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Literature summary for 3.6.1.6 extracted from

  • Imae, K.; Saito, Y.; Kizaki, H.; Ryuno, H.; Mao, H.; Miyashita, A.; Suzuki, Y.; Sekimizu, K.; Kaito, C.
    Novel nucleoside diphosphatase contributes to Staphylococcus aureus virulence (2016), J. Biol. Chem., 291, 18608-18619 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
gene SA1684, quantitative reverse transcription-PCR analysis, recombinant expression of C-terminally His-tagged SA1684 protein in Escherichia coli strain BL21(DE3)-RIPL Staphylococcus aureus

Protein Variants

Protein Variants Comment Organism
D106A site-directed mutagenesis, the mutant shows highly reduced nucleoside diphosphatase activity Staphylococcus aureus
D123A/E124A site-directed mutagenesis, the mutant shows highly reduced nucleoside diphosphatase activity Staphylococcus aureus
additional information introduction of the wild-type SA1684 gene restores the hemolysin production of the SA1684-deletion mutant, whereas none of the alanine-substituted SA1684 mutant genes restores the hemolysin production. Construction of a DELTASA1684 deletion mutant, which exhibits drastically decreased virulence in a silkworm (Bombyx mori) infection model, in which the LD50 against silkworm larvae is more than 10times that of the parent strain. The SA1684-deletion mutant also exhibits decreased exotoxin production and colony-spreading ability. Introduction of wild-type SA1684 to the SA1684-deletion mutant restores the hemolysin production, nuclease production, and the colony-spreading activity Staphylococcus aureus
Y88A site-directed mutagenesis, the mutant shows highly reduced nucleoside diphosphatase activity Staphylococcus aureus

Inhibitors

Inhibitors Comment Organism Structure
GDP substrate inhibition at high concentration Staphylococcus aureus
GDPbetaS a non-hydrolyzable analogue of GDP Staphylococcus aureus
GTPgammaS a non-hydrolyzable analogue of GTP Staphylococcus aureus
UDP substrate inhibition at high concentration Staphylococcus aureus

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information Michaelis-Menten kinetics Staphylococcus aureus
0.0963
-
ADP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
0.102
-
GDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
0.203
-
TDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
0.221
-
UDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
0.303
-
CDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus

Metals/Ions

Metals/Ions Comment Organism Structure
Co2+ dependent on Staphylococcus aureus
Mn2+ dependent on, activates at 0.5-12.5 mM Staphylococcus aureus
additional information no or very poor activation by Cu2+, Ni2+, Zn2+, Mg2+, and Ca2+ at 0.5 mM Staphylococcus aureus

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
GDP + H2O Staphylococcus aureus
-
GMP + phosphate
-
?
GDP + H2O Staphylococcus aureus N315
-
GMP + phosphate
-
?
UDP + H2O Staphylococcus aureus
-
UMP + phosphate
-
?

Organism

Organism UniProt Comment Textmining
Staphylococcus aureus Q7A4T2
-
-
Staphylococcus aureus N315 Q7A4T2
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant C-terminally His-tagged SA1684 protein from Escherichia coli strain BL21(DE3)-RIPL by nickel affinity chromatography to over 90% purity Staphylococcus aureus

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ADP + H2O high activity Staphylococcus aureus AMP + phosphate
-
?
ADP + H2O high activity Staphylococcus aureus N315 AMP + phosphate
-
?
CDP + H2O moderate activity Staphylococcus aureus CMP + phosphate
-
?
dADP + H2O low activity Staphylococcus aureus dAMP + phosphate
-
?
dCDP + H2O low activity Staphylococcus aureus dCMP + phosphate
-
?
dGDP + H2O low activity Staphylococcus aureus dGMP + phosphate
-
?
GDP + H2O
-
Staphylococcus aureus GMP + phosphate
-
?
GDP + H2O moderate activity Staphylococcus aureus GMP + phosphate
-
?
GDP + H2O
-
Staphylococcus aureus N315 GMP + phosphate
-
?
GDP + H2O moderate activity Staphylococcus aureus N315 GMP + phosphate
-
?
IDP + H2O low activity Staphylococcus aureus IMP + phosphate
-
?
additional information purified SA1684 protein has Mn2+- or Co2+-dependent hydrolyzing activity against nucleoside diphosphates. The enzyme shows or poor activity with NTPS or NMPs, substrate specificity, overview Staphylococcus aureus ?
-
-
additional information purified SA1684 protein has Mn2+- or Co2+-dependent hydrolyzing activity against nucleoside diphosphates. The enzyme shows or poor activity with NTPS or NMPs, substrate specificity, overview Staphylococcus aureus N315 ?
-
-
TDP + H2O moderate activity Staphylococcus aureus TMP + phosphate
-
?
TDP + H2O moderate activity Staphylococcus aureus N315 TMP + phosphate
-
?
UDP + H2O
-
Staphylococcus aureus UMP + phosphate
-
?
UDP + H2O best substrate Staphylococcus aureus UMP + phosphate
-
?

Subunits

Subunits Comment Organism
? x * 28000, recombinant His-tagged enzyme, SDS-PAGE Staphylococcus aureus

Synonyms

Synonyms Comment Organism
Nucleoside diphosphatase
-
Staphylococcus aureus
SA1684
-
Staphylococcus aureus
SA1684 protein
-
Staphylococcus aureus

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
26
-
assay at Staphylococcus aureus

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.357
-
GDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
0.415
-
ADP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
0.68
-
TDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
0.763
-
CDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
1.388
-
UDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
assay at Staphylococcus aureus

General Information

General Information Comment Organism
evolution the SA1684 gene product carries theDUF402domain, which is found in RNA-binding proteins, and has amino acid sequence similarity with a nucleoside diphosphatase, Streptomyces coelicolor SC4828 protein Staphylococcus aureus
malfunction the SA1684-deletion mutant exhibits drastically decreased virulence in a silkworm (Bombyx mori) infection model, in which the LD50 against silkworm larvae is more than 10times that of the parent strain. The SA1684-deletion mutant also exhibits decreased exotoxin production and colony-spreading ability. Introduction of wild-type SA1684 to the SA1684-deletion mutant restores the hemolysin production, nuclease production, and the colony-spreading activity Staphylococcus aureus
additional information the amino acid residues Tyr88, Asp106, and Asp123/Glu124 of SA1684 protein are required for NDP hydrolysis and Staphylococcus aureus virulence Staphylococcus aureus
physiological function RNA sequence analysis reveals thatSA1684 is required for the expression of the virulence regulatory genes agr, sarZ, and sarX, as well as metabolic genes involved in glycolysis and fermentation pathways. These findings suggest that the nucleoside diphosphatase SA1684 links metabolic pathways and virulence gene expression and plays an important role in Staphylococcus aureus virulence in silkworms. Introduction of wild-type SA1684 to the SA1684-deletion mutant restores the hemolysin production, nuclease production, and the colony-spreading activity. Enzyme SA1684 is required for hemolysin production, nuclease production, and colony spreading Staphylococcus aureus

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.518
-
CDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
3.35
-
TDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
3.5
-
GDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
4.31
-
ADP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus
6.281
-
UDP recombinant enzyme, pH 7.5, 26°C Staphylococcus aureus