Table 1.

Summary of Af1521-GFP fusion proteins evaluated in this study and their ADP-ribosyl hydrolase rate constants.

Construct	Mutations*	kcat **	R2
eAf1521	K35E, Y74C, F97L, S105G, E110G, Y145R, N162D	0.095 ± 0.007	0.9882
Af1521-c001	wild type	0.153 ± 0.018	0.9638
Af1521-c002	K35E,Y145R	0.107 ± 0.008	0.9935
Af1521-c003	K35E,G143E,Y145R	0.051 ± 0.012	0.9483
Af1521-c004	N34S,K35E,Y145R	0.098 ± 0.008	0.9904
Af1521-c005	K35E,R36N,Y145R	0.097 ± 0.008	0.9907
Af1521-c006	G42E, I144R, Y145N	0.146 ± 0.017	0.9778
Af1521-c007 (“MacroGreen”)	K35E, G42E, I144R, Y145R	0.075 ± 0.007	0.9812
Af1521-c008	K35E, G42E, I144R, Y145N	0.139 ± 0.018	0.9744

*Bold: eAf1521 mutations essential for nanomolar binding of free ADP-ribose

** Determined by fitting to a first order equation. N = 2 to 4; n = 3 (original data exemplified graphically in Supplementary Fig. 1b).