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. 2018 Aug 13;7:e38064. doi: 10.7554/eLife.38064

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© 2018, Kronert et al

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Figure 5. — (A) The frequency at which small amplitude sinusoidal power was maximal, f_max, was measured for a range of ATP concentrations for control (PwMhc2 homozygotes), heterozygous (R146N-15/+) and homozygous (R146N-15/R146N-15) fibers. The curves were fit with the Michaelis-Menten equation and V_max and K_m values determined (see inset). Student’s t-test with p<0.05 significantly different from control (*p<0.05, **p<0.01, ***p<0.001) or heterozygote (^xp<0.05, ^xxp<0.01, ^xxxp<0.001). (B) f_max relative to inorganic phosphate concentration. f_max decreased with increasing [Pi], suggesting phosphate is competing with ATP for the rigor state. * indicates statistically significant difference from 0 mM Pi (Student’s t-test with p<0.05 significantly different (*p<0.05, **p<0.01). Linear regression analysis showed mutants had a steeper decline of f_max and larger negative slopes than the control. Seven control, 12 heterozygous, and seven homozygous fibers were tested. (C) Change in active stiffness with increasing ATP concentration. In-phase stiffness (elastic modulus) was measured at pCa 5.0 at 500 Hz ML oscillation using small amplitude sinusoidal analysis. All R146N stiffness values are statistically greater than the control’s by a Student’s t-test at p<0.05. (D) Active isometric tension generation as a function of ATP concentration at pCa 5.0. For the ATP study, N = 12 control, 12 heterozygous and 14 homozygous fibers. Black and purple horizontal lines indicate [ATP] where heterozygous and homozygous tension values are statistically greater than control’s (t-test, p<0.05). Full genotypes are shown in parentheses: PwMhc2 homozygote control (P{PwMhc2}/P{PwMhc2}; Mhc¹⁰/Mhc¹⁰); R146N-15/+ heterozygote (Mhc¹⁰/+; P{R146N-15}); R146N-15 homozygote (Mhc¹⁰/Mhc¹⁰; P{R146N-15}/P{R146N-15}).

Figure 5—figure supplement 1. — (A) The frequency at which small amplitude sinusoidal power was maximal, f_max, was measured for a range of ATP concentrations for control (PwMhc2 homozygotes), heterozygous (R146N-15/+) and homozygous (R146N-15/R146N-15) fibers. The curves were fit with the Michaelis-Menten equation and V_max and K_m values determined (see inset). Student’s t-test with p<0.05 significantly different from control (*p<0.05, **p<0.01, ***p<0.001) or heterozygote (^xp<0.05, ^xxp<0.01, ^xxxp<0.001). (B) f_max relative to inorganic phosphate concentration. f_max decreased with increasing [Pi], suggesting phosphate is competing with ATP for the rigor state. * indicates statistically significant difference from 0 mM Pi (Student’s t-test with p<0.05 significantly different (*p<0.05, **p<0.01). Linear regression analysis showed mutants had a steeper decline of f_max and larger negative slopes than the control. Seven control, 12 heterozygous, and seven homozygous fibers were tested. (C) Change in active stiffness with increasing ATP concentration. In-phase stiffness (elastic modulus) was measured at pCa 5.0 at 500 Hz ML oscillation using small amplitude sinusoidal analysis. All R146N stiffness values are statistically greater than the control’s by a Student’s t-test at p<0.05. (D) Active isometric tension generation as a function of ATP concentration at pCa 5.0. For the ATP study, N = 12 control, 12 heterozygous and 14 homozygous fibers. Black and purple horizontal lines indicate [ATP] where heterozygous and homozygous tension values are statistically greater than control’s (t-test, p<0.05). Full genotypes are shown in parentheses: PwMhc2 homozygote control (P{PwMhc2}/P{PwMhc2}; Mhc¹⁰/Mhc¹⁰); R146N-15/+ heterozygote (Mhc¹⁰/+; P{R146N-15}); R146N-15 homozygote (Mhc¹⁰/Mhc¹⁰; P{R146N-15}/P{R146N-15}).