f60

Stability/Folding

Mole-Fraction of Unfolded Protein (fT)

unitless

Fluorescence Intensity

None

60 °C

None

None

None

None

Fluorescence intensity ratio at 350 nm: 330 nm vs. temperature was fitted to a two-state transition model:

IT = {IN+aT+(ID+bT)exp[(ΔHvh/R)(1/Tm−1/T)]} / {1+exp[(ΔHvh/R)(1/Tm−1/T)]}

where IT is the observed signal; IN and ID, the native and denatured baseline intercepts; a and b, the native and denatured baseline slopes; T, the temperature; ∆Hvh, the van’t Hoff enthalpy; R, the gas constant (1.987 cal⋅mol−1⋅K−1); and Tm, the thermal transition midpoint. The mole-fraction, fT, of unfolded protein at any temperature T was calculated from:

fT = {exp[(ΔHvh/R)(1/Tm−1/T)]} / {1+exp[(ΔHvh/R)(1/Tm−1/T)]}