Activating Compound | Comment | Organism | Structure |
---|---|---|---|
HCO3- | Km about 50% increased in presence of sodium bicarbonate, crystallized enzyme structure complexed with HCO3- in its activated form | Geobacillus kaustophilus |
Cloned (Comment) | Organism |
---|---|
expressed in Escherichia coli BL21, pET17b expression vector | Geobacillus kaustophilus |
expressed in Escherichia coli, PCR-amplified genomic DNA from Bacillus subtilis strain 168 ligated into pET15b encoding an N-terminal six-His tag vector | Bacillus subtilis |
Crystallization (Comment) | Organism |
---|---|
hanging drop method, 17 A resolution, X-ray coordinates and structure factors determined, complexes with phosphate, with Mg2+, with Mg2+ and HCO3-, with Mg2+ and the alternate substrate 2,3-diketohexane 1-phosphate identified, activated enolase carboxylated on Lys173, conserved Lys98 in the N-terminal domain determined for C1 proton abstraction | Geobacillus kaustophilus |
Protein Variants | Comment | Organism |
---|---|---|
K147A | requires a 10fold greater concentration of protein for observation of enolization | Geobacillus kaustophilus |
K173A | detectable activity of about 3% of that of wild-type enolase, retains ability to enolize the desthio substrate | Geobacillus kaustophilus |
K98A | unable to catalyze the enolase reaction | Geobacillus kaustophilus |
additional information | site-directed mutagenesis of active site residues, spectrophotometric activity assay performed with elevated concentrations of the mutant enzymes | Geobacillus kaustophilus |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | activity depends on | Geobacillus kaustophilus |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
2,3-diketo-5-methylthiopentane 1-phosphate | Geobacillus kaustophilus | methionine salvage pathway | 2-hydroxy-3-keto-5-methylthiopent-1-ene 1-phosphate | - |
? | |
2,3-diketo-5-methylthiopentane 1-phosphate | Bacillus subtilis | methionine salvage pathway | 2-hydroxy-3-keto-5-methylthiopent-1-ene 1-phosphate | - |
? | |
2,3-diketo-5-methylthiopentane 1-phosphate | Bacillus subtilis 168 | methionine salvage pathway | 2-hydroxy-3-keto-5-methylthiopent-1-ene 1-phosphate | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Bacillus subtilis | P37869 | - |
- |
Bacillus subtilis 168 | P37869 | - |
- |
Geobacillus kaustophilus | Q5KVE7 | - |
- |
Purification (Comment) | Organism |
---|---|
recombinant protein, gel filtration | Geobacillus kaustophilus |
recombinant protein, gel filtration | Bacillus subtilis |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
additional information | - |
spectrophotometric assay described, D-ribulose 1-phosphate and 5-methylthio-D-ribulose 1-phosphate in the presence of limiting 5-methylthio-D-ribulose 1-phosphate dehydratase analyzed, enzyme concentrations from 0.1 to 10 microM used | Geobacillus kaustophilus |
additional information | - |
spectrophotometric assay described, generation of reaction products determined by NMR | Bacillus subtilis |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
2,3-diketo-5-methylthiopentane 1-phosphate | methionine salvage pathway | Geobacillus kaustophilus | 2-hydroxy-3-keto-5-methylthiopent-1-ene 1-phosphate | - |
? | |
2,3-diketo-5-methylthiopentane 1-phosphate | methionine salvage pathway | Bacillus subtilis | 2-hydroxy-3-keto-5-methylthiopent-1-ene 1-phosphate | - |
? | |
2,3-diketo-5-methylthiopentane 1-phosphate | methionine salvage pathway, stereochemical course of the reaction catalyzed by enolase determined, C1 proton abstraction | Geobacillus kaustophilus | 2-hydroxy-3-keto-5-methylthiopent-1-ene 1-phosphate | - |
? | |
2,3-diketo-5-methylthiopentane 1-phosphate | methionine salvage pathway, stereochemical course of the reaction catalyzed by enolase determined, C1 proton abstraction | Bacillus subtilis | 2-hydroxy-3-keto-5-methylthiopent-1-ene 1-phosphate | - |
? | |
2,3-diketo-5-methylthiopentane 1-phosphate | methionine salvage pathway | Bacillus subtilis 168 | 2-hydroxy-3-keto-5-methylthiopent-1-ene 1-phosphate | - |
? | |
2,3-diketo-5-methylthiopentane 1-phosphate | methionine salvage pathway, stereochemical course of the reaction catalyzed by enolase determined, C1 proton abstraction | Bacillus subtilis 168 | 2-hydroxy-3-keto-5-methylthiopent-1-ene 1-phosphate | - |
? | |
2,3-diketohexane 1-phosphate | alternate substrate, C1 proton abstraction | Geobacillus kaustophilus | ? | - |
? | |
2,3-diketohexane 1-phosphate | alternate substrate, C1 proton abstraction | Bacillus subtilis | ? | - |
? | |
2,3-diketohexane 1-phosphate | alternate substrate, C1 proton abstraction | Bacillus subtilis 168 | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
More | composed of a N-terminal alpha,beta domain and a C-terminal domain consisting of eight alpha,alpha barrels, polypeptides packed as tight dimers | Geobacillus kaustophilus |
Synonyms | Comment | Organism |
---|---|---|
enolase | - |
Geobacillus kaustophilus |
enolase | - |
Bacillus subtilis |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.5 | - |
assay at | Geobacillus kaustophilus |