Table 3.

α-Helical Content and Parameters of Thermal and GdnHCl-Induced Denaturation of WT and Mutant Forms of ApoA-I bound to rHDL Determined by Far-UV CD ^a.

apoA-I	α-helix d (%)	# residues		T1/2f (°C)	Tdmf (°C)	ΔGDo*g (kcal/mol)	ΔGDog (kcal/mol)	D1/2g (M)
		in protein	in helix e
WT b	71 ± 2	248	176	79 ± 0.5	91± 0.5	4.4 ± 0.2	4.1 ± 0.2	2.3 ± 0.05
apoA-I[D102A/D103A] b	62 ± 4*	248	154(-22)	76 ± 1*	89 ± 1	3.0 ± 0.2***	2.4 ± 0.1***	1.9 ± 0.07**
apoA-I[R116V/ K118A] b	66 ± 2*	248	164(-12)	80 ± 1	92 ± 1	4.2 ± 0.3	3.7 ± 0.3	2.0 ± 0.1*
apoA-I[R160V/H162A] b	70 ± 4	248	174	78 ± 0.5	90 ± 1	4.1± 0.3	3.5 ± 0.3	2.3 ± 0.1
apoA-I[Δ(61-78)] b	63 ± 3**	230(-18)	145(-31)	76 ± 1*	89 ± 1	3.1± 0.2***	2.6± 0.2***	1.9 ± 0.1**
WT c	67 ± 2	249	167	-	-	4.6 ± 0.2	4.1± 0.3	2.3 ± 0.0
apoA-I[E110A/E111A] c	63 ± 2*	249	157(-10)	-	-	3.9 ± 0.2*	3.6 ± 0.1*	2.3 ± 0.1

^aValues are the average ± standard deviation from 3 to 6 experiments for two or three independent preparations of each protein. Significance of differences from the value for WT:

^*p<0.05,

^**p<0.01,

^***p<0.005.

^bProteins expressed in the baculovirus expression systems.

^cProteins expressed in the adenovirus expression system.

^dEstimated from the value [Θ₂₂₂] at 25 °C. The values are the average ± S.D. from 8 to 13 measurements for each protein.

^eThe number of residues in the helical conformation as estimated by multiplying the number of residues in the protein by its α-helial content. Values in parentheses show changes in the number of residues as compared to WT.

^fParameters determined from the thermal unfolding curves. T_1/2 is a midpoint of thermal unfolding, at which a half of the total change in CD signal is observed; T^d_m is a maximum of the derivative function d[Θ₂₂₂]/dT.

^gParameters determined from GdnHCl-induced unfolding curves recorded after 68-72 h incubation of rHDL aliquots with various concentrations of GndHCl. Free energy of denaturation, ΔG_D^o*, was determined using denaturant binding model as described in ref 40; ΔG_D^o was determined by the linear extrapolation method (ref 12); the midpoint of denaturation, D_1/2, was determined graphically from the curves and by the linear extrapolation method.