Micromechanical Thermal Assays of Ca²⁺-Regulated Thin-Filament Function and Modulation by Hypertrophic Cardiomyopathy Mutants of Human Cardiac Troponin

Table 1

Metabolite concentrations in standard and modified motility buffers.

Metabolites	Standard^1	Modified solutions for motility assays^2
	Motility buffer	Control	High Pi	Low ATP
[ATP]	2 mM	2 mM	2 mM	0.2 mM
Sucrose phosphorylase	—	5.3 μg/mL	—	5.3 μg/mL
Sucrose	—	10 mM	10 mM	10 mM
[Pi]	~10 μM	~1 μM	4 mM	~1 μM
Creatine kinase (CK)	—	0.1 mg/mL	0.1 mg/mL	0.1 mg/mL
Creatine (Cr)	—	30 μM	30 μM	12 μM
Creatine phosphate (CP)	—	1 mM	1 mM	1 mM
[ADP]	~10 μM	0.42–0.57 μM	0.42–0.57 μM	~0.02 μM

Values for [ATP], [Pi], and [ADP] represent final concentrations in motility buffers. [Pi] in columns 3 and 5 were estimated using 𝐾_eq=0.06 for the reaction sucrose + Pi ↔α-D-glucose-1-phosphate + D-fructose [52, 54]. [ADP] estimates are based on the equilibrium constant (𝐾_eq) for the reaction ATP + Cr ↔ ADP + CP, and are given in columns 3-4 with lowest and highest value corresponding to the lowest (27°C) and highest (42°C) temperatures employed, respectively. The estimate of [ADP] in column 2 was based on Chase and Kushmerick [53].
¹Motility buffer (MB) employed in the majority of experiments (data in Figures 1–4) did not include sucrose phosphorylase and creatine kinase.
²Modified motility buffers were used for experiments shown in Figure 5.