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Literature summary for 3.1.3.89 extracted from

  • Zimmerman, M.D.; Proudfoot, M.; Yakunin, A.; Minor, W.
    Structural insight into the mechanism of substrate specificity and catalytic activity of an HD-domain phosphohydrolase: the 5-deoxyribonucleotidase YfbR from Escherichia coli (2008), J. Mol. Biol., 378, 215-226.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
-
Escherichia coli

Crystallization (Commentary)

Crystallization (Comment) Organism
free state and two complexes with natural substrates, to 2.1 A resolution. The free-state structure contains a large cavity accommodating the metal-coordinating HD motif of residues H33, H68, D69, and D137 and other conserved residues, i.e. R18, E72, and D77. These residues are important for activity. Residue R18 stabilizes the phosphate on the Co2+, and residue D77 forms a strong hydrogen bond critical for binding the ribose. The indole side chain of W19 is located close to the 2'-carbon atom of the deoxyribose moiety and is proposed to act as the selectivity switch for deoxyribonucleotide. The nucleotide bases of both dAMP and TMP make no specific hydrogen bonds with the protein, explaining the lack of nucleotide base selectivity. The E72A substrate complex structures also suggest a plausible single-step nucleophilic substitution mechanism Escherichia coli

Protein Variants

Protein Variants Comment Organism
D137A complete loss of activity Escherichia coli
D69A almost complete loss of activity Escherichia coli
D77A complete loss of activity Escherichia coli
E122A almost complete loss of activity Escherichia coli
E72A catalytically inactive. Structures complexed with Co2+ and either TMP or dAMP disclose the unusual binding mode of deoxyribonucleotides in the active site Escherichia coli
H33A about 20% of wild-type activity Escherichia coli
H68A almost complete loss of activity Escherichia coli
R18A almost complete loss of activity Escherichia coli
V30A about 80% of wild-type activity Escherichia coli
V37A about 85% of wild-type activity Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.016
-
5'-dAMP mutant V37A, pH 7.5, 37°C Escherichia coli
0.017
-
5'-dAMP wild-type, pH 7.5, 37°C Escherichia coli
0.034
-
5'-dAMP mutant E122A, pH 7.5, 37°C Escherichia coli
0.042
-
5'-dAMP mutant V30A, pH 7.5, 37°C Escherichia coli

Organism

Organism UniProt Comment Textmining
Escherichia coli P76491
-
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
5'-dAMP + H2O
-
Escherichia coli deoxyadenosine + phosphate
-
?

Synonyms

Synonyms Comment Organism
YfbR
-
Escherichia coli

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.06
-
5'-dAMP mutant E122A, pH 7.5, 37°C Escherichia coli
0.08
-
5'-dAMP mutant V30A, pH 7.5, 37°C Escherichia coli
0.13
-
5'-dAMP mutant V37A, pH 7.5, 37°C Escherichia coli
0.14
-
5'-dAMP wild-type, pH 7.5, 37°C Escherichia coli

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
1.72
-
5'-dAMP mutant E122A, pH 7.5, 37°C Escherichia coli
1.98
-
5'-dAMP mutant V30A, pH 7.5, 37°C Escherichia coli
8.1
-
5'-dAMP wild-type, pH 7.5, 37°C Escherichia coli
8.41
-
5'-dAMP mutant V37A, pH 7.5, 37°C Escherichia coli