Table 1.

Kinetic constants for the native and evolved enzymes

Substrate	Kinetic constant	Native galactosidase	Evolved fucosidase
PNPG	kcat, s−1	268	30.9
	Km, mM	0.04	0.18
	kcat/Km, mM−1⋅s−1	6,700	172
PNPF	kcat, s−1	209	96.6
	Km, mM	31	1.5
	kcat/Km, mM−1⋅s−1	6.7	64.4
Specificity		1,000	2.7
ONPG	kcat, s−1	765	14.5
	Km, mM	0.11	0.11
	kcat/Km, mM−1⋅s−1	6,950	132
ONPF	kcat, s−1	—	24.1
	Km, mM	—	0.55
	kcat/Km, mM−1⋅s−1	(2)*	43.9
Specificity		3,200	3.0

The native galactosidase and the evolved fucosidase were purified, and the enzymes were assayed on four different substrates. 

^*The k_cat/K_m value for the native galactosidase on ONPF was estimated to be about 2 mM⁻¹⋅s⁻¹ by measuring the hydrolysis rate relative to that of ONPG.