Table 2.

Calculated association constants at transition temperatures for EI(H189A) and wild-type, dephospho-EI binding ligands from DSC and fluorescence measurementsa

		ΔHobsb	ΔHcc
Protein μM subunit	Ligands (mM)	(kcal/mole)		Tmb (°C)	KA′ at Tm (M−1)
EI(H189A), 3.15	—	45	—	45.4	—
EI(H189A), 4.09	1 PEPd	70	59.85	49.9	2.17 × 103
EI(H189A), 3.31	5 Pyr/2 Mg2+	83	60.84	50.2	4.91 × 102
EI(H189A), 3.46	1 PEP/2 Mg2+	112	105.39	63.7	6.05 × 105
EI(wt), 2.36-7.23	—	75.5	—	46.3	—
EI(wt), 3.30	0.65 PPyr/2 Mg2+	136	130.94	63.1	5.02 × 106

^a Proteins are in 20 mM K-phosphate and 100 mM KCl (pH 7.5), and scan rates at 30°C/h.

^b Enthalpy changes at T_m (ΔH_obs) and transition temperatures (T_m) are averages of 2–4 separate determinations from two, two-state analysis of DSC data in the absence of effectors when two endotherms are resolved and, in the presence of effectors, from two-state analyses of Trp fluorescence changes as a function of increasing temperature for C-terminal domain unfolding (Nosworthy et al. 1998).

^c Values of ΔH_c are calculated from Equation 2.

^d From Dimitrova et al. (2002).