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Literature summary for 1.4.1.9 extracted from

  • Sekimoto, T.; Fukui, T.; Tanizawa, K.
    Role of the conserved glycyl residues located at the active site of leucine dehydrogenase from Bacillus stearothermophilus (1994), J. Biochem., 116, 176-182.
    View publication on PubMed
Search for more literature for 1.4.1.9

Protein Variants

Protein Variants Comment Organism
G77A turnover numver in oxidative deamination of L-Leu is 36% of that of the wild-type enzyme. In reductive amination the turnover number is comparable to that of the wild-type enzyme. The Km-value for 2-oxoisohexanoate is 6.3fold higher and the Km-value for NH4+ is 2.8fold higher than that of the wild-type enzyme. Mutant enzyme shows lowered unfolding temperature compared with the wild-type enzyme. Faster degradation than wild-type enzyme after incubation at 37°C for 15 h with trypsin or subtilisin at a protease-to-substrate ratio of 1:1 Geobacillus stearothermophilus
G78A turnover number in oxidative deamination of L-Leu is 5.4% of that of the wild-type enzyme. In reductive amination the turnover number is comparable to that of the wild-type enzyme. The Km-value for 2-oxoisohexanoate is 8.8fold higher and the Km-value for NH4+ is 10fold higher than that of the wild-type enzyme. Mutant enzyme shows lowered unfolding temperature compared with the wild-type enzyme. Faster degradation than wild-type enzyme after incubation at 37°C for 15 h with trypsin or subtilisin at a protease-to-substrate ratio of 1:1 Geobacillus stearothermophilus
G79A turnover number in oxidative deamination of L-Leu is 40% of that of the wild-type enzyme. In reductive amination the turnover number is comparable to that of the wild-type enzyme. The Km-value for 2-oxoisohexanoate is 6.4fold higher and the Km-value for NH4+ is 3.9fold higher than that of the wild-type enzyme. Mutant enzyme shows lowered unfolding temperature compared with the wild-type enzyme Geobacillus stearothermophilus

General Stability

General Stability Organism
mutant enzymes G77A and G78A show faster degradation than wild-type enzyme after incubation at 37°C for 15 h with trypsin or subtilisin at a protease-to-substrate ratio of 1:1. Wild-type enzyme and mutant enzyme G79A are degraded at almost the same rate Geobacillus stearothermophilus

Inhibitors

Inhibitors Comment Organism Structure
4-methylpentanoate competitive inhibition of wild-type enzyme and mutant enzyme K80A Geobacillus stearothermophilus
pyridoxal 5'-phosphate
-
 Geobacillus stearothermophilus

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.035
-
 NADH wild-type enzyme Geobacillus stearothermophilus
0.038
-
 NADH mutant enzyme G77A Geobacillus stearothermophilus
0.039
-
 NAD+ mutant enzyme G77A Geobacillus stearothermophilus
0.054
-
 NADH mutant enzyme G78A Geobacillus stearothermophilus
0.055
-
 NADH mutant enzyme G79A Geobacillus stearothermophilus
0.059
-
 NAD+ mutant enzyme G78A Geobacillus stearothermophilus
0.063
-
 NAD+ wild-type enzyme Geobacillus stearothermophilus
0.071
-
 NAD+ mutant enzyme G79A Geobacillus stearothermophilus
0.88
-
 2-Oxoisohexanoate wild-type enzyme Geobacillus stearothermophilus
3.5
-
 L-Leu mutant enzyme G77A Geobacillus stearothermophilus
3.6
-
 L-Leu mutant enzyme G79A Geobacillus stearothermophilus
4.3
-
 L-Leu mutant enzyme G78A Geobacillus stearothermophilus
5.1
-
 L-Leu wild-type enzyme Geobacillus stearothermophilus
5.5
-
 2-Oxoisohexanoate mutant enzyme G77A Geobacillus stearothermophilus
5.6
-
 2-Oxoisohexanoate mutant enzyme G79A Geobacillus stearothermophilus
7.7
-
 2-Oxoisohexanoate mutant enzyme G78A Geobacillus stearothermophilus
75
-
 NH4+ wild-type enzyme Geobacillus stearothermophilus
210
-
 NH4+ mutant enzyme G77A Geobacillus stearothermophilus
290
-
 NH4+ mutant enzyme G79A Geobacillus stearothermophilus
750
-
 NH4+ mutant enzyme G78A Geobacillus stearothermophilus

Organism

Organism UniProt Comment Textmining
Geobacillus stearothermophilus
-
-
-

Purification (Commentary)

Purification (Comment) Organism
-
 Geobacillus stearothermophilus

Reaction

Reaction Comment Organism Reaction ID
L-leucine + H2O + NAD+ = 4-methyl-2-oxopentanoate + NH3 + NADH + H+ ordered bi-ter mechanism, in which NAD+ and L-Leu are bound and NH4+, 2-oxoisohexanoate, and NADH are released in that order Geobacillus stearothermophilus
a

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
L-Leu + H2O + NAD+
-
 Geobacillus stearothermophilus 4-methyl-2-oxopentanoate + NH3 + NADH
-
 r

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
53
-
 unfolding temperature of mutant enzyme G78A Geobacillus stearothermophilus
60
-
 unfolding temperature of mutant enzyme G77A Geobacillus stearothermophilus
76
-
 unfolding temperature of mutant enzyme G79A Geobacillus stearothermophilus
80
-
 unfolding temperature of the wild-type enzyme Geobacillus stearothermophilus

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
additional information
-
 additional information turnover numbers of wild-type enzyme and mutant enzymes Geobacillus stearothermophilus

Cofactor

Cofactor Comment Organism Structure
NAD+
-
 Geobacillus stearothermophilus
NADH
-
 Geobacillus stearothermophilus

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
20
-
 4-methylpentanoate mutant enzyme G77A Geobacillus stearothermophilus
25
-
 4-methylpentanoate wild-type enzyme Geobacillus stearothermophilus
25
-
 4-methylpentanoate mutant enzyme G78A Geobacillus stearothermophilus
30
-
 4-methylpentanoate mutant enzyme G79A Geobacillus stearothermophilus