Literature summary for 2.7.4.22 extracted from

Jensen, K.S.; Johansson, E.; Jensen, K.F.
Structural and enzymatic investigation of the Sulfolobus solfataricus uridylate kinase shows competitive UTP inhibition and the lack of GTP stimulation (2007), Biochemistry, 46, 2745-2757.

Activating Compound

Activating Compound	Comment	Organism	Structure
GTP	no influence, does not bind the enzyme	Saccharolobus solfataricus
additional information	GTP no influence, GTP does not bind, unlike most bacteria, is not activated by GTP, UTP acts as a competitive inhibitor against both substrates, high concentration of UMP abolishes the inhibition of UTP at low ATP concentrations, indicating that UTP binds to the acceptor site (UMP site)	Saccharolobus solfataricus

Cloned(Commentary)

Cloned (Comment)	Organism
expression in Escherichia coli	Saccharolobus solfataricus
plasmid vector pUHE23-2, expression in Escherichia coli strain NF1830	Saccharolobus solfataricus

Crystallization (Commentary)

Crystallization (Comment)	Organism
ternary complex with UMP and the nonhydrolyzable ATP analogue alpha,beta-methylene-ATP (SsUMPK-UMP-AMPPCP), resolution 2.1 A, a complex with UMP (SsUMPK-UMP), resolution 2.2 A, a complex with UTP (SsUMPK-UTP), resolution 2.8 A, hanging drop vapor diffusion, protein solution (2UL) in 10 mM TRIS/Cl pH 7.6 with 4.6 mg/ml SSUMPK and 2 mM UMP and 5 mM MgCl2 mixed with 2 UL mother solution (0.65 M sodium acetate, 100 mM CdCl2, 0.1 M HEPES), pH 7.5	Saccharolobus solfataricus
with ATP and UMP, with UMP, with UTP	Saccharolobus solfataricus

Crystallization (Comment)

Organism

ternary complex with UMP and the nonhydrolyzable ATP analogue alpha,beta-methylene-ATP (SsUMPK-UMP-AMPPCP), resolution 2.1 A, a complex with UMP (SsUMPK-UMP), resolution 2.2 A, a complex with UTP (SsUMPK-UTP), resolution 2.8 A, hanging drop vapor diffusion, protein solution (2UL) in 10 mM TRIS/Cl pH 7.6 with 4.6 mg/ml SSUMPK and 2 mM UMP and 5 mM MgCl2 mixed with 2 UL mother solution (0.65 M sodium acetate, 100 mM CdCl2, 0.1 M HEPES), pH 7.5

Saccharolobus solfataricus

with ATP and UMP, with UMP, with UTP

Saccharolobus solfataricus

Inhibitors

Inhibitors	Comment	Organism
ATP	-	Saccharolobus solfataricus
CTP	no influence	Saccharolobus solfataricus
additional information	CTP is not inhibitory	Saccharolobus solfataricus
UMP	-	Saccharolobus solfataricus
UTP	binds the enzyme, competitive inhibitor for both substrates; feedback inhibition, competitive inhibitor against both substrates	Saccharolobus solfataricus

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
0.014	-	UMP	in presence of 0.5 mM ATP at 60°C	Saccharolobus solfataricus
0.014	-	UMP	radioactive assay, 500 microM ATP and 100 microM UMP, at 60°C, the more commonly used photometric assay using auxiliary enzymes is incompatible with high temperature	Saccharolobus solfataricus
0.081	-	ATP	in presence of 0.1 mM UMP, at 60°C	Saccharolobus solfataricus
0.081	-	ATP	radioactive assay, 500 microM ATP and 100 microM UMP, at 60°C, the more commonly used photometric assay using auxiliary enzymes is incompatible with high temperature	Saccharolobus solfataricus

Metals/Ions

Metals/Ions	Comment	Organism
Cd2+	precipitates in phosphate buffer	Saccharolobus solfataricus
Cd2+	causes severe precipitation problems in the phosphate buffer	Saccharolobus solfataricus
Co2+	-	Saccharolobus solfataricus
Mg2+	-	Saccharolobus solfataricus
Mn2+	-	Saccharolobus solfataricus
additional information	only one metal ion in the active site. This ion is coordinated with phosphate groups of both ATP and UMP	Saccharolobus solfataricus

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
140000	-	gel filtration	Saccharolobus solfataricus
140000	-	gel flitration	Saccharolobus solfataricus

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.	Reac.
ATP + UMP	Saccharolobus solfataricus	high substrate specificity toward UMP and ATP	ADP + UDP	-	?

Organism

Organism	UniProt	Comment	Textmining
Saccharolobus solfataricus	-	-	-
Saccharolobus solfataricus	Q97ZE2	strain P2, plasmid vector pUHE23-2, expression in Escherichia coli strain NF1830	-

Purification (Commentary)

Purification (Comment)	Organism
heating at 70°C, ammonium sulfate precipitation, and column chromatography on Dyematrex Gel Red A and the anion exchange materials DE52 and Q6. Minor amounts (around 2%) of a smaller 15 kDa protein, presumably an SsUMPK degradation product, co-purified with SsUMP.	Saccharolobus solfataricus
of the recombinant protein	Saccharolobus solfataricus

Reaction

Reaction	Comment	Organism	Reaction ID
ATP + UMP = ADP + UDP	a sequential bi-bi reaction mechanism and, as a nucleophilic substitution reaction while both substrates are bound to the enzyme	Saccharolobus solfataricus	a

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
ATP + dUMP	3.7% of the activity detected with UMP	Saccharolobus solfataricus	ADP + dUDP	-	?
ATP + UMP	low activity with dUMP as substrate, no activity with CMP, dTMP, AMP, IMP, XMP, GMP, low activity with dATP, GTP, CTP and UTP as substrate	Saccharolobus solfataricus	ADP + UDP	-	r
ATP + UMP	high substrate specificity toward UMP and ATP	Saccharolobus solfataricus	ADP + UDP	-	?
additional information	no activity with CMP, dTMP, or the purine nucleotides (AMP, IMP, XMP, and GMP), ATP is by far the most effective phosphoryl donor, although a very weak activity (less than 1% of ATP activity) is seen with dATP, GTP, CTP, and UTP	Saccharolobus solfataricus	?	-	?

Subunits

Subunits	Comment	Organism
hexamer	gel flitration	Saccharolobus solfataricus
hexamer	gel filtration and sedimentation experiments	Saccharolobus solfataricus

Synonyms

Synonyms	Comment	Organism
pyrH	-	Saccharolobus solfataricus
SsUMPK	-	Saccharolobus solfataricus
UMP kinase	-	Saccharolobus solfataricus
UMPK	-	Saccharolobus solfataricus
uridine monophosphate kinase	-	Saccharolobus solfataricus
uridylate kinase	-	Saccharolobus solfataricus

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism	Structure
19	-	UMP	radioactive assay, at 60°C, the more commonly used photometric assay using auxiliary enzymes is incompatible with high temperature	Saccharolobus solfataricus

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7	-	pH variation influences only turnover rate, at 60°C	Saccharolobus solfataricus
7	-	variation of pH only influences the turnover number, at 60°C, 50 mM succinate/phosphate buffer, phosphate buffer, glycine buffer, assays are performed with 100 microM UMP, 500 microM ATP, and 10 mM MgCl2	Saccharolobus solfataricus

pH Range

pH Minimum	pH Maximum	Comment	Organism
additional information	-	variation of pH only influences the turnover number	Saccharolobus solfataricus
4.5	10.7	various buffers, at 60°C	Saccharolobus solfataricus
6	9	-	Saccharolobus solfataricus

Ki Value [mM]

Ki Value [mM]	Ki Value maximum [mM]	Inhibitor	Comment	Organism	Structure
0.014	-	UMP	in presence of 0.5 mM ATP	Saccharolobus solfataricus
0.097	-	ATP	in presence of 0.1 mM UMP	Saccharolobus solfataricus