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Literature summary for 2.7.1.23 extracted from

  • Shi, F.; Huan, X.; Wang, X.; Ning, J.
    Overexpression of NAD kinases improves the L-isoleucine biosynthesis in Corynebacterium glutamicum ssp. lactofermentum (2012), Enzyme Microb. Technol., 51, 73-80.
    View publication on PubMed

Application

Application Comment Organism
synthesis overexpressing NAD kinase is a useful metabolic engineering strategy to improve NADPH supply and isoleucine biosynthesis Corynebacterium glutamicum
synthesis overexpressing NAD kinase is a useful metabolic engineering strategy to improve NADPH supply and isoleucine biosynthesis Corynebacterium glutamicum subsp. lactofermentum

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21(DE3) cells Corynebacterium glutamicum
expressed in Escherichia coli BL21(DE3) cells Corynebacterium glutamicum subsp. lactofermentum

Protein Variants

Protein Variants Comment Organism
P117S the mutation results in increased ATP-NAD+ kinase activity and ATP-NADH kinase activity Corynebacterium glutamicum
P117S the mutation results in increased ATP-NAD+ kinase activity and ATP-NADH kinase activity Corynebacterium glutamicum subsp. lactofermentum
P57S the mutation results in increased ATP-NAD+ kinase activity and ATP-NADH kinase activity Corynebacterium glutamicum
P57S the mutation results in increased ATP-NAD+ kinase activity and ATP-NADH kinase activity Corynebacterium glutamicum subsp. lactofermentum
P57S/P117S the double mutation only exhibits a little higher activity than P117S-single point mutation. The catalytic efficiency of the mutant improves greatly, which is 6.8 (for NAD+) and 3.2fold (for ATP) higher than that of the wild type enzyme Corynebacterium glutamicum
P57S/P117S the double mutation only exhibits a little higher activity than P117S-single point mutation. The catalytic efficiency of the mutant improves greatly, which is 6.8 (for NAD+) and 3.2fold (for ATP) higher than that of the wild type enzyme Corynebacterium glutamicum subsp. lactofermentum
S57P the variation is associated with the decreased enzyme activity Corynebacterium glutamicum
S57P the variation is associated with the decreased enzyme activity Corynebacterium glutamicum subsp. lactofermentum

Inhibitors

Inhibitors Comment Organism Structure
NADH
-
Corynebacterium glutamicum
NADH
-
Corynebacterium glutamicum subsp. lactofermentum
NADP+
-
Corynebacterium glutamicum
NADP+
-
Corynebacterium glutamicum subsp. lactofermentum
NADPH
-
Corynebacterium glutamicum
NADPH
-
Corynebacterium glutamicum subsp. lactofermentum

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.07
-
ATP mutant enzyme P57S/P117S, at pH 6.0 and 30°C Corynebacterium glutamicum subsp. lactofermentum
1.4
-
NAD+ wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum
1.41
-
NAD+ mutant enzyme P57S/P117S, at pH 6.0 and 30°C Corynebacterium glutamicum subsp. lactofermentum
1.95
-
ATP wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum
2.11
-
NAD+ mutant enzyme P57S/P117S, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum
2.15
-
ATP wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum
3.73
-
ATP mutant enzyme P57S/P117S, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum
4.02
-
NAD+ wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
36000
-
4 * 36000, SDS-PAGE Corynebacterium glutamicum
36000
-
4 * 36000, SDS-PAGE Corynebacterium glutamicum subsp. lactofermentum
140000
-
gel filtration Corynebacterium glutamicum
140000
-
gel filtration Corynebacterium glutamicum subsp. lactofermentum

Organism

Organism UniProt Comment Textmining
Corynebacterium glutamicum
-
subsp. lactofermentum
-
Corynebacterium glutamicum ATCC 13869
-
subsp. lactofermentum
-
Corynebacterium glutamicum subsp. lactofermentum
-
-
-
Corynebacterium glutamicum subsp. lactofermentum JHI3-156
-
-
-

Purification (Commentary)

Purification (Comment) Organism
Ni-NTA column chromatography, gel filtration Corynebacterium glutamicum
Ni-NTA column chromatography, gel filtration Corynebacterium glutamicum subsp. lactofermentum

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + NAD+ ATP is the preferred phosphoryl donor and NAD+ is the preferred acceptor Corynebacterium glutamicum ADP + NADP+
-
?
ATP + NAD+ ATP is the preferred phosphoryl donor and NAD+ is the preferred acceptor Corynebacterium glutamicum subsp. lactofermentum ADP + NADP+
-
?
ATP + NAD+ ATP is the preferred phosphoryl donor and NAD+ is the preferred acceptor Corynebacterium glutamicum ATCC 13869 ADP + NADP+
-
?
ATP + NAD+ ATP is the preferred phosphoryl donor and NAD+ is the preferred acceptor Corynebacterium glutamicum subsp. lactofermentum JHI3-156 ADP + NADP+
-
?
hexaphosphate + NAD+
-
Corynebacterium glutamicum subsp. lactofermentum pentaphosphate + NADP+
-
?
hexaphosphate + NAD+
-
Corynebacterium glutamicum subsp. lactofermentum JHI3-156 pentaphosphate + NADP+
-
?
hexapolyphosphate + NAD+
-
Corynebacterium glutamicum pentapolyphosphate + NADP+
-
?
hexapolyphosphate + NAD+
-
Corynebacterium glutamicum ATCC 13869 pentapolyphosphate + NADP+
-
?

Subunits

Subunits Comment Organism
homotetramer 4 * 36000, SDS-PAGE Corynebacterium glutamicum
homotetramer 4 * 36000, SDS-PAGE Corynebacterium glutamicum subsp. lactofermentum

Synonyms

Synonyms Comment Organism
NAD kinase
-
Corynebacterium glutamicum
NAD kinase
-
Corynebacterium glutamicum subsp. lactofermentum
poly(P)/ATPdependent NAD kinase
-
Corynebacterium glutamicum
poly(P)/ATPdependent NAD kinase
-
Corynebacterium glutamicum subsp. lactofermentum
Ppnk
-
Corynebacterium glutamicum
Ppnk
-
Corynebacterium glutamicum subsp. lactofermentum

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
70
-
preincubating the enzyme for 2 h at 70°C results in a loss of 50% of its activity Corynebacterium glutamicum
70
-
preheating the enzyme for 2 h at temperatures up to 70°C does not alter the ATP-NAD+ kinase activity significantly (less than 10% of the activity lost) Corynebacterium glutamicum subsp. lactofermentum

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.5
-
NAD+ mutant enzyme P57S/P117S, at pH 6.0 and 30°C Corynebacterium glutamicum subsp. lactofermentum
0.52
-
ATP wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum
0.54
-
NAD+ wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum
0.6
-
ATP mutant enzyme P57S/P117S, at pH 6.0 and 30°C Corynebacterium glutamicum subsp. lactofermentum
2.54
-
ATP wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum
3.8
-
NAD+ wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum
15.42
-
NAD+ mutant enzyme P57S/P117S, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum
20.51
-
ATP mutant enzyme P57S/P117S, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
-
Corynebacterium glutamicum subsp. lactofermentum
8
-
-
Corynebacterium glutamicum

pH Stability

pH Stability pH Stability Maximum Comment Organism
7
-
the enzyme remains stable at pH 7.0 Corynebacterium glutamicum subsp. lactofermentum
7 8 the enzyme is precipitated and denatured at pH 7.0. Even at pH 8.0, the enzyme is depolymerized and denatured after long-time storing Corynebacterium glutamicum

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.17
-
NADPH at pH 8.0 and 30°C Corynebacterium glutamicum
0.21
-
NADP+ at pH 8.0 and 30°C Corynebacterium glutamicum
0.22
-
NADPH at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum
0.24
-
NADH at pH 8.0 and 30°C Corynebacterium glutamicum
0.43
-
NADP+ at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum
0.62
-
NADH at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum

General Information

General Information Comment Organism
metabolism NAD kinase is the key enzyme for the de novo biosynthesis of NADP+ and NADPH Corynebacterium glutamicum
metabolism NAD kinase is the key enzyme for the de novo biosynthesis of NADP+ and NADPH Corynebacterium glutamicum subsp. lactofermentum

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.24
-
ATP wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum
0.36
-
NAD+ mutant enzyme P57S/P117S, at pH 6.0 and 30°C Corynebacterium glutamicum subsp. lactofermentum
0.39
-
NAD+ wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum
0.56
-
ATP mutant enzyme P57S/P117S, at pH 6.0 and 30°C Corynebacterium glutamicum subsp. lactofermentum
0.94
-
NAD+ wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum
1.3
-
ATP wild type enzyme, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum
5.5
-
ATP mutant enzyme P57S/P117S, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum
7.32
-
NAD+ mutant enzyme P57S/P117S, at pH 7.5 and 30°C Corynebacterium glutamicum subsp. lactofermentum