Cloned (Comment) | Organism |
---|---|
gene deoD, DNA and amino acid sequence determination and analysis of clinical isolate PNP-Zg encoded by a deoD gene, sequence comparisons, recombinant expression of PNP-Zg in Escherichia coli strain BL21-CodonPlus(DE3)-RIL | Helicobacter pylori |
Crystallization (Comment) | Organism |
---|---|
purified recombinant enzyme PNP-Zg in ternary complex with hypoxanthine and phosphate molecules, obtained from three different crystallization conditions: B8 (0.2 M MgCl2, 0.1 M Tris-HCl, pH 7.0, 10 % PEG 8000 for structure HpPNP-1), A1 (0.2 M Li2SO4, 0.1 M Na acetate, pH 4.5, 50 % PEG 400 for structure HpPNP-2) and D3 (0.2 M Na chloride, 0.1 M Na/K phosphate, pH 6.2, 50% PEG 200 for structure HpPNP-3), 20°C, X-ray diffraction structure determination and analysis at 2.3-2.4 A resolution, molecular replacement using the structure of Escherichia coli PNP (PDB ID 1K9S) as template | Helicobacter pylori |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
142000 | - |
recombinant enzyme, gel filtration | Helicobacter pylori |
154716 | - |
sequence comparison | Helicobacter pylori |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
7-methylguanosine + phosphate | Helicobacter pylori | low activity | 7-methylguanine + alpha-D-ribose 1-phosphate | - |
r | |
adenosine + phosphate | Helicobacter pylori | very low activity | adenine + alpha-D-ribose 1-phosphate | - |
r | |
guanosine + phosphate | Helicobacter pylori | - |
guanine + alpha-D-ribose 1-phosphate | - |
r | |
inosine + phosphate | Helicobacter pylori | - |
hypoxanthine + alpha-D-ribose 1-phosphate | - |
r |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Helicobacter pylori | A0A0G2R4X7 | multifunctional fusion protein; clinical isolate PNP-Zg | - |
Purification (Comment) | Organism |
---|---|
recombinant enzyme PNP-Zg from Escherichia coli strain BL21-CodonPlus(DE3)-RIL by formycin affinity chromatography (elution with inosine), dialysis, and gel filtration, to homogeneity | Helicobacter pylori |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
purine ribonucleoside + phosphate = purine + alpha-D-ribose 1-phosphate | molecular reaction mechanism, overview | Helicobacter pylori |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
2.2 | - |
purified recombinant enzyme, pH 7.0, 25°C, substrate adenosine | Helicobacter pylori |
7.5 | - |
purified recombinant enzyme, pH 7.0, 25°C, substrate 7-methylguanosine | Helicobacter pylori |
80.4 | - |
purified recombinant enzyme, pH 7.0, 25°C, substrate inosine | Helicobacter pylori |
Storage Stability | Organism |
---|---|
storage of the purified enzyme at 4°C for one month at 0.4 mg/ml in 100 mM Tris-HCl, pH 7.6, results in an about 50% decrease in activity | Helicobacter pylori |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
7-methylguanosine + phosphate | low activity | Helicobacter pylori | 7-methylguanine + alpha-D-ribose 1-phosphate | - |
r | |
adenosine + phosphate | very low activity | Helicobacter pylori | adenine + alpha-D-ribose 1-phosphate | - |
r | |
guanosine + phosphate | - |
Helicobacter pylori | guanine + alpha-D-ribose 1-phosphate | - |
r | |
inosine + phosphate | - |
Helicobacter pylori | hypoxanthine + alpha-D-ribose 1-phosphate | - |
r |
Subunits | Comment | Organism |
---|---|---|
homohexamer | 6 * 24900, recombinant enzyme, SDS-PAGE, 6 * 25786, sequence calculation | Helicobacter pylori |
More | each monomer requires a portion of other monomer for the completion of the active site. The two monomers connected by approximate two-fold symmetry in this way form a dimer, where the monomers mutually donate to the active site amino acids His4 and Arg43. Such dimers are repeated by an approximate three-fold axis that is perpendicular to the two-fold axis to complete a set of three dimers that make the whole homohexamer of PNP, secondary structure analysis, overview | Helicobacter pylori |
Synonyms | Comment | Organism |
---|---|---|
DeoD | - |
Helicobacter pylori |
EC592_06680 | locus name | Helicobacter pylori |
HpPNP | - |
Helicobacter pylori |
PNP | - |
Helicobacter pylori |
PNP-Zg | - |
Helicobacter pylori |
purine nucleoside orthophosphate ribosyl transferase | - |
Helicobacter pylori |
purine nucleoside phosphorylase | - |
Helicobacter pylori |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
assay at | Helicobacter pylori |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
20 | 80 | purified recombinant enzyme, pH 7.0, 1 h. The enzyme is stable retaining over 90% of the initial activity at up to 37°C at pH 7.0, the activity is completely abolished at 60°C | Helicobacter pylori |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7 | - |
assay at | Helicobacter pylori |
pH Stability | pH Stability Maximum | Comment | Organism |
---|---|---|---|
2 | 12 | purified recombinant enzyme, 25°C, 1 h. The enzyme is stable retaining over 90% of the initial activity in the pH range 6.0-9.0 at 25°C. Stability decreases rapidly below pH 5.5 and above pH 9.5 | Helicobacter pylori |
General Information | Comment | Organism |
---|---|---|
metabolism | the purine metabolism in Helicobacter pylori is solely dependant on the salvage pathway and one of the key enzymes in this pathway is purine nucleoside phosphorylase (PNP) | Helicobacter pylori |
additional information | the structure of Helicobacter pylori PNP is typical for high molecular mass PNPs. Active site structure and ligand binding structure analysis, overview. The structure of Helicobacter pylori PNP-Zg represents the first reported case of a true dead-end complex of bacterial hexameric PNP bound with one of the reaction substrates, phosphate, and one of the reaction products, hypoxanthine, which is produced by the enzyme turnover during the purification procedure | Helicobacter pylori |
physiological function | PNP is the key enzyme in the purine salvage pathway. It catalyses the reversible phosphorolytic cleavage of the glycosidic bond of purine nucleosides (and some analogues) in the presence of phosphate as a second substrate | Helicobacter pylori |