Application | Comment | Organism |
---|---|---|
drug development | the enzyme is a potential target for development of antibiotic compounds | Clostridioides difficile |
Cloned (Comment) | Organism |
---|---|
gene alr, sequence comparisons, recombinant expression of codon-optimized wild-type and mutant enzymes in Escherichia coli strain Rosetta 2 (pLysS)DE3, recombinant expression of wild-type and mutant enzymes in Escherichia coli alr/dadX double-knockout mutant strain MB2159, the wild-type and K271T mutant CdAlr proteins are able to complement the D-alanine auxotrophy and restore its growth in D-alanine-free medium | Clostridioides difficile |
Crystallization (Comment) | Organism |
---|---|
wild-type enzyme in complex with pyridoxal-5'-phosphate or with D-cycloserine, and enzyme mutant K271T in complex with pyridoxal 5'-phosphate, sitting drop vapor diffusion method, mixing of 0.002 ml of 32 m/ml protein in 10 mM PLP, 1 mM TCEP, and 50 mM Tris-HCl, pH 8.0, with 0.0015 ml of reservoir solution containing 0.2 M ammonium sulfate, 0.1 M Bis-Tris, pH 6.5, 25% w/v PEG 3350 for the wild-type/PLP complex, and additional 100 mM cycloserine, 200 mM sodium formate, 20% w/v PEG 3350 for the wild-type enzyme/inhibitor complex, or containing 0.17 M lithium sulfate, 0.085 M Tris-HCl, pH 8.5, 25.5% w/v PEG 4000, and 20% v/v glycerol for the enzyme mutant/PLP complex, X-ray diffraction structure determination and analysis at 2.1-2.6 A resolution | Clostridioides difficile |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
D-cycloserine | active site bound inhibitor, binding structure, overview | Clostridioides difficile |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | Michaelis-Menten kinetics | Clostridioides difficile | |
5.4 | - |
D-alanine | pH 8.5, temperature not specified in the publication, recombinant mutant K271T | Clostridioides difficile | |
7 | - |
D-alanine | pH 8.5, temperature not specified in the publication, recombinant wild-type enzyme | Clostridioides difficile | |
13.8 | - |
L-alanine | pH 8.5, temperature not specified in the publication, recombinant mutant K271T | Clostridioides difficile | |
17.6 | - |
L-alanine | pH 8.5, temperature not specified in the publication, recombinant wild-type enzyme | Clostridioides difficile |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
80530 | - |
mutant enzyme, gel filtration | Clostridioides difficile |
81090 | - |
wild-type enzyme, gel filtration | Clostridioides difficile |
86400 | - |
mutant enzyme, about, sequence calculation | Clostridioides difficile |
86600 | - |
wild-type enzyme, about, sequence calculation | Clostridioides difficile |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
L-alanine | Clostridioides difficile | - |
D-alanine | - |
r | |
L-alanine | Clostridioides difficile 630 | - |
D-alanine | - |
r |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Clostridioides difficile | Q180W0 | strain 630 is virulent and multidrug-resistant | - |
Clostridioides difficile 630 | Q180W0 | strain 630 is virulent and multidrug-resistant | - |
Purification (Comment) | Organism |
---|---|
recombinant codon-optimized wild-type and mutant enzymes from Escherichia coli strain Rosetta 2 (pLysS) DE3 by anion exchange chromatography, gel filtration, and dialysis | Clostridioides difficile |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
L-alanine | - |
Clostridioides difficile | D-alanine | - |
r | |
L-alanine | reversible racemization | Clostridioides difficile | D-alanine | - |
r | |
L-alanine | - |
Clostridioides difficile 630 | D-alanine | - |
r | |
L-alanine | reversible racemization | Clostridioides difficile 630 | D-alanine | - |
r |
Subunits | Comment | Organism |
---|---|---|
homodimer | 2 x 43306, recombinant wild-type enzyme, mass spectrometry, 2 * 43279, recombinant mutant enzyme, mass spectrometry | Clostridioides difficile |
More | the enzyme forms a homodimer with two active sites in which the cofactor pyridoxal 5'-phosphate is bound. Intermolecular interactions, overview | Clostridioides difficile |
Synonyms | Comment | Organism |
---|---|---|
ALR | - |
Clostridioides difficile |
CdAlr | - |
Clostridioides difficile |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
19.25 | - |
D-alanine | pH 8.5, temperature not specified in the publication, recombinant wild-type enzyme | Clostridioides difficile | |
59.3 | - |
L-alanine | pH 8.5, temperature not specified in the publication, recombinant wild-type enzyme | Clostridioides difficile | |
1298.4 | - |
D-alanine | pH 8.5, temperature not specified in the publication, recombinant mutant K271T | Clostridioides difficile | |
3275.5 | - |
L-alanine | pH 8.5, temperature not specified in the publication, recombinant mutant K271T | Clostridioides difficile |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8.5 | - |
assay at | Clostridioides difficile |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
pyridoxal 5'-phosphate | - |
Clostridioides difficile |
General Information | Comment | Organism |
---|---|---|
additional information | analysis of the enzyme structure, active site structure, and intermolecular interactions, structure comparisons, overview. The N-terminal and C-terminal domains of all reported Alr structures are connected by a short hinge region, but the hinge angles between the N- and C-terminal of Alrs vary | Clostridioides difficile |
physiological function | alanine racemase (Alr) is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that catalyzes the reversible racemization of L- and D-alanine. D-alanine is an essential component of the bacterial cell-wall peptidoglycan | Clostridioides difficile |