Table 1.

Summary of the dissociation constants, the changes in the heat capacity upon binding, and the kinetic rate constants of the TRTK12 wild-type peptide and alanine variants binding to S100B at 25°C.

Peptide	Kda μM	ΔCp, expb kJ·mol−1 K−1	ΔCp, calcc kJ·mol−1 K−1	koff (s−1)d	kon (1×107 M−1s−1)e
TRTK12	1.7 ± 0.1 (3 ± 1)	−1.2 ± 0.2	−1.4 ± 0.1	53 ± 4	3.1 ± 0.3
TRTKM1	2.9 ± 1.0 (0.2 ± 0.1)	−1.2 ± 0.1	−1.3 ± 0.1	34 ± 2	1.2 ± 0.1
TRTKM5	1.2 ± 0.4 (0.6 ± 0.6)	−1.0 ± 0.1	−1.2 ± 0.1	66 ± 7	5.5 ± 1.9
TRTKM6	1.3 ± 0.6 (5 ± 1)	−1.3 ± 0.1	−1.5 ± 0.1	31 ± 1	6.6 ± 1.0
TRTKM9	1.7 ± 0.1 (2 ± 1)	−1.5 ± 0.1	−1.5 ± 0.1	53 ± 8	3.1 ± 0.5
TRTKM10	3.4 ± 1.0 (1 ± 0.1)	−1.0 ± 0.1	−1.1 ± 0.1	36 ± 8	1.1 ± 0.4
TRTKM11	3.8 ± 1.0 (10 ± 3)	−1.6 ± 0.2	−1.3 ± 0.1	n.a.	n.a.

^aDissociation constants obtained using ITC. Values in parentheses were obtained using emission fluorescence assays.

^bThe experimentally determined changes in the heat capacity upon binding (Figure 3).

^cStructure-based calculations (see Eq. 7) using the average value from homology models based on the human S100B dimer binding to two peptides using PDB: 1MQ1 ⁽⁴³⁾, 1MWN ⁽⁴⁰⁾, and 3IQQ ⁽⁴⁴⁾.

^dk_off values obtained using fluorescence stopped-flow spectroscopy (Eq 9).

^ek_on values were calculated using k_off rate constants and the dissociation constants obtained from ITC.