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Literature summary for 1.2.1.12 extracted from
- Sperm-specific glyceraldehyde-3-phosphate dehydrogenase is stabilized by additional proline residues and an interdomain salt bridge (2014), Biochim. Biophys. Acta, 1844, 1820-1826 .
Cloned(Commentary)
| Cloned (Comment) | Organism |
|---|---|
| recombinant expression of mutant enzymes in Escherichia coli strain BL21(DE3), subcloning in Escherichia coli strain HB101 | Homo sapiens |
| recombinant expression of wild-type enzyme in Escherichia coli strain BL21 (DE3), subcloning in Escherichia coli strain HB101 | Homo sapiens |
Protein Variants
| Protein Variants | Comment | Organism |
|---|---|---|
| D311N | site-directed mutagenesis, the mutation breaks the salt bridge between the catalytic and NAD+-binding domains, the inactivation rate constant in the presence of GdnHCl increases 6fold, and the value of GdnHCl concentration corresponding to the protein half-denaturation decreases from 1.83 to 1.35 M. The mutation D311N enhances the enzymatic activity of the protein 2fold | Homo sapiens |
| E244Q | site-directed mutagenesis, mutation at the interdomain salt bridge | Homo sapiens |
| E96Q | site-directed mutagenesis, mutation at the interdomain salt bridge | Homo sapiens |
| additional information | construction of a plasmid encoding truncated GAPDS lacking 68 N-terminal amino acids (dN-GAPDS) | Homo sapiens |
| P111A | site-directed mutagenesis, mutation at first position of alpha-helix | Homo sapiens |
| P157A | site-directed mutagenesis, mutation at first position of alpha-helix | Homo sapiens |
| P164A | site-directed mutagenesis, mutation at beta-turn, the mutant shows reduced thermostability and reduced resistance against guanidine hydrochloride. The Tm value of the heat-absorption curve decreases by 3.3°C compared to the wild-type protein | Homo sapiens |
| P197A | site-directed mutagenesis, mutation at beta-turn | Homo sapiens |
| P213A | site-directed mutagenesis, mutation at beta-turn | Homo sapiens |
| P326A | site-directed mutagenesis, mutation at first position of alpha-helix, the mutant shows reduced thermostability and reduced resistance against guanidine hydrochloride. The Tm value of the heat-absorption curve decreases by 6.0°C compared to the wild-type protein | Homo sapiens |
Inhibitors
| Inhibitors | Comment | Organism | Structure |
|---|---|---|---|
| guanidine hydrochloride | unfolding of both wild type and mutant dN-GAPDS proteins is described by a single [GdnHCl]50 value. For the truncated mutant dN-GAPDS, it constitutes 1.83 M. Different mutations of dN-GAPDS alter this parameter to various extents. The most pronounced effect is observed in the case of mutants P111A, P157A, and D311N. The mutation P111A increases the value of [GdnHCl]50 by 0.43 M, the mutations P157A and D311N decrease the GdnHCl50 value by 0.36 and 0.48 M, respectively. In other mutants, the [GdnHCl]50 value is less affected or does not change, overview; unfolding of muscle isoenzyme GAPD is a two step process | Homo sapiens |  |
Molecular Weight [Da]
| Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
|---|---|---|---|
| 150000 | - |
native PAGE | Homo sapiens |
Natural Substrates/ Products (Substrates)
| Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| D-glyceraldehyde 3-phosphate + phosphate + NAD+ | Homo sapiens | - |
3-phospho-D-glyceroyl phosphate + NADH + H+ | - |
r | |
Organism
| Organism | UniProt | Comment | Textmining |
|---|---|---|---|
| Homo sapiens | O14556 | - |
- |
| Homo sapiens | P04406 | - |
- |
Purification (Commentary)
| Purification (Comment) | Organism |
|---|---|
| recombinant truncated mutant dN-GAPDS by cation exchange chromatography from Escherichia coli strain BL21(DE3). Recombinant enzyme mutants P197A and P164A by ammonium sulfate fractionation and dialysis, followed by anion exchange chromatography, the flow-through is further purified by hydrophobic interaction chromatography | Homo sapiens |
| recombinant wild-type enzyme from Escherichia coli strain BL21(DE3) | Homo sapiens |
Source Tissue
| Source Tissue | Comment | Organism | Textmining |
|---|---|---|---|
| skeletal muscle | - |
Homo sapiens | - |
| sperm cell | - |
Homo sapiens | - |
Specific Activity [micromol/min/mg]
| Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
|---|---|---|---|
| 45 | - |
purified recombinant truncated enzyme mutant dN-GAPDS, pH 8.9, 20°C, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
| 46 | - |
purified recombinant truncated enzyme mutant dN-GAPDS P213A, pH 8.9, 20°C, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
| 47 | - |
purified recombinant truncated enzyme mutant dN-GAPDS P111A, pH 8.9, 20°C, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
| 47 | - |
purified recombinant truncated enzyme mutant dN-GAPDS P164A, pH 8.9, 20°C, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
| 47 | - |
purified recombinant truncated enzyme mutant dN-GAPDS P326A, pH 8.9, 20°C, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
| 50 | - |
purified recombinant truncated enzyme mutant dN-GAPDS P157A, pH 8.9, 20°C, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
| 60 | - |
purified recombinant truncated enzyme mutant dN-GAPDS E244Q, pH 8.9, 20°C, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
| 64 | - |
purified recombinant truncated enzyme mutant dN-GAPDS E96Q, pH 8.9, 20°C, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
| 64 | - |
purified recombinant truncated enzyme mutant dN-GAPDS P197A, pH 8.9, 20°C, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
| 92 | - |
purified recombinant truncated enzyme mutant dN-GAPDS D311N, pH 8.9, 20°C, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
| 95 | - |
purified recombinant wild-type enzyme, pH 8.9, 20°C, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
Substrates and Products (Substrate)
| Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
|---|---|---|---|---|---|---|
| D-glyceraldehyde 3-phosphate + phosphate + NAD+ | - |
Homo sapiens | 3-phospho-D-glyceroyl phosphate + NADH + H+ | - |
r | |
Subunits
| Subunits | Comment | Organism |
|---|---|---|
| homotetramer | - |
Homo sapiens |
| homotetramer | 4 * 37000, about, SDS-PAGE | Homo sapiens |
| More | the residues P164 (beta-turn), P326 (first position of alpha-helix), and the interdomain salt bridge D311-H124 are significant for the enhanced stability of GAPDS. The salt bridge D311-H124 enhances stability of the active site of GAPDS at the expense of the catalytic activity. The N-terminal domain is hidden inside the cytoskeleton structures and does not interact with the catalytic part of the enzyme | Homo sapiens |
Synonyms
| Synonyms | Comment | Organism |
|---|---|---|
| GAPD | - |
Homo sapiens |
| GAPDS | - |
Homo sapiens |
| glyceraldehyde-3-phosphate dehydrogenase | - |
Homo sapiens |
| sperm-specific glyceraldehyde-3-phosphate dehydrogenase | - |
Homo sapiens |
Temperature Optimum [°C]
| Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
|---|---|---|---|
| 20 | - |
assay at, oxidation | Homo sapiens |
| 20 | - |
assay at, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
pH Optimum
| pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
|---|---|---|---|
| 8.9 | - |
assay at, oxidation | Homo sapiens |
| 8.9 | - |
assay at, oxidation of D-glyceraldehyde 3-phosphate | Homo sapiens |
Cofactor
| Cofactor | Comment | Organism | Structure |
|---|---|---|---|
| NAD+ | - |
Homo sapiens | |
| NADH | - |
Homo sapiens | |
pI Value
| Organism | Comment | pI Value Maximum | pI Value |
|---|---|---|---|
| Homo sapiens | below | - |
7 |
| Homo sapiens | recombinant truncated enzyme mutant dN-GAPDS | - |
8.3 |
General Information
| General Information | Comment | Organism |
|---|---|---|
| evolution | both GAPDS and GAPD are homotetramers with the sequence identity of about 70%. They are encoded by different genes which have emerged after duplication of the original gene during the early evolution of chordates | Homo sapiens |
| evolution | both GAPDS and GAPD are homotetramers with the sequence identity of about 70%. They are encoded by different genes which have emerged after duplication of the original gene during the early evolution of chordates. The GAPDS gene is lost by most lineages, and specialized to a testis-specific protein in reptilians and mammals | Homo sapiens |
| additional information | comparison of the sequences of muscle GAPD and sperm GAPDS isozymes reveals seven additional proline residues in the catalytic part of GAPDS | Homo sapiens |
| additional information | sperm-specific glyceraldehyde-3-phosphate dehydrogenase, GAPDS, is stabilized by additional proline residues and an interdomain salt bridge. Residues P164, P326, and the interdomain salt bridge D311-H124 are significant for the enhanced stability of GAPDS. The salt bridge D311-H124 enhances stability of the active site of GAPDS at expense of the catalytic activity. Comparison of the sequences of muscle GAPD and sperm GAPDS isozymes reveals seven additional proline residues in the catalytic part of GAPDS | Homo sapiens |
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