Table III.

Parameters determined from the versus pH profiles of wild-type eCBL and the R58A, R58K and R372K site-directed mutants^a

Enzyme	pKa1	pKa2
eCBL	8.28 ± 0.06	10.20 ± 0.06
R58A	7.93 ± 0.08	9.53 ± 0.09
R58K	8.94 ± 0.09	9.8 ± 0.1
R372K	8.44 ± 0.09	9.8 ± 0.1

^aKinetic measurements for the eCBL-catalyzed hydrolysis of l-Cth were carried out from pH 6.4–10.6 in MBP buffer containing 0.01–6.25 mM l-Cth, 20 μM PLP, 2 mM DTNB and 0.068–4.5 μM enzyme at 25°C. The k_cat/ versus pH data were fitted to equation 2 to obtain the values for pK_a1 and pK_a2.