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Literature summary for 1.1.1.271 extracted from

  • Lau, S.T.; Tanner, M.E.
    Mechanism and active site residues of GDP-fucose synthase (2008), J. Am. Chem. Soc., 130, 17593-17602.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
wild-type and mutants introduced into the pT7GFS vector. Plasmid encodes for the expression of GFS bearing N-terminal hexahistidine tags, expressed in Escherichia coli BL21 (DE3) Escherichia coli K-12

Protein Variants

Protein Variants Comment Organism
C109S active site mutant, produces GDP-6-deoxy-D-altrose as its major product (75%) besides GDP-L-fucose (25%) out of GDP-4-keto-6-deoxy-D-mannose Escherichia coli
C109S produces GDP-6-deoxy-D-altrose as its major product, thus C-3'' epimerization occurs first and premature reduction of the GDP-4-keto-6-deoxy-D-altrose intermediate becomes competitive with GDP-L-fucose production. Results in the appearance of a kinetic isotope effect when [3''-2H]-GDP-6-deoxy-4-ketomannose is used as a substrate. The mutant catalyzes a rapid wash-in of solvent derived deuterium into the C-5'' position of GDP-fucose in the presence of NADP+ Escherichia coli K-12
H179Q is inactive, readily catalyzes the wash-out of deuterium from the C-3'' position of [3''-2H]-GDP-6-deoxy-4-keto-mannose Escherichia coli K-12
H179Q the mutant catalyzes isotope exchange into starting material indicates that its active site has not been dramatically perturbed and it is still able to bind substrate and catalyze deprotonation events Escherichia coli

Organism

Organism UniProt Comment Textmining
Escherichia coli P32055
-
-
Escherichia coli K-12
-
-
-

Purification (Commentary)

Purification (Comment) Organism
by metal affinity chromatography Escherichia coli K-12

Storage Stability

Storage Stability Organism
-80°C, 3 months Escherichia coli K-12

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
GDP-4-keto-6-deoxy-D-mannose + NADPH + H+
-
Escherichia coli K-12 GDP-L-fucose + NADP+
-
?
GDP-4-keto-6-deoxy-D-mannose + NADPH + H+
-
Escherichia coli GDP-L-fucose + NADP+
-
?

Synonyms

Synonyms Comment Organism
GDP-4-keto-6-deoxy-D-mannose epimerase/reductase
-
Escherichia coli K-12
GDP-4-keto-6-deoxy-D-mannose epimerase/reductase
-
Escherichia coli
GDP-fucose synthase
-
Escherichia coli K-12
GDP-fucose synthase
-
Escherichia coli
GFS
-
Escherichia coli K-12
GFS
-
Escherichia coli
GMER
-
Escherichia coli K-12
GMER
-
Escherichia coli

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1
-
GDP-6-deoxy-4-keto-mannose mutant C109S, isotope [5''-2H]-GDP-6-deoxy-4-keto-mannose Escherichia coli K-12
1
-
GDP-6-deoxy-4-keto-mannose wild-type, isotope [3''-2H]-GDP-6-deoxy-4-keto-mannose Escherichia coli K-12
1
-
GDP-6-deoxy-4-keto-mannose wild-type, isotope [5''-2H]-GDP-6-deoxy-4-keto-mannose Escherichia coli K-12
1.5
-
GDP-6-deoxy-4-keto-mannose mutant C109S, isotope [3''-2H]-GDP-6-deoxy-4-keto-mannose Escherichia coli K-12

Cofactor

Cofactor Comment Organism Structure
NADPH
-
Escherichia coli K-12
NADPH
-
Escherichia coli