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Literature summary for 5.1.3.1 extracted from

  • Akana, J.; Fedorov, A.A.; Fedorov, E.; Novak, W.R.; Babbitt, P.C.; Almo, S.C.; Gerlt, J.A.
    D-Ribulose 5-phosphate 3-epimerase: functional and structural relationships to members of the ribulose-phosphate binding (beta/alpha)8-barrel superfamily (2006), Biochemistry, 45, 2493-2503.
    View publication on PubMed

Activating Compound

Activating Compound Comment Organism Structure
Zn2+ RPE is activated by Zn2+ which binds with a stoichiometry of one ion per polypeptide Streptococcus pyogenes

Cloned(Commentary)

Cloned (Comment) Organism
the gene encoding RPE is expressed in the Escherichia coli strain BL21 (DE3) Streptococcus pyogenes

Crystallization (Commentary)

Crystallization (Comment) Organism
crystal structure of the RPE is solved at 1.8 A resolution in the presence of D-xylitol 5-phosphate, an inert analogue of the D-xylulose 5-phosphate substrate. This structure suggests that the 2,3-enediolate intermediate in the 1,1-proton transfer reaction is stabilized by bidentate coordination to the Zn2+ that also is liganded to His 34, Asp 36, His 67, and Asp 176, the carboxylate groups of the Asp residues are positioned also to function as the acid/base catalysts Streptococcus pyogenes

Protein Variants

Protein Variants Comment Organism
D176A no detectable activity, substrate= 10 mM D-ribulose 5-phosphate Streptococcus pyogenes
D36A no detectable activity, substrate= 10 mM D-ribulose 5-phosphate Streptococcus pyogenes
H34A mutant discloses decreased affinity for Zn2+, kcat = 200/sec, substrate = 10 mM D-ribulose 5-phosphate, +10 micromol/l ZnCl2 Streptococcus pyogenes
H67A mutant discloses decreased affinity for Zn2+, kcat = 270/sec, substrate = 10 mM D-ribulose 5-phosphate, +10 micromol/l ZnCl2 Streptococcus pyogenes
additional information H34A, D36A, H67A, and D176A mutants are constructed since these His and Asp residues are strictly conserved in all RPEs and are observed to coordinate Zn2+ in the reported structures of the RPE from rice Streptococcus pyogenes

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.2
-
D-ribulose 5-phosphate
-
Streptococcus pyogenes

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ assay carried out at 10 mM Mg2+ Streptococcus pyogenes

Organism

Organism UniProt Comment Textmining
Streptococcus pyogenes Q9A1H8
-
-

Purification (Commentary)

Purification (Comment) Organism
lysate is applied to a chelating sepharose fast flow column charged with Ni2+, the N-terminal His tag is removed by thrombin cleavage Streptococcus pyogenes

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
D-Ribulose 5-phosphate
-
Streptococcus pyogenes D-Xylulose 5-phosphate
-
?

Synonyms

Synonyms Comment Organism
D-ribulose 5-phosphate 3-epimerase
-
Streptococcus pyogenes
RPE
-
Streptococcus pyogenes

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
100
-
D-ribulose 5-phosphate RPE metal-free, stoichiometry of Zn2+: 0.1 ion per polypeptide, substrate 10 mM D-ribulose 5-phosphate Streptococcus pyogenes
200
-
D-ribulose 5-phosphate mutant H34A, + 10 micromol/l ZnCl2 Streptococcus pyogenes
270
-
D-ribulose 5-phosphate mutant H67A, + 10 micromol/l ZnCl2 Streptococcus pyogenes
480
-
D-ribulose 5-phosphate RPE as isolated, stoichiometry of Zn2+: 0.3 ion per polypeptide, substrate 10 mM D-ribulose 5-phosphate Streptococcus pyogenes
840
-
D-ribulose 5-phosphate RPE as isolated + 0.5 mM ZnCl2 present in assay, stoichiometry of Zn2+: 0.9 ion per polypeptide, substrate 10 mM D-ribulose 5-phosphate Streptococcus pyogenes
870
-
D-ribulose 5-phosphate RPE metal-free + 0.5 mM ZnCl2 present in assay, stoichiometry of Zn2+: 0.9 ion per polypeptide, substrate 10 mM D-ribulose 5-phosphate Streptococcus pyogenes

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
assay at Streptococcus pyogenes