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Literature summary for 3.5.4.25 extracted from

  • Spoonamore, J.E.; Bandarian, V.
    Understanding functional divergence in proteins by studying intragenomic homologues (2008), Biochemistry, 47, 2592-2600.
    View publication on PubMed
Search for more literature for 3.5.4.25

Cloned(Commentary)

Cloned (Comment) Organism
overexpression and purification of wild type and site-directed variants of GCH II Streptomyces coelicolor

Protein Variants

Protein Variants Comment Organism
D127A about 10fold less efficient than the wild-type protein Streptomyces coelicolor
G209D about 10fold less efficient than the wild-type protein Streptomyces coelicolor
M120A generated by site-directed mutagenesis Streptomyces coelicolor
M120C generated by site-directed mutagenesis Streptomyces coelicolor
M120D generated by site-directed mutagenesis Streptomyces coelicolor
M120E generated by site-directed mutagenesis Streptomyces coelicolor
M120F generated by site-directed mutagenesis Streptomyces coelicolor
M120G generated by site-directed mutagenesis Streptomyces coelicolor
M120H generated by site-directed mutagenesis Streptomyces coelicolor
M120I generated by site-directed mutagenesis Streptomyces coelicolor
M120K generated by site-directed mutagenesis Streptomyces coelicolor
M120N generated by site-directed mutagenesis Streptomyces coelicolor
M120Q generated by site-directed mutagenesis Streptomyces coelicolor
M120S generated by site-directed mutagenesis Streptomyces coelicolor
M120T generated by site-directed mutagenesis Streptomyces coelicolor
M120W generated by site-directed mutagenesis Streptomyces coelicolor
M120Y generated by site-directed mutagenesis Streptomyces coelicolor
R83H mutant substantially impaired in overall activity Streptomyces coelicolor
Y123F inactive Streptomyces coelicolor
Y123M about 10fold less efficient than the wild-type protein, produces exclusively 2-amino-5-formylamino-6-ribosylamino-4(3H)-pyrimidinone 5'-phosphate Streptomyces coelicolor
Y326F generated by site-directed mutagenesis, catalytically inactive variant Streptomyces coelicolor
Y326M generated by site-directed mutagenesis, catalytically inactive variant Streptomyces coelicolor

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.008
-
 GTP M120Y variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.008
-
 GTP variant M120Y, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.0089
-
 GTP M120K variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.011
-
 GTP M120Q variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.015
-
 GTP M120I variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.019
-
 GTP M120F variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.019
-
 GTP variant M120F, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.024
-
 GTP wild type, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.033
-
 GTP M120S variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.048
-
 GTP variant Y123M, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.05
-
 GTP M120H variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.056
-
 GTP wild type, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.059
-
 GTP M120A variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.063
-
 GTP wild type, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.067
-
 GTP M120C variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.067
-
 GTP M120N variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.085
-
 GTP variant D127A, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.092
-
 GTP variant R123M, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.15
-
 GTP M120T variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.19
-
 GTP M120G variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
0.23
-
 GTP variant G209D, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor

Metals/Ions

Metals/Ions Comment Organism Structure
Zn2+ content 1 molecule per monomer enzyme, enzyme requires a zinc ion which presumably activates a water molecule for excision of C-8 of GTP Streptomyces coelicolor
Zn2+ content of 1 molecule per enzyme monomer, enzyme requires a zinc ion which presumably activates a water molecule for excision of C-8 of GTP Streptomyces coelicolor

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
46100
-
 monomer, determined by SDS-PAGE Streptomyces coelicolor

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
GTP + H2O Streptomyces coelicolor
-
 2-amino-5-formylamino-6-ribosylamino-4(3H)-pyrimidinone 5'-phosphate + diphosphate + ?
-
 ?
GTP + H2O Streptomyces coelicolor enzyme catalyzes the first step in the biosynthesis of riboflavin 2,5-diamino-6-ribosylamino-4(3H)-pyrimidinone 5'-phosphate + diphosphate + ?
-
 ?
additional information Streptomyces coelicolor enzyme may be involved in the biosynthesis of toxoflavin, which is produced from GTP in a series of steps that are likely initiated by GCH II ?
-
 ?
additional information Streptomyces coelicolor substitution of Met with His leads to a protein that catalyzes the production of both 2-amino-5-formylamino-6-ribosylamino-4(3H)-pyrimidinone 5'-phosphate and 2,5-diamino-6-ribosylamino-4(3H)-pyrimidinone 5'-phosphate with equal efficiency ?
-
 ?

Organism

Organism UniProt Comment Textmining
Streptomyces coelicolor O88011 homologue SCO 6655
-
Streptomyces coelicolor Q9EWJ8 homologue SCO1441
-
Streptomyces coelicolor Q9KYI5 homologue SCO 2687
-

Purification (Commentary)

Purification (Comment) Organism
purification of wild type enzyme and its mutants by nickel affinity chromatography, site-directed variants are expressed in the native state and purified to homogeneity by ion exchange and hydrophobic interaction chromatographic steps Streptomyces coelicolor

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
GTP + H2O
-
 Streptomyces coelicolor 2-amino-5-formylamino-6-ribosylamino-4(3H)-pyrimidinone 5'-phosphate + diphosphate + ?
-
 ?
GTP + H2O enzyme catalyzes the first step in the biosynthesis of riboflavin Streptomyces coelicolor 2,5-diamino-6-ribosylamino-4(3H)-pyrimidinone 5'-phosphate + diphosphate + ?
-
 ?
additional information enzyme may be involved in the biosynthesis of toxoflavin, which is produced from GTP in a series of steps that are likely initiated by GCH II Streptomyces coelicolor ?
-
 ?
additional information substitution of Met with His leads to a protein that catalyzes the production of both 2-amino-5-formylamino-6-ribosylamino-4(3H)-pyrimidinone 5'-phosphate and 2,5-diamino-6-ribosylamino-4(3H)-pyrimidinone 5'-phosphate with equal efficiency Streptomyces coelicolor ?
-
 ?

Synonyms

Synonyms Comment Organism
3,4-dihydroxy-2-butanone 4-phosphate synthase
-
 Streptomyces coelicolor
GCH II
-
 Streptomyces coelicolor
GTP cyclohydrolase II
-
 Streptomyces coelicolor

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.42
-
 GTP M120D variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
1.2
-
 GTP variant Y326F, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
1.8
-
 GTP M120W variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
3.18
-
 GTP M120E variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
3.18
-
 GTP SCO 2687, variant Y123F, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
4.86
-
 GTP variant Y326M, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
10.8
-
 GTP M120H variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
20.4
-
 GTP M120T variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
42
-
 GTP M120A variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
46.8
-
 GTP variant M120F, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
57
-
 GTP M120G variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
66
-
 GTP M120C variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
78
-
 GTP M120I variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
78
-
 GTP SCO 2687, variant D127A, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
84
-
 GTP M120S variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
84
-
 GTP variant M120Y, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
108
-
 GTP M120N variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
120
-
 GTP M120Q variant, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
150
-
 GTP wild type, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
168
-
 GTP SCO 2687, variant G209D, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
204
-
 GTP SCO 2687, wild type, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
228
-
 GTP SCO 2687, variant R123M, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
348
-
 GTP SCO 2687, variant Y123M, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor
432
-
 GTP wild type, 1 mM substrate, 0.1 M Tris-HCl, pH 8, 5 mM MgCl2, 0.5 mM dithiothreitol Streptomyces coelicolor