Table 1.

Computational Details of the 2D‐US Simulations

	Residues	Linker conformation	Kinase domain conformation	# Windows	ns/window	k kcal/(mol·Å2)	d w (Å)
WT	253‐523	Restricted↔released	Inactive	245	30	10	0.5
WT	253‐523	Restricted↔released	Active	238	30	10	0.5
W260A	253‐523	Restricted↔released	Inactive	245	40	10	0.5
WT	260‐521	n/a	Inactive↔active	1101	15	20	0.25
W260A	260‐521	n/a	Inactive↔active	681	16	20	0.5

Supporting Information Fig. S3 shows the sequence and the structural motifs of Src which are utilized in the present work. The conformation of the kinase domain in each one of the top three constructs was not subject to any restraining potential and was found to be stable during umbrella sampling calculations of the linker region. Y416 in the A‐loop is not phosphorylated in all five sets of umbrella sampling calculations. “n/a” stands for not applicable.