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Literature summary for 2.7.1.146 extracted from

  • Castro-Fernandez, V.; Bravo-Moraga, F.; Herrera-Morande, A.; Guixe, V.
    Bifunctional ADP-dependent phosphofructokinase/glucokinase activity in the order Methanococcales - biochemical characterization of the mesophilic enzyme from Methanococcus maripaludis (2014), FEBS J., 281, 2017-2029.
    View publication on PubMed

Activating Compound

Activating Compound Comment Organism Structure
D-glucosamine 10 mM, 40% activation of glucokinase activity Methanococcus maripaludis

Cloned(Commentary)

Cloned (Comment) Organism
expression in Escherichia coli Methanococcus maripaludis

Crystallization (Commentary)

Crystallization (Comment) Organism
molecular modeling of structure. for binding of ADP, residues M347, I431 and L441 create a hydrophobic pocket around the adenine group. R194 makes a hydrogen bond with alpha and beta phosphates, carbonyl and NH groups from V432 peptide bond make a hydrogen bond with the NH2 group of C6 and the N1 atom of adenine Methanococcus maripaludis

Inhibitors

Inhibitors Comment Organism Structure
ADP strong inhibition effect in the reverse glucokinase reaction Methanococcus maripaludis
AMP strong substrate inhibition effect in the reverse glucokinase reaction Methanococcus maripaludis
D-fructose
-
Methanococcus maripaludis
D-fructose 6-phosphate strong inhibition of glucokinase activity, competitive inhibion versus D-glucose as variable substrate Methanococcus maripaludis
D-glucose substrate inhibition above 200 mM. At 10 mM 20% inhibition of phosphofructokinase activity Methanococcus maripaludis
EDTA 50 mM, complete inhibition; complete loss of activity; completely abolishes activity Methanococcus maripaludis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.065
-
D-fructose 6-phosphate pH 6.5, 30°C Methanococcus maripaludis
0.065
-
D-fructose 6-phosphate pH 7.0, 30°C Methanococcus maripaludis
0.1 2 D-glucose 6-phosphate pH 6.5, 30°C Methanococcus maripaludis
0.1 2 D-fructose 1,6-bisphosphate pH 6.5, 30°C Methanococcus maripaludis
0.12
-
D-fructose 1,6-bisphosphate pH 7.0, 30°C Methanococcus maripaludis
8
-
ADP pH 6.5, 30°C Methanococcus maripaludis
8
-
ADP pH 7.0, 30°C Methanococcus maripaludis
40
-
D-glucose pH 6.5, 30°C Methanococcus maripaludis

Metals/Ions

Metals/Ions Comment Organism Structure
Co2+ 2 mM, divalent metal ion required, activation of phosphofructokinase activity with Co2+ is about 70% compared to the activation with Mg2+, activation of glucokinase activity is about 35% compared to the activation with Mg2+ Methanococcus maripaludis
Co2+ among the divalent metal cations tested, the highest activity is observed in the presence of Mg2+, although, in the presence of Co2+, Ni2+ and Mn2+, significant activity is also measured Methanococcus maripaludis
Co2+ 2 mM, about 70% of the activity with Mg2+ Methanococcus maripaludis
Mg2+ highest activity in the presence of Mg2+ Methanococcus maripaludis
Mg2+ 2 mM, divalent metal ion required, highest activity is observed in the presence of Mg2+ Methanococcus maripaludis
Mg2+ among the divalent metal cations tested, the highest activity is observed in the presence of Mg2+, although, in the presence of Co2+, Ni2+ and Mn2+, significant activity is also measured Methanococcus maripaludis
Mn2+ 2 mM, divalent metal ion required, activation of phosphofructokinase activity with Co2+ is about 25% compared to the activation with Mg2+, activation of glucokinase activity is about 20% compared to the activation with Mg2+ Methanococcus maripaludis
Mn2+ among the divalent metal cations tested, the highest activity is observed in the presence of Mg2+, although, in the presence of Co2+, Ni2+ and Mn2+, significant activity is also measured Methanococcus maripaludis
Mn2+ 2 mM, about 25% of the activity with Mg2+ Methanococcus maripaludis
additional information divalent cation required, with highest activity in the presence of Mg2+ Methanococcus maripaludis
Ni2+ 2 mM, divalent metal ion required, activation of phosphofructokinase activity with Co2+ is about 30% compared to the activation with Mg2+, activation of glucokinase activity is less than 10% compared to the activation with Mg2+ Methanococcus maripaludis
Ni2+ among the divalent metal cations tested, the highest activity is observed in the presence of Mg2+, although, in the presence of Co2+, Ni2+ and Mn2+, significant activity is also measured Methanococcus maripaludis

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ADP + D-fructose 6-phosphate Methanococcus maripaludis
-
AMP + D-fructose 1,6-bisphosphate
-
ir
ADP + D-glucose Methanococcus maripaludis the enzyme phosphorylates both D-glucose and D-fructose 6-phosphate. Binding of both substrates to the same active site. At a sugar concentration of 10 mM the acctivity with D-fructose 6-phosphate is about 75% compared to the activity with D-glucose. No activity in presence of ATP. kcat/KM for the phosphorylation of D-fructose 6-phosphate is 440fold higher than the kcat/Km for the phosphorylation of glucose. Analysis of the kcat/Km ratios shows that the glucose dephosphorylation is 2fold more effective than the phosphorylation AMP + D-glucose 6-phosphate
-
r

Organism

Organism UniProt Comment Textmining
Methanococcus maripaludis Q6LXQ3
-
-
Methanococcus maripaludis Q6LXQ3 bifunctional ADP-dependent phosphofructokinase/glucokinase, reactions of EC 2.7.1.147 and EC 2.7.1.146, respectively
-

Purification (Commentary)

Purification (Comment) Organism
-
Methanococcus maripaludis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ADP + D-fructose 6-phosphate
-
Methanococcus maripaludis AMP + D-fructose 1,6-bisphosphate
-
ir
ADP + D-fructose 6-phosphate the bifunctional enzyme is able to phosphorylate D-glucose and beta-D-fructose 6-phosphate. The results of molecular modeling show that both sugars are bound to the enzyme by essentially the same residues except for N203, which establishes an interaction only when the substrate is D-fructose 6-phosphate, and E79, which interacts only with glucose. The enzyme shows higher activity with glucose compared to that obtained with beta-D-fructose 6-phosphate. beta-D-Fructose 6-phosphate shows 75% of the activity measured with glucose. In the presence of ATP, no activity is detected. Phosphatase activity is 67-fold lower than the kinase activity Methanococcus maripaludis AMP + D-fructose 1,6-bisphosphate
-
ir
ADP + D-fructose 6-phosphate the enzyme phosphorylates both D-glucose and D-fructose 6-phosphate. Binding of both substrates to the same active site. At a sugar concentration of 10 mM the activity with D-fructose 6-phosphate is about 75% compared to the activity with D-glucose. No activity in presence of ATP. kcat/KM for the phosphorylation of D-fructose 6-phosphate is 440fold higher than the kcat/Km for the phosphorylation of glucose Methanococcus maripaludis AMP + D-fructose 1,6-bisphosphate
-
ir
ADP + D-fructose 6-phosphate kinetics show hyperbolic behavior Methanococcus maripaludis AMP + D-fructose 1,6-bisphosphate
-
r
ADP + D-glucose the enzyme phosphorylates both D-glucose and D-fructose 6-phosphate. Binding of both substrates to the same active site. At a sugar concentration of 10 mM the acctivity with D-fructose 6-phosphate is about 75% compared to the activity with D-glucose. No activity in presence of ATP. kcat/KM for the phosphorylation of D-fructose 6-phosphate is 440fold higher than the kcat/Km for the phosphorylation of glucose. Analysis of the kcat/Km ratios shows that the glucose dephosphorylation is 2fold more effective than the phosphorylation Methanococcus maripaludis AMP + D-glucose 6-phosphate
-
r
ADP + D-glucose the enzyme phosphorylates both D-glucose and D-fructose 6-phosphate.Binding of both substrates to the same active site. At a sugar concentration of 10 mM the acctivity with D-fructose 6-phosphate is about 75% compared to the activity with D-glucose. No activity in presence of ATP. kcat/KM for the phosphorylation of D-fructose 6-phosphate is 440fold higher than the kcat/Km for the phosphorylation of glucose. Analysis of the kcat/Km ratios shows that the glucose dephosphorylation is 2fold more effective than the phosphorylation Methanococcus maripaludis AMP + D-glucose 6-phosphate
-
r
AMP + D-fructose 1,6-bisphosphate
-
Methanococcus maripaludis ADP + D-fructose 6-phosphate
-
r
CDP + beta-D-fructose 6-phosphate about 15% compared to the activity with ADP Methanococcus maripaludis CMP + D-fructose 1,6-bisphosphate
-
ir
additional information less than 10% activity compared to the activity with D-glucose and ADP: L-rhamnose, D-arabinose, D-lyxose, D-fucose, D-galactose, D-mannose, D-fructose, 2-deoxyglucose, D-glucosamine, D-xylose, maltose, lactose Methanococcus maripaludis ?
-
?
additional information bifunctional ADP-dependent phosphofructokinase/glucokinase, reactions of EC 2.7.1.147 and EC 2.7.1.146, respectively. The rate at which fructose 6-phosphate is phosphorylated is 440fold higher than the glucose phosphorylation rate Methanococcus maripaludis ?
-
?
UDP + D-glucose the enzyme phosphorylates both D-glucose and D-fructose 6-phosphate. Activity with UDP and D-glucose is about 20% compared to the activity with ADP and D-glucose Methanococcus maripaludis UMP + D-glucose 6-phosphate
-
r

Synonyms

Synonyms Comment Organism
bifunctional ADP-dependent phosphofructokinase/glucokinase
-
Methanococcus maripaludis
MmPFK/GK
-
Methanococcus maripaludis
pfkC
-
Methanococcus maripaludis

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.45
-
D-fructose 1,6-bisphosphate pH 7.0, 30°C Methanococcus maripaludis
0.45
-
D-glucose 6-phosphate pH 6.5, 30°C Methanococcus maripaludis
0.45
-
D-fructose 1,6-bisphosphate pH 6.5, 30°C Methanococcus maripaludis
16.5
-
D-fructose 6-phosphate pH 7.0, 30°C Methanococcus maripaludis
16.5
-
D-fructose 6-phosphate pH 6.5, 30°C Methanococcus maripaludis
23
-
D-glucose pH 6.5, 30°C Methanococcus maripaludis
30
-
ADP pH 7.0, 30°C Methanococcus maripaludis
30
-
ADP pH 6.5, 30°C Methanococcus maripaludis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6.5
-
-
Methanococcus maripaludis
6.5
-
phosphofructokinase activity Methanococcus maripaludis
7
-
-
Methanococcus maripaludis
7
-
glucokinase activity Methanococcus maripaludis

pH Range

pH Minimum pH Maximum Comment Organism
5 7 pH 5.0: about 50% of maximal activity, pH 7.0: about 65% of maximal activity Methanococcus maripaludis
5 7 pH 5.0: about 50% of maximal phosphofructokinase activity, pH 7.0: about 65% of maximal phosphofructokinase activity Methanococcus maripaludis
5.5 7.5 pH 5.5: about 50% of maximal phosphofructokinase activity, pH 7.5: about 75% of maximal phosphofructokinase activity Methanococcus maripaludis

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.226
-
D-fructose 6-phosphate pH 7.5, 30°C, glucokinase reaction Methanococcus maripaludis
0.4
-
AMP pH 7.5, 30°C, reverse glucokinase reaction Methanococcus maripaludis
54
-
ADP pH 7.0, 30°C Methanococcus maripaludis
54
-
ADP pH 6.5, 30°C Methanococcus maripaludis
895
-
D-glucose pH 7.5, 30°C Methanococcus maripaludis

IC50 Value

IC50 Value IC50 Value Maximum Comment Organism Inhibitor Structure
0.08
-
pH 7.5, 30°C, reverse glucokinase reaction Methanococcus maripaludis ADP
0.22
-
pH 6.5, 30°C Methanococcus maripaludis D-fructose 6-phosphate
0.22
-
pH 7.5, 30°C, glucokinase reaction Methanococcus maripaludis D-fructose 6-phosphate
40
-
pH 6.5, 30°C Methanococcus maripaludis D-glucose
40
-
pH 7.5, 30°C, phosphopfructokinase reaction Methanococcus maripaludis D-fructose

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.57
-
D-glucose pH 6.5, 30°C Methanococcus maripaludis
1.09
-
D-glucose 6-phosphate pH 6.5, 30°C Methanococcus maripaludis
3
-
ADP pH 6.5, 30°C Methanococcus maripaludis
3.75
-
ADP pH 7.0, 30°C Methanococcus maripaludis
3.75
-
D-fructose 1,6-bisphosphate pH 7.0, 30°C Methanococcus maripaludis
3.75
-
D-fructose 1,6-bisphosphate pH 6.5, 30°C Methanococcus maripaludis
253
-
D-fructose 6-phosphate pH 7.0, 30°C Methanococcus maripaludis
253
-
D-fructose 6-phosphate pH 6.5, 30°C Methanococcus maripaludis