Table 1. Calculated K_ds and derived ΔG° (Gibbs free energy of binding) for the interactions between eIF4E and the hybrid eIF4G1/PHAGESOL peptides.

Peptide	Sequence	Thermal Shift (Tm, °C)	Estimated Kd derived from the Tm (nM)	SPR derived Kd (nM)		(ΔG°, cal mol−1)
				Keq	Kkin	Keq derived	Kkin derived
eIF4G1-WT	1KKRYDREFLLGF12	7.03±0.1	460	580.2±16.7	523.9±61.6	−8500±20	−8550±70
PHAGESOL	1KKRYSRDQLVAL12	9.83±0.1	77	76.7±3.4	77.1±9.0	−9700±20	−9700±70
eIF4G1-D5S	1KKRYSREFLLGF12	8.97±0.07	100	103.0±2.3	99.9±6.2	−9520±10	−9540±40
eIF4G1-G11A	1KKRYDREFLLAF12	8.13±0.12	200	308.7±11.6	282.0±17.6	−8870±20	−8930±40
eIF4G1-L10V	1KKRYDREFLVGF12	3.9±0.03	4400	4537.0±621.7	2684.7±444.5	−7280±90	−7590±110
eIF4G1-F12L	1KKRYDREFLLGL12	6.87±0.12	520	761.4±63.8	718.1±98.0	−8340±50	−8370±90
eIF4G1-F8Q	1KKRYDREQLLGF12	5. 00±0.03	2000	1633.3±75.1	1777.0±406.1	−7890±30	−7840±150
eIF4G1-E7D	1KKRYDRDFLLGF12	6.87±0.18	520	784.4±21.5	758.8±70.7	−8320±20	−8340±60

The table shows the peptide sequences used to study the relevance of individual amino acid changes observed in the phage derived sequence. The peptides were characterized using a fluorescence based thermal denaturation method and by using SPR with eIF4E amine coupled to the chip surface. K_ds were also derived from the respective thermal shift and SPR data. K_ds were derived from the equilibrium responses (K_eq) and from the association and dissociation phases (K_kin) of the SPR data. The Gibbs free energy of binding (ΔG°) was calculated with the equation ΔG = −RT ln K_a using both dissociation constant values determined.