Table 2.

Predicted Relative Binding Free Energy of Each Mutation of 3HFM at Different Transition Times between 1 to 8 ns for 100 Independent Transitions^a

	ΔΔG (kcal/mol)
mutations (3HFM)	experimental	1 ns	2 ns	3 ns	4 ns	5 ns	6 ns	7 ns	8 ns	9 ns	10 ns
Y20F	−0.48	−7.53	−6.34	−3.45	−2.95	−0.34	−1.02	−0.83	0.07	−0.69	−0.98
D32N	0.17 ± 0.3	−3.47	−5.67	−2.99	−1.32	−1.59	−1.98	−1.14	0.53
R21A	0.90	5.82	7.54	4.56	3.98	1.23	2.34	1.74	1.35
D101K	2.13	16.98	13.27	7.43	3.59	−0.46	−1.27	0.87	0.12
W98F	3.25 ± 0.16	7.49	5.89	6.78	2.35	−0.16	−0.87	0.46	2.23
Y50L	4.39 ± 0.12	9.65	6.37	2.36	1.33	0.10	1.37	2.89	3.26
N31E	5.71 ± 0.13	13.28	8.73	9.67	4.78	−1.26	0.56	2.34	3.52
K97D	6.77 ± 0.14	10.25	10.47	7.09	5.36	9.00	7.20	8.33	6.83	7.86	8.24

^aEstimated ΔΔG values of all eight mutations of the 3HFM system for 100 independent transitions. The predicted ΔΔG values were compared with the corresponding experimental data. ΔΔG values beyond 8 ns of transition time are for test mutations Y20F and K97D as a part of the convergence study.