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Literature summary for 2.4.1.1 extracted from

  • Schwarz, A.; Pierfederici, F.M.; Nidetzky, B.
    Catalytic mechanism of alpha-retaining glucosyl transfer by Corynebacterium callunae starch phosphorylase: the role of histidine-334 examined through kinetic characterization of site-directed mutants (2005), Biochem. J., 387, 437-445.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 2.4.1.1

Cloned(Commentary)

Cloned (Comment) Organism
expression of wild-type and mutant enzymes in Escherichia coli Corynebacterium callunae

Protein Variants

Protein Variants Comment Organism
H334A site-directed mutagenesis, catalytic efficiency in the phosphorolysis reaction is similar to the wild-type enzyme, 50fold reduced binding of alpha-D-glucose 1-phosphate compared to the wild-type enzyme Corynebacterium callunae
H334N site-directed mutagenesis, 50fold reduced catalytic efficiency in the phosphorolysis reaction and highly reduced selectivity for alpha-D-glucose 1-phosphate, versus alpha-D-xylose 1-phosphate, in the synthesis reaction with 10fold reduced binding of alpha-D-glucose 1-phosphate compared to the wild-type enzyme Corynebacterium callunae
H334Q site-directed mutagenesis, 150fold reduced catalytic efficiency in the phosphorolysis reaction and highly reduced selectivity for alpha-D-glucose 1-phosphate, versus alpha-D-xylose 1-phosphate, in the synthesis reaction with 10fold reduced binding of alpha-D-glucose 1-phosphate compared to the wild-type enzyme Corynebacterium callunae

Inhibitors

Inhibitors Comment Organism Structure
D-gluconic acid 1,5-lactone
-
 Corynebacterium callunae

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information stopped-flow and steady-state kinetics, wild-type and mutant enzymes Corynebacterium callunae
0.23
-
 starch pH 7.0, 30°C, recombinant wild-type enzyme, phosphorolysis Corynebacterium callunae
0.25
-
 starch pH 7.0, 30°C, recombinant mutant H334A, phosphorolysis Corynebacterium callunae
0.32
-
 starch pH 7.0, 30°C, recombinant mutant H334Q, phosphorolysis Corynebacterium callunae
0.85
-
 alpha-D-glucose 1-phosphate pH 7.0, 30°C, recombinant wild-type enzyme, synthesis Corynebacterium callunae
1
-
 starch pH 7.0, 30°C, recombinant mutant H334N, phosphorolysis Corynebacterium callunae
1.1
-
 alpha-D-glucose 1-phosphate pH 7.0, 30°C, recombinant mutant H334Q, synthesis Corynebacterium callunae
1.3
-
 alpha-D-glucose 1-phosphate pH 7.0, 30°C, recombinant mutant H334N, synthesis Corynebacterium callunae
1.6
-
 glycogen pH 7.0, 30°C, recombinant wild-type enzyme and mutant H334A, phosphorolysis Corynebacterium callunae
1.7
-
 glycogen pH 7.0, 30°C, recombinant mutant H334Q, phosphorolysis Corynebacterium callunae
1.8
-
 alpha-D-glucose 1-phosphate pH 7.0, 30°C, recombinant mutant H334A, synthesis Corynebacterium callunae
3.5
-
 maltopentaose pH 7.0, 30°C, recombinant mutant H334A, phosphorolysis Corynebacterium callunae
3.8
-
 maltopentaose pH 7.0, 30°C, recombinant wild-type enzyme, phosphorolysis Corynebacterium callunae
4.1
-
 maltopentaose pH 7.0, 30°C, recombinant mutant H334Q, phosphorolysis Corynebacterium callunae
6.8
-
 phosphate pH 7.0, 30°C, recombinant wild-type enzyme, phosphorolysis Corynebacterium callunae
12
-
 phosphate pH 7.0, 30°C, recombinant mutants H334A and H334Q, phosphorolysis Corynebacterium callunae
13
-
 phosphate pH 7.0, 30°C, recombinant mutant H334N, phosphorolysis Corynebacterium callunae
16
-
 glycogen pH 7.0, 30°C, recombinant mutant H334N, phosphorolysis Corynebacterium callunae
103
-
 maltopentaose pH 7.0, 30°C, recombinant mutant H334N, phosphorolysis Corynebacterium callunae

Organism

Organism UniProt Comment Textmining
Corynebacterium callunae
-
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and mutant enzymes from Escherichia coli Corynebacterium callunae

Reaction

Reaction Comment Organism Reaction ID
[(1->4)-alpha-D-glucosyl]n + phosphate = [(1->4)-alpha-D-glucosyl]n-1 + alpha-D-glucose 1-phosphate catalytic mechanism of alpha-retaining glucosyl transfer, His334 is involved in selectively stabilizing the oxycarbenium ion-like transition state of the reaction, ligand binding Corynebacterium callunae
a

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
(1,4-alpha-D-glucosyl)n + phosphate polysaccharide substrates are starch and glycogen, phosphorolysis is the preferred reaction, maltotetraose is the smallest substrate accepted Corynebacterium callunae (1,4-alpha-D-glucosyl)n-1 + alpha-D-glucose 1-phosphate
-
 r
(1,4-alpha-D-glucosyl)n-1 + alpha-D-xylose 1-phosphate polysaccharide substrate is starch, very low activity Corynebacterium callunae alpha-D-xylose-(1,4-alpha-D-glucosyl)n + phosphate
-
 r
glycogen + glucose 1-phosphate
-
 Corynebacterium callunae glycogen + phosphate
-
 ?
maltopentaose + phosphate
-
 Corynebacterium callunae maltotetraose + alpha-D-glucose 1-phosphate
-
 r
starch + glucose 1-phosphate
-
 Corynebacterium callunae starch + phosphate
-
 ?

Synonyms

Synonyms Comment Organism
starch phosphorylase
-
 Corynebacterium callunae

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
30
-
 assay at Corynebacterium callunae

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.00048
-
 alpha-D-xylose 1-phosphate pH 7.0, 30°C, recombinant mutant H334N, synthesis Corynebacterium callunae
0.0026
-
 alpha-D-xylose 1-phosphate pH 7.0, 30°C, recombinant wild-type enzyme, synthesis Corynebacterium callunae
0.0028
-
 alpha-D-xylose 1-phosphate pH 7.0, 30°C, recombinant mutant H334A, synthesis Corynebacterium callunae
0.0066
-
 alpha-D-xylose 1-phosphate pH 7.0, 30°C, recombinant mutant H334Q, synthesis Corynebacterium callunae
0.52
-
 starch pH 7.0, 30°C, recombinant mutant H334Q, phosphorolysis Corynebacterium callunae
0.68
-
 alpha-D-glucose 1-phosphate pH 7.0, 30°C, recombinant mutant H334Q, synthesis Corynebacterium callunae
1.27
-
 alpha-D-glucose 1-phosphate pH 7.0, 30°C, recombinant mutant H334N, synthesis Corynebacterium callunae
1.5
-
 starch pH 7.0, 30°C, recombinant mutant H334N, phosphorolysis Corynebacterium callunae
49
-
 alpha-D-glucose 1-phosphate pH 7.0, 30°C, recombinant mutant H334A, synthesis Corynebacterium callunae
52
-
 alpha-D-glucose 1-phosphate pH 7.0, 30°C, recombinant wild-type enzyme, synthesis Corynebacterium callunae
74
-
 starch pH 7.0, 30°C, recombinant wild-type enzyme, phosphorolysis Corynebacterium callunae
80
-
 starch pH 7.0, 30°C, recombinant mutant H334A, phosphorolysis Corynebacterium callunae

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7
-
 assay at Corynebacterium callunae

Cofactor

Cofactor Comment Organism Structure
pyridoxal 5'-phosphate
-
 Corynebacterium callunae