Table 2. Reduction rates of F₄₂₀H₂-dependent menadione and nitroimidazole reductase activities of Ddn mutants.

	Rate of Reaction (nmol min-1 μmolenzyme-1)a
Ddn Mutants	Menadione	Pretomanid	Delamanid
M. tuberculosis mutants selected for pretomanid resistance in laboratory
M1T	N/A	N/A	N/A
S22L	5.03×102 ± 4.34×101	N/D	N/D
L49P*	4.99×101 ± 7.87	N/D	N/D
L64P	4.13×101 ± 1.12	N/D	N/D
W88R*	2.16×101 ± 7.18×10−2	N/D	N/D
InsD108	N/A	N/A	N/A
R112W	1.96×103 ± 2.82×102	N/D	N/D
C149Y	2.36×102 ± 1.25×101	N/D	2.13×101 ± 1.65
Ddn and orthologs
Ddn Mtb	8.49×104 ± 9.32×103	3.66×102 ± 3.3×101	1.03×102 ± 7.59
MMAR_5035	7.99×103 ± 9.11×102	2.3×102 ± 1.44×101	3.26×101 ± 7.11
MUL_4109	2.21×101 ± 2.71	N/D	N/D
MVAN_5261	5.74×103 ± 7.87×101	N/D	N/D
MAV_0613	4.51×103 ± 1.24×103	N/D	N/D
MSMEG_5998	7.35×103 ± 7.91×102	N/D	N/D
Mutations based on naturally occurring sequence polymorphisms in M. tuberculosis
P6S	3.74×104 ± 8.74×103	3.29×102 ± 1.63×101	8.94×101 ± 4.64
P6T	5.07×104 ± 5.16×103	5.07×102 ± 3.18×101	1.13×102 ± 4.17
P6L	8.24×104 ± 1.46×103	1.94×102 ± 2.84	5.84×101 ± 1.23×101
M21T	1.52×104 ± 1.26×103	3.65×102 ± 8.99	7.29×101 ± 3.60
R23L	4.59×104 ± 3.28×103	4.95×102 ± 1.02×101	1.20×102 ± 3.93
R23W	9.86×104 ± 9.98×103	2.00×102 ± 1.37×101	9.15×101 ± 3.25×101
T26P	3.29×103 ± 3.43×102	6.21×101 ± 9.18	5.82 ± 3.10
W27C	1.44×104 ± 2.10×102	1.66×102 ± 5.98	4.19×101 ± 1.91
Y29H	1.51×104 ± 1.54×103	5.11×101 ± 7.60	1.77×101 ± 4.81
Y29S	7.25×103 ± 6.67×102	3.88×101 ± 9.51×10−1	7.15 ± 2.69
R30S	6.04×104 ± 1.86×103	1.44×102 ± 1.81×101	7.11×101 ± 7.87
G34E	6.65×104 ± 3.10×103	7.39×101 ± 4.72	5.29×101 ± 6.98
G34R	9.14×103 ± 7.79×102	3.36×101 ± 5.92	4.19×101 ± 2.77
G36V	4.78×104 ± 5.09×102	8.16×101 ± 1.36×101	4.91×101 ± 1.88
P45L	7.69×102 ± 5.92×101	8.29×101 ± 1.52×101	7.78×101 ± 1.05
L49P	4.99×101 ± 7.87	N/D	N/D
T50P	3.36×102 ± 6.07×101	5.37×101 ± 7.75	5.65×101 ± 5.90
T51P	1.35×103 ± 1×102	3.70×101 ± 4.80	2.87×101 ± 5.04×10−1
T52N	7.61×104 ± 1.01×104	2.74×102 ± 6.21	1.19×102 ± 7.13
T52P	9.66×103 ± 3.38×102	8.73×101 ± 6.03	5.07×101 ± 3.04
T56P	1.20×104 ± 2.84×103	1.01×101 ± 8.60×10−1	2.79×101 ± 9.43×10−1
G57A	5.60×104 ± 1.47×103	2.65×102 ± 1.32×101	9.69×101 ± 9.43
V61G	4.98×104 ± 3.05×103	1.01×102 ± 2.25	5.46×101 ± 6.00
N62D	1.22×103 ± 1.62×102	1.68×101 ±9.60×10−1	2.19×101 ± 1.80
Y65S	1.12×103 ± 8.63×101	N/D	N/D
L67P	6.86×102 ± 8.52	3.58×101 ± 6.73×10−1	1.55×101 ± 1
D69N	6.33×104 ± 1.28×103	2.94×102 ± 4.06	1.25×102 ± 2.00×101
G71R	9.49×104 ± 4.63×103	3.71×102 ± 6.81	1.31×102 ± 1.86
R72Q	5.20×104 ± 9.28×102	2.11×102 ± 1.14×101	8.32×101 ± 4.33
R72W	5.08×104 ± 1.31×103	2.27×102 ± 1.47	1.07×102 ± 1.99
S78Y	1.41×101 ± 3.05	N/D	1.21×101 ± 2.72
K79Q	7.95×103 ± 8.05×102	N/D	N/D
G81S	1.52×104 ± 1.26×103	3.00×101 ± 5.93	2.28×101 ± 2.81
E83D	6.93×104 ± 3.28×103	4.22×102 ± 6.09×101	1.56×102 ± 1.04×101
W88R	2.16×101 ± 7.18×10−2	N/D	N/D
L90V	6.61×104 ± 2.60×103	2.49×102 ± 1.23×101	7.68×101 ± 8.40
N91T	2.24×103 ± 5.07×102	3.19×101 ± 5.85	3.00×101 ± 1.45
I102V	7.60×104 ± 4.82×103	2.88×102 ± 1.10×101	8.25×101 ± 1.54×101
E105Q	1.08×105 ± 1.60×104	3.49×102 ± 7.56	1.23×102 ± 1.78
A111V	5.62×102 ± 2.25×101	4.22×101 ± 7.77	2.24×101 ± 3.64
D113N	3.45×104 ± 1.31×103	1.94×102 ± 1.26×101	4.48×101 ± 5.85
E117K	7.07×104 ± 2.11×103	3.13×102 ± 5.83	9.15×101 ± 8.51
P124S	7.18×104 ± 2.48×103	2.92×102 ± 3.35	8.75×101 ± 1.03×101
Y133C	1.82×103 ± 1.54×102	N/D	3.90 ± 1.95
T140I	7.28×104 ± 4.09×102	1.66×102 ± 2.40×101	1.03×102 ± 3.50
V147M	4.38×104 ± 4.29×103	1.51×102 ± 5.97	6.40×101 ± 7.73
Potential spontaneous mutations within the substrate binding site of Ddn
Y65L	4.07×104 ± 3.22×103	N/D	6.66×101 ± 5.23
Y65M	6.72×104 ± 5.78×103	N/D	2.25×101 ± 2.82
Y65S*	1.12×103 ± 8.63×101	N/D	N/D
Y65C	3.11×104 ± 9.27×102	N/D	4.43×101 ± 6.39
Y65F	3.59×104 ± 2.09×103	2.93×102 ± 1.82×101	2.41×102 ± 1.35×101
S78A	4.17×104 ± 2.47×103	N/D	1.20×101 ± 1.68
S78C	4.08×103 ± 1.54×102	N/D	2.40×101 ± 4.13
S78T	6.15×104 ± 6.77×103	N/D	1.90×101 ± 1.75
S78V	1.12×103 ± 2.00×102	N/D	6.54 ± 3.87
S78Y*	1.41×101 ± 3.05	N/D	1.21×101 ± 2.72
Y130C	3.31×103 ± 3.92×102	7.62×101 ± 2.49	5.15×101 ± 3.85
Y130D	7.08×103 ± 2.17×103	3.48×101 ± 2.57	2.42×101 ± 1.3
Y130F	1.41×104 ± 2.12×103	2.75×101 ± 7.45	4.92×101 ± 4.82×10−1
Y130H	4.63×103 ± 4.05×102	4.02×101 ± 8.18	3.19×101 ± 1.02×101
Y130N	3.65×103 ± 2.07×102	7.66×101 ± 8.82	3.57×101 ± 4.03
Y130S	1.35×104 ± 1.46×103	2.19×102 ± 2.24 ×101	3.66×101 ± 1.31×101
Y130W	1.28×104 ± 1.39×103	9.77×101 ± 2.57	5.53×101 ± 4.63
Y133C*	1.82×103 ± 1.54×102	N/D	3.90 ± 1.95
Y133F	4.08×104 ± 3.03×103	1.41×102 ± 6.54	7.82×101 ± 3.58
Y133L	8.87×104 ± 1.10×103	N/D	2.78×101 ±5.64
Y133M	5.82×104 ± 3.57×103	N/D	1.64×101 ± 2.32
Y133W	1.45×104 ± 6.57×102	N/D	7.80 ± 4.47×10−1
Y136E	1.24×102 ± 6.10	N/D	4.51×101 ± 1.93
Y136T	6.86×102 ± 1.85×101	N/D	3.67 ± 1.17×10−1
Y136F	5.08×104 ± 5.64×103	3.88×102 ± 3.07×101	1.05×102 ±1.29×101
Y136S	5.13×102 ± 1.38×101	N/D	N/D

N/D–Below limit of detection, N/A–Enzyme was unable to express,

*—found in naturally occurring sequence polymorphisms

^a–Data from this table is also used in Table 3, and Figs 4 and 6