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Literature summary for 4.1.3.3 extracted from

  • Campeotto, I.; Bolt, A.H.; Harman, T.A.; Dennis, C.; Trinh, C.H.; Phillips, S.E.; Nelson, A.; Pearson, A.R.; Berry, A.
    Structural insights into substrate specificity in variants of N-acetylneuraminic acid lyase produced by directed evolution (2010), J. Mol. Biol., 404, 56-69.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
His-tagged version expressed in Escherichia coli BL21 Escherichia coli

Crystallization (Commentary)

Crystallization (Comment) Organism
sitting drop diffusion method, PEG 3350 Escherichia coli

Protein Variants

Protein Variants Comment Organism
E192A strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192C strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192D strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192F strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192G strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192H strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192I strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192K strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192L strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192M strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192N strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192P strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192Q strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192R strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192S strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192T strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192V strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192W strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli
E192Y strong interaction of Glu192 (wild type) with hydroxyl groups at C8 and C9 of N-acetylneuraminic acid Escherichia coli

Inhibitors

Inhibitors Comment Organism Structure
(2R,3R)-2,3,4-trihydroxy-N,N-dipropylbutanamide competitive inhibitor Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.2
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192M mutant protein, pH 7.5, 25°C Escherichia coli
0.2
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192P mutant protein, pH 7.5, 25°C Escherichia coli
0.3
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192F mutant protein, pH 7.5, 25°C Escherichia coli
0.4
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192N mutant protein, pH 7.7.5, 25°C Escherichia coli
0.4
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192V mutant protein, pH 7.5, 25°C Escherichia coli
0.5
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192H mutant protein, pH 7.5, 25°C Escherichia coli
0.6
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192S mutant protein, pH 7.5, 25°C Escherichia coli
0.7
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192Q mutant protein, pH 7.5, 25°C Escherichia coli
1.1
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192D mutant protein, pH 7.5, 25°C Escherichia coli
3.5
-
N-acetylneuraminic acid E192Q mutant protein, pH 7.5, 25°C Escherichia coli
4.4
-
N-acetylneuraminic acid wild type (E192), pH 7.5, 25°C Escherichia coli
4.6
-
N-acetylneuraminic acid E192D mutant protein, pH 7.5, 25°C Escherichia coli
5
-
N-acetylneuraminic acid E192M mutant protein, pH 7.5, 25°C Escherichia coli
5.9
-
N-acetylneuraminic acid E192F mutant protein, pH 7.5, 25°C Escherichia coli
6
-
N-acetylneuraminic acid E192P mutant protein, pH 7.5, 25°C Escherichia coli
6
-
N-acetylneuraminic acid E192S mutant protein, pH 7.5, 25°C Escherichia coli
6.2
-
N-acetylneuraminic acid E192H mutant protein, pH 7.5, 25°C Escherichia coli
11
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid wild type (E192), pH 7.5, 25°C Escherichia coli
34
-
N-acetylneuraminic acid E192V mutant protein, pH 7.5, 25°C Escherichia coli
38
-
N-acetylneuraminic acid E192N mutant protein, pH 7.7.5, 25°C Escherichia coli

Organism

Organism UniProt Comment Textmining
Escherichia coli
-
-
-

Purification (Commentary)

Purification (Comment) Organism
immobilized metal ion affinity chromatography (Ni2+), gel filtration Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid
-
Escherichia coli pyruvate + (2R,3S)-2,3-dihydroxy-4-oxo-N,N-dipropylbutanamide
-
r
N-acetylmannosamine + pyruvate
-
Escherichia coli N-acetylneuraminic acid
-
r
N-acetylneuraminic acid
-
Escherichia coli N-acetylmannosamine + pyruvate
-
r

Synonyms

Synonyms Comment Organism
N-Acetylneuraminic acid aldolase
-
Escherichia coli
N-Acetylneuraminic acid lyase
-
Escherichia coli
Sialic acid aldolase
-
Escherichia coli

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.1
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192D mutant protein, pH 7.5, 25°C Escherichia coli
0.42
-
N-acetylneuraminic acid E192P mutant protein, pH 7.5, 25°C Escherichia coli
0.43
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192S mutant protein, pH 7.5, 25°C Escherichia coli
0.63
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192H mutant protein, pH 7.5, 25°C Escherichia coli
0.82
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192P mutant protein, pH 7.5, 25°C Escherichia coli
0.83
-
N-acetylneuraminic acid E192S mutant protein, pH 7.5, 25°C Escherichia coli
1.17
-
N-acetylneuraminic acid E192M mutant protein, pH 7.5, 25°C Escherichia coli
1.17
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192M mutant protein, pH 7.5, 25°C Escherichia coli
1.22
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid wild type (E192), pH 7.5, 25°C Escherichia coli
1.47
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192F mutant protein, pH 7.5, 25°C Escherichia coli
1.58
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192V mutant protein, pH 7.5, 25°C Escherichia coli
2
-
N-acetylneuraminic acid E192D mutant protein, pH 7.5, 25°C Escherichia coli
2
-
N-acetylneuraminic acid E192H mutant protein, pH 7.5, 25°C Escherichia coli
2
-
N-acetylneuraminic acid E192V mutant protein, pH 7.5, 25°C Escherichia coli
2.17
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192N mutant protein, pH 7.7.5, 25°C Escherichia coli
2.67
-
N-acetylneuraminic acid E192F mutant protein, pH 7.5, 25°C Escherichia coli
2.83
-
N-acetylneuraminic acid E192N mutant protein, pH 7.7.5, 25°C Escherichia coli
2.83
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192Q mutant protein, pH 7.5, 25°C Escherichia coli
4.33
-
N-acetylneuraminic acid wild type (E192), pH 7.5, 25°C Escherichia coli
5
-
N-acetylneuraminic acid E192Q mutant protein, pH 7.5, 25°C Escherichia coli

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
1.3
-
(2R,3R)-2,3,4-trihydroxy-N,N-dipropylbutanamide E192N mutant protein , pH 7.5, 25°C Escherichia coli

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.06
-
N-acetylneuraminic acid E192V mutant protein, pH 7.5, 25°C Escherichia coli
0.07
-
N-acetylneuraminic acid E192N mutant protein, pH 7.7.5, 25°C Escherichia coli
0.07
-
N-acetylneuraminic acid E192P mutant protein, pH 7.5, 25°C Escherichia coli
0.08
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192D mutant protein, pH 7.5, 25°C Escherichia coli
0.12
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid wild type (E192), pH 7.5, 25°C Escherichia coli
0.13
-
N-acetylneuraminic acid E192S mutant protein, pH 7.5, 25°C Escherichia coli
0.23
-
N-acetylneuraminic acid E192M mutant protein, pH 7.5, 25°C Escherichia coli
0.32
-
N-acetylneuraminic acid E192H mutant protein, pH 7.5, 25°C Escherichia coli
0.43
-
N-acetylneuraminic acid E192D mutant protein, pH 7.5, 25°C Escherichia coli
0.45
-
N-acetylneuraminic acid E192F mutant protein, pH 7.5, 25°C Escherichia coli
0.7
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192S mutant protein, pH 7.5, 25°C Escherichia coli
0.98
-
N-acetylneuraminic acid wild type (E192), pH 7.5, 25°C Escherichia coli
1.33
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192H mutant protein, pH 7.5, 25°C Escherichia coli
1.42
-
N-acetylneuraminic acid E192Q mutant protein, pH 7.5, 25°C Escherichia coli
4.17
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192Q mutant protein, pH 7.5, 25°C Escherichia coli
4.33
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192P mutant protein, pH 7.5, 25°C Escherichia coli
4.5
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192V mutant protein, pH 7.5, 25°C Escherichia coli
5.17
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192F mutant protein, pH 7.5, 25°C Escherichia coli
5.33
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192M mutant protein, pH 7.5, 25°C Escherichia coli
5.67
-
(5R,6R)-7-(dipropylamino)-4,5,6-trihydroxy-2,7-dioxoheptanoic acid E192N mutant protein, pH 7.7.5, 25°C Escherichia coli