. 2014 Aug 19;289(40):28006–28018. doi: 10.1074/jbc.M114.589077

TABLE 1.

Kinetic and equilibrium binding parameters for the formation of SK·Pg and SKΔK414·Pg complexes

Kinetic constants obtained from simultaneous numerical integration of the forward and reverse reactions^a, forward reactions measured as anisotropy changes^aa, and SK dependences of the fast and slow phases of the forward reactions^b are listed for reaction Scheme 1 in the absence of kringle ligands (no effector) and in the presence of saturating 6-AHA or benzamidine. K_D, _overall was calculated from the individual kinetic parameters^c and measured by fluorescence titration^d. For analysis of near-linear SK dependences K₁ was fixed to the value obtained by numerical analysis^fixed. Amplitudes of change in fluorescence intensity (ΔF_max/F_o) were from numerical analysis (f₂ and f₃ for SK·Pg 2 and SK·Pg 3, respectively) and equilibrium binding (overall value). Reported errors are 2 × S.D. and were calculated by error propagation for compound parameters.

	K_{1 (encounter)}	k₂ (≈k_{lim 1})	k₋₂ (≈k_{off 1})	k₃ (≈k_{lim 2})	k₋₃ (≈k_{off 2})	K_D, _overall	ΔF_max/F_o
	μm	s⁻¹	s⁻¹	s⁻¹	s⁻¹	μm	%

[5F]FFR-[Lys]Pg, no effector
nSK	2.8 ± 0.3^a	34 ± 2^a	3.5 ± 0.4^a	0.34 ± 0.04^a	0.15 ± 0.01^a	0.086 ± 0.018^a,c	−30 ± 9^a, −25 ± 6^a
	1.8 ± 0.2^aa	33 ± 2^aa	1.8 ± 0.1^aa	0.26 ± 0.06^aa	0.12 ± 0.02^aa	0.030 ± 0.018^aa,c
	3.4 ± 1.0^b	33 ± 3^b	4.0 ± 1.0^b	0.90 ± 0.30^b	0.25 ± 0.10^b	0.044 ± 0.009^d	−28 ± 1^d
WT-SK	0.94 ± 0.01^a	27 ± 1^a	3.5 ± 0.2^a	0.44 ± 0.03^a	0.22 ± 0.01^a	0.039 ± 0.005^a,c	−26 ± 8^a, −24 ± 4^a
	2.1 ± 1.2^b	30 ± 6^b	4.5 ± 2.0^b	3.60 ± 1.60^b	0.24 ± 0.16^b	0.028 ± 0.006^d	−23 ± 1^d
SKΔK414	20.3 ± 0.2^a	31 ± 1^a	2.5 ± 0.1^a	0.17 ± 0.01^a	0.20 ± 0.01^a	0.84 ± 0.09^a,c	−24 ± 4^a, −30 ± 5^a
	19.0 ± 4.0^b	25 ± 4^b	2.3 ± 0.6^b	0.10 ± 0.08^b	0.30 ± 0.06^b	0.60 ± 0.20^d	−24 ± 2^d

6-AHA
nSK	10.5 ± 0.7^a	28 ± 3^a	3.7 ± 0.2^a	0.10 ± 0.02^a	0.23 ± 0.01^a	0.89 ± 0.20^a,c	−22 ± 8^a, −37 ± 8^a
	7.0 ± 5.0^b	25 ± 8^b	2.0 ± 1.0^b	2.00 ± 2.00^b	0.19 ± 0.09^b	0.56 ± 0.09^d	−23 ± 1^d
SKΔK414	16.3 ± 0.6^a	31 ± 2^a	2.7 ± 0.1^a	0.19 ± 0.02^a	0.16 ± 0.01^a	0.64 ± 0.10^a,c	−23 ± 6^a, −29 ± 3^a
	14.0 ± 2.0^b	31 ± 3^b	1.0 ± 0.6^b	0.15 ± 0.18^b	0.23 ± 0.02^b	0.75 ± 0.35^d	−24 ± 3^d

Benzamidine
nSK	11.4 ± 0.3^a	156 ± 3^a	3.2 ± 0.1^a	0.07 ± 0.01^a	0.15 ± 0.02^a	0.16 ± 0.04^a,c	−18 ± 7^a, −17 ± 8^a
	12.0 ± 2.0^b	150 ± 15^b	2.0 ± 1.0^b			0.20 ± 0.02^d	−30 ± 1^d
SKΔK414	16.0 ± 0.3^a	69 ± 4^a	4.8 ± 0.2^a	0.26 ± 0.02^a	0.24 ± 0.02^a	0.52 ± 0.06^a,c	−27 ± 7^a, −23 ± 3^a
	16.0 ± 10.0^b	70 ± 30^b	4.0 ± 2.0^b	1.30 ± 0.40^b	0.27 ± 0.10^b	0.80 ± 0.10^d	−38 ± 1^d

[5F]FFR-[Glu]Pg, no effector
nSK	18.1 ± 0.2^a	42 ± 1^a	2.8 ± 0.1^a	0.10 ± 0.01^a	0.19 ± 0.01^a	0.76 ± 0.20^a,c	−31 ± 3^a, −25 ± 4^a
	18.1^b,fixed	46 ± 5^b	2.0 ± 0.5^b	1.00 ± 0.20^b	0.17 ± 0.02^b	0.58 ± 0.08^d	−32 ± 1^d

6-AHA
nSK	15.4 ± 0.2^a	30 ± 1^a	3.2 ± 0.1^a	0.35 ± 0.01^a	0.19 ± 0.01^a	0.56 ± 0.08^a,c	−18 ± 8^a, −16 ± 7^a
	15.4^b,fixed	36 ± 5^b	1.8 ± 0.6^b	2.94 ± 0.21^b	0.12 ± 0.03^b	0.60 ± 0.07^d	−23 ± 1^d