. 2019 Jun 24;9:9081. doi: 10.1038/s41598-019-45282-0

Table 2.

Dissociation constants for the monomer-dimer-tetramer equilibrium of the exosite mutant human m-NAD(P)-ME in the presence of NAD⁺ or ATP.

m-NAD(P)-ME	Protein only	With NAD⁺	With ATP
WT	0.24 ± 0.002	0.35 ± 0.003	0.66 ± 0.01 (2.8)	0.05 ± 0.001 (0.14)	0.64 ± 0.01 (2.7)	2.72 ± 0.06 (7.8)
H154V	0.03 ± 0.002	0.46 ± 0.003	0.07 ± 0.001 (2.3)	0.16 ± 0.001 (0.3)	0.46 ± 0.01 (15.3)	4.03 ± 0.04 (8.7)
K156A	0.12 ± 0.004	0.32 ± 0.01	1.36 ± 0.05 (11.3)	0.08 ± 0.003 (0.3)	0.32 ± 0.01 (2.7)	0.80 ± 0.02 (2.5)
G192A	0.09 ± 0.002	0.30 ± 0.001	0.06 ± 0.002 (0.7)	0.18 ± 0.01 (0.6)	0.21 ± 0.01 (2.3)	0.40 ± 0.01 (1.3)
R194N	0.31 ± 0.01	0.23 ± 0.01	0.17 ± 0.01 (0.5)	0.04 ± 0.001 (0.2)	0.08 ± 0.001 (0.3)	1.44 ± 0.03 (6.3)
R197E	0.03 ± 0.001	2.33 ± 0.08	0.10 ± 0.003 (3.3)	1.11 ± 0.04 (0.5)	0.10 ± 0.003 (3.3)	0.59 ± 0.02 (0.3)
R197D	0.80 ± 0.02	0.26 ± 0.01	0.06 ± 0.003 (0.1)	0.81 ± 0.04 (3.1)	0.69 ± 0.02 (0.9)	0.11 ± 0.003 (0.4)
N482G	0.05 ± 0.01	0.18 ± 0.002	0.38 ± 0.002 (7.6)	0.04 ± 0.01 (0.2)	0.10 ± 0.003 (2.0)	0.06 ± 0.002 (0.3)
N482A	0.29 ± 0.01	0.95 ± 0.03	0.58 ± 0.03 (2.0)	0.97 ± 0.05 (1.0)	1.44 ± 0.04 (49.7)	0.35 ± 0.01 (0.4)
R542V	0.01 ± 0.001	0.44 ± 0.01	1.87 ± 0.08 (187.0)	0.21 ± 0.01 (0.5)	0.02 ± 0.001 (2.0)	9.87 ± 0.38 (22.4)
Y552F	0.05 ± 0.001	0.51 ± 0.01	1.57 ± 0.11 (31.4)	0.09 ± 0.01 (0.2)	0.01 ± 0.001 (0.2)	2.58 ± 0.07 (5.1)
R556Q	0.01 ± 0.003	1.75 ± 0.06	0.18 ± 0.01 (18.0)	0.91 ± 0.03 (0.5)	0.40 ± 0.01 (40.0)	0.18 ± 0.01 (0.1)
R556V	0.13 ± 0.001	2.30 ± 0.02	0.60 ± 0.01 (4.6)	4.07 ± 0.08 (1.8)	2.03 ± 0.06 (15.6)	0.18 ± 0.01 (0.08)

m-NAD(P)-ME

Protein only

With NAD⁺

With ATP

K_d,12^a (µM)

K_d,24^b (µM)

K_d,12(NAD)^c (µM) (K_d,12(NAD)/K_d,12)

K_d,24(NAD)^d (µM) (K_d,24(NAD)/K_d,24)

K_d,12(ATP)^e (µM) (K_d,12(ATP)/K_d,12)

K_d,24(ATP)^f (µM) (K_d,24(ATP)/K_d,24)

0.24 ± 0.002

0.35 ± 0.003

0.66 ± 0.01

(2.8)

0.05 ± 0.001

(0.14)

0.64 ± 0.01

(2.7)

2.72 ± 0.06

(7.8)

H154V

0.03 ± 0.002

0.46 ± 0.003

0.07 ± 0.001

(2.3)

0.16 ± 0.001

(0.3)

0.46 ± 0.01

(15.3)

4.03 ± 0.04

(8.7)

K156A

0.12 ± 0.004

0.32 ± 0.01

1.36 ± 0.05

(11.3)

0.08 ± 0.003

(0.3)

0.32 ± 0.01

(2.7)

0.80 ± 0.02

(2.5)

G192A

0.09 ± 0.002

0.30 ± 0.001

0.06 ± 0.002

(0.7)

0.18 ± 0.01

(0.6)

0.21 ± 0.01

(2.3)

0.40 ± 0.01

(1.3)

R194N

0.31 ± 0.01

0.23 ± 0.01

0.17 ± 0.01

(0.5)

0.04 ± 0.001

(0.2)

0.08 ± 0.001

(0.3)

1.44 ± 0.03

(6.3)

R197E

0.03 ± 0.001

2.33 ± 0.08

0.10 ± 0.003

(3.3)

1.11 ± 0.04

(0.5)

0.10 ± 0.003

(3.3)

0.59 ± 0.02

(0.3)

R197D

0.80 ± 0.02

0.26 ± 0.01

0.06 ± 0.003

(0.1)

0.81 ± 0.04

(3.1)

0.69 ± 0.02

(0.9)

0.11 ± 0.003

(0.4)

N482G

0.05 ± 0.01

0.18 ± 0.002

0.38 ± 0.002

(7.6)

0.04 ± 0.01

(0.2)

0.10 ± 0.003

(2.0)

0.06 ± 0.002

(0.3)

N482A

0.29 ± 0.01

0.95 ± 0.03

0.58 ± 0.03

(2.0)

0.97 ± 0.05

(1.0)

1.44 ± 0.04

(49.7)

0.35 ± 0.01

(0.4)

R542V

0.01 ± 0.001

0.44 ± 0.01

1.87 ± 0.08

(187.0)

0.21 ± 0.01

(0.5)

0.02 ± 0.001

(2.0)

9.87 ± 0.38

(22.4)

Y552F

0.05 ± 0.001

0.51 ± 0.01

1.57 ± 0.11

(31.4)

0.09 ± 0.01

(0.2)

0.01 ± 0.001

(0.2)

2.58 ± 0.07

(5.1)

R556Q

0.01 ± 0.003

1.75 ± 0.06

0.18 ± 0.01

(18.0)

0.91 ± 0.03

(0.5)

0.40 ± 0.01

(40.0)

0.18 ± 0.01

(0.1)

R556V

0.13 ± 0.001

2.30 ± 0.02

0.60 ± 0.01

(4.6)

4.07 ± 0.08

(1.8)

2.03 ± 0.06

(15.6)

0.18 ± 0.01

(0.08)

^aK_d,12 and ^bK_d,24 indicate the monomer-dimer and dimer-tetramer dissociation constants, respectively; ^cK_d,12(NAD) and ^dK_d,24(NAD) indicate the monomer-dimer and dimer-tetramer dissociation constants, respectively, with 0.4 mM NAD⁺ treatment; ^eK_d,12(ATP) and ^fK_d,24(ATP) indicate the monomer-dimer and dimer-tetramer dissociation constants, respectively, with 0.5 mM ATP treatment.