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Literature summary for 2.7.1.40 extracted from

  • Zoraghi, R.; See, R.H.; Gong, H.; Lian, T.; Swayze, R.; Finlay, B.B.; Brunham, R.C.; McMaster, W.R.; Reiner, N.E.
    Functional analysis, overexpression, and kinetic characterization of pyruvate kinase from methicillin-resistant Staphylococcus aureus (2010), Biochemistry, 49, 7733-7747.
    View publication on PubMed

Activating Compound

Activating Compound Comment Organism Structure
AMP allosteric activator Staphylococcus aureus
ATP slight activation of the full-length enzyme, not the C-terminally truncated enzyme Staphylococcus aureus
D-ribose 5-phosphate allosteric activator Staphylococcus aureus
additional information no activation by fructose 1,6-bisphosphate Staphylococcus aureus

Cloned(Commentary)

Cloned (Comment) Organism
gene pyk, phylogenetic tree, expression of His-tagged full-length and C-terminally truncated enzymes in Escherichia coli strain BL-21(DE3) Staphylococcus aureus
gene pykA, phylogenetic tree, expression of His-tagged isozyme PK2 in Escherichia coli strain BL21(DE3) Escherichia coli
gene pykf, phylogenetic tree, expression of His-tagged isozyme PK1 in Escherichia coli strain BL21(DE3) Escherichia coli

Protein Variants

Protein Variants Comment Organism
additional information construction of a C-terminally truncated mutant PKCT with a stop after residue 390. The catalytic activities of PKCT toward both phophoenolpyruvate and ADP are profoundly decreased compared to those of wild-type enzyme Staphylococcus aureus

Inhibitors

Inhibitors Comment Organism Structure
ATP inhibition of full-length enzyme at concentration above 2.5 mM Staphylococcus aureus
D-fructose 1,6-bisphosphate inhibition of full-length enzyme at 10 mM Staphylococcus aureus
phosphate
-
Staphylococcus aureus

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information steady-state kinetics of recombinant wild-type and truncated PKin the absence or presence of 1 mM AMP as a function of phosphoenolpyruvate Staphylococcus aureus

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Staphylococcus aureus
Mg2+ required Escherichia coli

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
53100
-
4 * 65100, about, full-length enzyme, sequence calculation, 4 * 53100, about, C-terminally truncated enzyme mutant, sequence calculation Staphylococcus aureus
65100
-
4 * 65100, about, full-length enzyme, sequence calculation, 4 * 53100, about, C-terminally truncated enzyme mutant, sequence calculation Staphylococcus aureus
215000
-
His-tagged C-terminally truncated enzyme mutant, gel filtration Staphylococcus aureus
250000
-
His-tagged full-length enzyme, gel filtration Staphylococcus aureus

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + pyruvate Staphylococcus aureus
-
ADP + phosphoenolpyruvate
-
?
ATP + pyruvate Escherichia coli
-
ADP + phosphoenolpyruvate
-
?
ATP + pyruvate Staphylococcus aureus MRSA252
-
ADP + phosphoenolpyruvate
-
?

Organism

Organism UniProt Comment Textmining
Escherichia coli A0A0H3JER5 isozyme PK2; gene pykA
-
Escherichia coli P0AD62 isozyme PK1; gene pykF
-
Staphylococcus aureus Q6GG09 a methicillin-resistant strain, gene pyk
-
Staphylococcus aureus MRSA252 Q6GG09 a methicillin-resistant strain, gene pyk
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged full-length and C-terminally truncated enzymes from Escherichia coli strain BL-21(DE3) by nickel affiniy chromatography Staphylococcus aureus
recombinant His-tagged isozyme PK1 from Escherichia coli strain BL21(DE3) by nickel affiniy chromatography Escherichia coli
recombinant His-tagged isozyme PK2 from Escherichia coli strain BL21(DE3) by nickel affiniy chromatography Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + pyruvate
-
Staphylococcus aureus ADP + phosphoenolpyruvate
-
?
ATP + pyruvate
-
Escherichia coli ADP + phosphoenolpyruvate
-
?
ATP + pyruvate
-
Staphylococcus aureus MRSA252 ADP + phosphoenolpyruvate
-
?

Subunits

Subunits Comment Organism
homotetramer
-
Escherichia coli
homotetramer 4 * 65100, about, full-length enzyme, sequence calculation, 4 * 53100, about, C-terminally truncated enzyme mutant, sequence calculation Staphylococcus aureus
More the C-terminal domain is not required for the tetramerization of the enzyme, homotetramerization also occurs in a truncated enzyme lacking the domain Staphylococcus aureus

Synonyms

Synonyms Comment Organism
MRSA PK
-
Staphylococcus aureus

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
assay at Staphylococcus aureus
37
-
assay at Escherichia coli

pH Optimum

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

Cofactor

Cofactor Comment Organism Structure
ATP
-
Staphylococcus aureus
ATP
-
Escherichia coli

General Information

General Information Comment Organism
evolution significant evolutionary distance existing between the type I and type II isoenzymes in Gram-negative bacteria Staphylococcus aureus
malfunction the catalytic activities of the C-terminally truncated mutant toward both phophoenolpyruvate and ADP are profoundly decreased compared to those of wild-type enzyme Staphylococcus aureus
additional information the C-terminally truncated enzyme exhibits high affinity toward both phophoenolpyruvate and ADP and exhibits hyperbolic kinetics toward phophoenolpyruvate in the presence of activators AMP and ribose 5-phosphate consistent with kinetic properties of full-length enzyme Staphylococcus aureus
physiological function the C-terminal domain is not required for substrate binding or allosteric regulation observed in the holoenzyme, the kinetic efficiency of the truncated enzyme is decreased by 24 and 16fold, in ligand-free state, toward phophoenolpyruvate and ADP, respectively, but is restored by 3fold in AMP-bound state. The C-terminal domain (Gly473-Leu585) plays a substantial role in enzyme activity and comformational stability, and the C-terminal domain is involved in maintaining the specificity of allosteric regulation Staphylococcus aureus