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Literature summary for 6.3.1.21 extracted from

  • Marolewski, A.; Mattia, K.; Warren, M.; Benkovic, S.
    Formyl phosphate a proposed intermediate in the reaction catalyzed by Escherichia coli PurT GAR transformylase (1997), Biochemistry, 36, 6709-6716 .
    View publication on PubMed
Search for more literature for 6.3.1.21

Cloned(Commentary)

Cloned (Comment) Organism
gene purT, recombinant expression of wild-type and mutant enzymes in TX680 auxotrophic cells Escherichia coli

Protein Variants

Protein Variants Comment Organism
G162I site-directed mutagenesis, the enzyme mutant releases formyl phosphate into solution, reduced activity compared to wild-type enzyme Escherichia coli

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information steady-state kinetics Escherichia coli
0.0101
-
 N1-(5-phospho-beta-D-ribosyl)glycinamide recombinant enzyme, pH 8.0, 25°C Escherichia coli
0.036
-
 ADP recombinant enzyme, pH 8.0, 25°C, reaction with formyl phosphate Escherichia coli
0.045
-
 ATP recombinant enzyme, pH 8.0, 25°C, with formate Escherichia coli
0.077
-
 ATP recombinant enzyme, pH 8.0, 25°C, with acetate Escherichia coli
0.26
-
 ADP recombinant enzyme, pH 8.0, 25°C, reaction with acetyl phosphate Escherichia coli
0.319
-
 formate recombinant enzyme, pH 8.0, 25°C Escherichia coli
0.48
-
 acetyl phosphate recombinant enzyme, pH 8.0, 25°C, reaction with ADP Escherichia coli
0.49
-
 ADP recombinant enzyme, pH 8.0, 25°C, reaction with carbamoyl phosphate Escherichia coli
1.17
-
 Carbamoyl phosphate recombinant enzyme, pH 8.0, 25°C, reaction with ADP Escherichia coli
1.3
-
 formyl phosphate recombinant enzyme, pH 8.0, 25°C, reaction with ADP Escherichia coli

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Escherichia coli

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + formate + N1-(5-phospho-beta-D-ribosyl)glycinamide Escherichia coli
-
 ADP + phosphate + N2-formyl-N1-(5-phospho-beta-D-ribosyl)glycinamide
-
 ?

Organism

Organism UniProt Comment Textmining
Escherichia coli P33221
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and mutant enzymes from TX680 auxotrophic cells by gel filtration, anion exchange chromatography, and dialysis, to over 90% purity Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ADP + acetyl phosphate only half-reaction Escherichia coli ?
-
 ?
ADP + carbamoyl phosphate only half-reaction Escherichia coli ?
-
 ?
ADP + formyl phosphate only half-reaction Escherichia coli ?
-
 ?
ATP + acetate + N1-(5-phospho-beta-D-ribosyl)glycinamide only half-reaction resulting in formation of ADP and acetylphosphate Escherichia coli ?
-
 ?
ATP + formate + N1-(5-phospho-beta-D-ribosyl)glycinamide
-
 Escherichia coli ADP + phosphate + N2-formyl-N1-(5-phospho-beta-D-ribosyl)glycinamide
-
 ?
additional information kinetic studies of the wild-type PurT enzyme demonstrate that formyl phosphate behaves as a chemically and kinetically competent intermediate. The requirement for ATP and glycinamide ribonucleotide (GAR) in these reactions is consistent with previous steady-state kinetic results, which have demonstrated that all substrates must be bound before catalysis. Kinetic analysis and positional isotope exchange studies also support the assignment of formyl phosphate as a plausible intermediate. Forward and reverse half-reactions utilizing the proposed intermediate are measured spectrophotometrically. PurT transformylase is capable of generating acetyl phosphate from ATP and acetate. The first half-reaction leads to production of acyl phosphate, the second half-reaction acylates GAR. Neither acetyl GAR nor carbamyl GAR is produced when the enzyme is incubated with ATP, GAR, and the appropriate acid. Neither the reverse first half-reaction with acetyl phosphate nor carbamyl phosphate requires GAR to be present, in contrast to the half-reactions involving formyl phosphate. No activity with aminoformate Escherichia coli ?
-
-

Synonyms

Synonyms Comment Organism
GAR transformylase
-
 Escherichia coli
glycinamide ribonucleotide transformylase
-
 Escherichia coli
PurT
-
 Escherichia coli

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
 assay at Escherichia coli

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8
-
 assay at Escherichia coli

Cofactor

Cofactor Comment Organism Structure
ATP
-
 Escherichia coli

General Information

General Information Comment Organism
evolution The purT encoded glycinamide ribonucleotide transformylase differs from the previously known purN encoded enzyme in size, sequence, and substrates, and ATP and formate are required as opposed to formyl tetrahydrofolate Escherichia coli
malfunction PurT transformylase mutant G162I catalyzes the production of formyl GAR two orders of magnitude less efficiently than the wild-type enzyme. This reduced rate is apparently sufficient to sustain cell growth under limiting purine conditions Escherichia coli
physiological function the Escherichia coli purT encoded glycinamide ribonucleotide transformylase (GAR transformylase) serves as an alternate enzyme in the production of formyl GAR for use in de novo purine biosynthesis Escherichia coli