADP release for β-S1. Overall, the cycle times are ten-fold faster for α-S1 but the portion of time each myosin spends tightly bound to actin (the duty ratio) is similar. Sequence analysis points to regions that might underlie the basis for this finding.

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Related in: MedlinePlus

Comparison of the rate and equilibrium constants for myosin S1 relative to those of Human β-S1. The values listed in Table 1 which discriminate between the α and β isoforms were divided by the equivalent value for human β-S1 and plotted on a log scale. Values contained between the two horizontal lines are within a factor of 2 of the values for human β-S1. This region contains almost all of the human and bovine β-S1 and bovine β-S1 isoform data and excludes all of the α-S1 and rabbit skeletal S1 data. All values plotted for the human and mouse α isoforms lie outside this range and are at least three-fold larger than the value for β-S1. Values for Rabbit skeletal S1 are shown for comparison and are similar to the α isoform values in each case. Other values in Table 1 are within a factor of 2 of the human β-S1 values for all isoforms
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Fig7: Comparison of the rate and equilibrium constants for myosin S1 relative to those of Human β-S1. The values listed in Table 1 which discriminate between the α and β isoforms were divided by the equivalent value for human β-S1 and plotted on a log scale. Values contained between the two horizontal lines are within a factor of 2 of the values for human β-S1. This region contains almost all of the human and bovine β-S1 and bovine β-S1 isoform data and excludes all of the α-S1 and rabbit skeletal S1 data. All values plotted for the human and mouse α isoforms lie outside this range and are at least three-fold larger than the value for β-S1. Values for Rabbit skeletal S1 are shown for comparison and are similar to the α isoform values in each case. Other values in Table 1 are within a factor of 2 of the human β-S1 values for all isoforms

Mentions: We repeated all of the measurements described above for the “humanized” β-sS1 construct. In almost all cases, the data show little difference between the two β-constructs (see example data in Supplementary Fig. S2A–F summarized in Table 1 and Fig. 7). The constants, K1k+2 and k+2 for S1 alone do not differ between the two constructs. All other constants differ by no more than two-fold with the exception of k+αD which is seven-fold faster for β-S1 than for β-sS1. Thus, as seen for other myosin constructs, in the absence of load the light chains have little influence on the biochemical kinetic properties of S1.Fig. 7


Identification of functional differences between recombinant human α and β cardiac myosin motors.

Deacon JC, Bloemink MJ, Rezavandi H, Geeves MA, Leinwand LA - Cell. Mol. Life Sci. (2012)

Comparison of the rate and equilibrium constants for myosin S1 relative to those of Human β-S1. The values listed in Table 1 which discriminate between the α and β isoforms were divided by the equivalent value for human β-S1 and plotted on a log scale. Values contained between the two horizontal lines are within a factor of 2 of the values for human β-S1. This region contains almost all of the human and bovine β-S1 and bovine β-S1 isoform data and excludes all of the α-S1 and rabbit skeletal S1 data. All values plotted for the human and mouse α isoforms lie outside this range and are at least three-fold larger than the value for β-S1. Values for Rabbit skeletal S1 are shown for comparison and are similar to the α isoform values in each case. Other values in Table 1 are within a factor of 2 of the human β-S1 values for all isoforms
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3375423&req=5

Fig7: Comparison of the rate and equilibrium constants for myosin S1 relative to those of Human β-S1. The values listed in Table 1 which discriminate between the α and β isoforms were divided by the equivalent value for human β-S1 and plotted on a log scale. Values contained between the two horizontal lines are within a factor of 2 of the values for human β-S1. This region contains almost all of the human and bovine β-S1 and bovine β-S1 isoform data and excludes all of the α-S1 and rabbit skeletal S1 data. All values plotted for the human and mouse α isoforms lie outside this range and are at least three-fold larger than the value for β-S1. Values for Rabbit skeletal S1 are shown for comparison and are similar to the α isoform values in each case. Other values in Table 1 are within a factor of 2 of the human β-S1 values for all isoforms
Mentions: We repeated all of the measurements described above for the “humanized” β-sS1 construct. In almost all cases, the data show little difference between the two β-constructs (see example data in Supplementary Fig. S2A–F summarized in Table 1 and Fig. 7). The constants, K1k+2 and k+2 for S1 alone do not differ between the two constructs. All other constants differ by no more than two-fold with the exception of k+αD which is seven-fold faster for β-S1 than for β-sS1. Thus, as seen for other myosin constructs, in the absence of load the light chains have little influence on the biochemical kinetic properties of S1.Fig. 7

Bottom Line: For these parameters, α-subfragment 1 (S1) is far more similar to adult fast skeletal muscle myosin isoforms than to the slow β isoform despite 91% sequence identity between the motor domains of α- and β-myosin.Among the features that differentiate α- from β-S1: the ATP hydrolysis step of α-S1 is ~ten-fold faster than β-S1, α-S1 exhibits ~five-fold weaker actin affinity than β-S1, and actin·α-S1 exhibits rapid ADP release, which is >ten-fold faster than ADP release for β-S1.Overall, the cycle times are ten-fold faster for α-S1 but the portion of time each myosin spends tightly bound to actin (the duty ratio) is similar.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular, Cellular and Developmental Biology and Biofrontiers Institute, University of Colorado, MCDB, Boulder, CO 80309, USA.

ABSTRACT
The myosin isoform composition of the heart is dynamic in health and disease and has been shown to affect contractile velocity and force generation. While different mammalian species express different proportions of α and β myosin heavy chain, healthy human heart ventricles express these isoforms in a ratio of about 1:9 (α:β) while failing human ventricles express no detectable α-myosin. We report here fast-kinetic analysis of recombinant human α and β myosin heavy chain motor domains. This represents the first such analysis of any human muscle myosin motor and the first of α-myosin from any species. Our findings reveal substantial isoform differences in individual kinetic parameters, overall contractile character, and predicted cycle times. For these parameters, α-subfragment 1 (S1) is far more similar to adult fast skeletal muscle myosin isoforms than to the slow β isoform despite 91% sequence identity between the motor domains of α- and β-myosin. Among the features that differentiate α- from β-S1: the ATP hydrolysis step of α-S1 is ~ten-fold faster than β-S1, α-S1 exhibits ~five-fold weaker actin affinity than β-S1, and actin·α-S1 exhibits rapid ADP release, which is >ten-fold faster than ADP release for β-S1. Overall, the cycle times are ten-fold faster for α-S1 but the portion of time each myosin spends tightly bound to actin (the duty ratio) is similar. Sequence analysis points to regions that might underlie the basis for this finding.

Show MeSH
Related in: MedlinePlus