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Information on EC 2.4.2.1 - purine-nucleoside phosphorylase and Organism(s) Escherichia coli and UniProt Accession P0ABP8
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2.4.2.1 purine-nucleoside phosphorylaseIUBMB Comments
Specificity not completely determined. Can also catalyse ribosyltransferase reactions of the type catalysed by EC 2.4.2.5, nucleoside ribosyltransferase.
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Word Map
- 2.4.2.1
- p-nitrophenol
- deaminase
- polynucleotide
- lymphocyte
- hypoxanthine
-
phosphorolysis
- rnase
-
salvage
- guanine
-
paraneoplastic
- t-cells
- pemphigus
- immunodeficiency
-
pincer
- deoxyguanosine
- exoribonuclease
- phosphoribosyltransferase
- polyneuropathy
- 5'-nucleotidase
-
practitioner
- dgtp
- nurse
-
degradosome
-
hypoxanthine-guanine
- parathion
- blister
-
bullous
-
mucocutaneous
- ribonucleosides
-
desmogleins
- deoxynucleosides
- deoxycytidine
- hgprt
-
phrenic
- castleman
- deoxyadenosine
- 2'-deoxyguanosine
-
metal-ligand
- obliterans
- pneumoperitoneum
- desmoplakins
- pemphigoid
-
square-planar
-
photocatalysts
- lesch-nyhan
-
iridium
-
photocatalytic
- acantholysis
- medicine
-
exonucleolytic
- p-aminophenol
- drug development
- analysis
- pharmacology
- synthesis
The taxonomic range for the selected organisms is: Escherichia coli
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Reaction Schemes
Synonyms
pnp, purine nucleoside phosphorylase, pnpase, pfpnp, purine-nucleoside phosphorylase, adenosine phosphorylase, hspnp, hppnp, tvpnp, inosine phosphorylase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
adenosine phosphorylase
-
-
-
-
inosine phosphorylase
-
-
-
-
inosine-guanosine phosphorylase
-
-
-
-
nucleotide phosphatase (2.4.2.1)
-
-
-
-
phosphorylase, purine nucleoside
-
-
-
-
PNPase
Pu-NPase
-
-
-
-
PUNP
-
-
-
-
PUNPI
-
-
-
-
PUNPII
-
-
-
-
purine deoxynucleoside phosphorylase
-
-
-
-
purine deoxyribonucleoside phosphorylase
-
-
-
-
purine nucleoside phosphorylase
purine ribonucleoside phosphorylase
-
-
-
-
PNPase
-
-
-
-
purine nucleoside phosphorylase
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
purine ribonucleoside + phosphate = purine + alpha-D-ribose 1-phosphate
molecular mechanism of catalysis involving protonation of the purine ring position N7, open and closed active site conformations, overview
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pentosyl group transfer
-
-
-
-
PATHWAY SOURCE
PATHWAYS
BRENDA
KEGG
-
-, -, -, -, -, -, -, -, -, -, -, -, -, -, -
SYSTEMATIC NAME
IUBMB Comments
purine-nucleoside:phosphate ribosyltransferase
Specificity not completely determined. Can also catalyse ribosyltransferase reactions of the type catalysed by EC 2.4.2.5, nucleoside ribosyltransferase.
CAS REGISTRY NUMBER
COMMENTARY
9030-21-1
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
1,N6-ethenoadenine + alpha-D-ribose 1-phosphate
1,N6-ethenoadenine N7-riboside + phosphate
-
-
-
r
1,N6-ethenoadenine + alpha-D-ribose 1-phosphate
1,N6-ethenoadenine N9-riboside + phosphate
-
-
-
r
1,N6-ethenoadenosine + phosphate
1,N6-ethenoadenine + beta-D-ribose 1-phosphate
i.e. 3-beta-D-ribosylimidazo[2,l-i]purine
-
-
r
1-methyladenosine + phosphate
1-methyladenine + alpha-D-ribose 1-phosphate
-
-
-
r
1-methylguanosine + phosphate
1-methylguanine + alpha-D-ribose 1-phosphate
-
-
-
r
2'-deoxyadenosine + phosphate
adenine + 2-deoxy-alpha-D-ribose 1-phosphate
54.7% of the activity with 2'-deoxyinosine
-
-
?
2'-deoxyguanosine + phosphate
guanine + 2-deoxy-alpha-D-ribose 1-phosphate
83.2% of the activity with 2'-deoxyinosine
-
-
?
2'-deoxyinosine + phosphate
hypoxanthine + 2-deoxy-alpha-D-ribose 1-phosphate
-
-
-
?
2,6-diamino-8-azapurine + alpha-D-ribose 1-phosphate
N7-D-ribosyl-2,6-diamino-8-azapurine + phosphate
enzyme PNP mutant D204N
-
-
r
2,6-diamino-8-azapurine + alpha-D-ribose 1-phosphate
N8-D-ribosyl-2,6-diamino-8-azapurine + phosphate
wild-type enzyme PNP
-
-
r
2,6-diamino-8-azapurine + alpha-D-ribose 1-phosphate
N9-D-ribosyl-2,6-diamino-8-azapurine + phosphate
enzyme PNP mutant D204N
-
-
r
2-amino-6-mercapto-7-methylpurine ribonucleoside + alpha-D-ribose 1-phosphate
?
-
-
-
?
2-amino-6-mercapto-7-methylpurine ribonucleoside + phosphate
alpha-D-ribose 1-phosphate + 2-amino-6-mercapto-7-methylpurine
-
-
-
?
2-fluoro-2-deoxyadenosine + phosphate
adenine + 2-fluoro-2-deoxy-alpha-D-ribose 1-phosphate
-
-
-
?
2-fluoroadenosine + phosphate
2-fluoroadenine + alpha-D-ribose 1-phosphate
-
-
-
?
3-(beta-D-ribofuranosyl)adenine + alpha-D-ribose 1-phosphate
?
-
-
-
?
3-(beta-D-ribofuranosyl)hyopxanthine + alpha-D-ribose 1-phosphate
?
-
-
-
?
6-mercaptopurine-2'-deoxyriboside + phosphate
6-mercaptopurine + 2-deoxy-alpha-D-ribose 1-phosphate
-
-
-
r
7-(beta-D-ribofuranosyl)guanine + alpha-D-ribose 1-phosphate
?
-
-
-
?
7-(beta-D-ribofuranosyl)hypoxanthine + alpha-D-ribose 1-phosphate
?
-
-
-
?
7-methyladenosine + phosphate
7-methyladenine + alpha-D-ribose 1-phosphate
-
-
-
?
7-methylinosine + phosphate
7-methylhypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
?
8-aminoguanine + alpha-D-ribose 1-phosphate
8-aminoguanosine + phosphate
-
-
-
?
8-azaadenine + alpha-D-ribose 1-phosphate
8-azaadenosine + phosphate
-
-
-
r
8-azaguanosine + phosphate
8-azaguanine + alpha-D-ribose 1-phosphate
-
-
-
r
9-beta-D-(2-deoxyribofuranosyl)-6-methylthiopurine + phosphate
6-methylthiopurine + 2-deoxy-alpha-D-ribose 1-phosphate
-
-
-
?
9-beta-D-arabinosyl-2-fluoroadenine + phosphate
2-fluoroadenine + beta-D-arabinose 1-phosphate
i.e. fludarabine
-
-
r
9-beta-D-ribofuranosyl-6-methylthiopurine + phosphate
6-methylthiopurine + alpha-D-ribose 1-phosphate
-
-
-
?
deoxyadenosine + phosphate
alpha-D-ribose 1-phosphate + adenine
-
-
-
?
deoxyguanosine + phosphate
guanine + alpha-D-deoxyribose 1-phosphate
-
-
-
?
deoxyinosine + phosphate
hypoxanthine + alpha-D-deoxyribose 1-phosphate
-
-
-
r
N7-D-ribosyl-2,6-diamino-8-azapurine + phosphate
2,6-diamino-8-azapurine + alpha-D-ribose 1-phosphate
enzyme PNP mutant D204N
-
-
r
N8-D-ribosyl-2,6-diamino-8-azapurine + phosphate
2,6-diamino-8-azapurine + alpha-D-ribose 1-phosphate
wild-type enzyme PNP
-
-
r
N9-D-ribosyl-2,6-diamino-8-azapurine + phosphate
2,6-diamino-8-azapurine + alpha-D-ribose 1-phosphate
enzyme PNP mutant D204N
-
-
r
nicotinamide riboside + phosphate
nicotinamide + alpha-D-ribose 1-phosphate
-
-
-
?
1,6-dihydropurine riboside + phosphate
alpha-D-ribose 1-phosphate + 1,6-dihydropurine
-
-
-
-
?
1,N2-ethenoguanine + alpha-D-ribose 1-phosphate
1,N2-ethenoguanosine + phosphate
-
-
-
r
1,N6-etheno-isoguanine + alpha-D-ribose 1-phosphate
1,N6-etheno-isoguanosine + phosphate
isoguanine is 2-hydroxy-6-aminopurine and showing a solvent-induced keto-enol tautomerism
-
-
r
1,N6-ethenoadenosine + phosphate
1,N6-ethenoadenine + beta-D-ribose 1-phosphate
i.e. 3-beta-D-ribosylimidazo[2,l-i]purine
-
-
r
1-methylguanosine + phosphate
alpha-D-ribose 1-phosphate + 1-methylguanine
-
-
-
-
?
1-methylinosine + phosphate
alpha-D-ribose 1-phosphate + 1-methylhypoxanthine
-
-
-
-
?
2',3'-dideoxyinosine + phosphate
hypoxanthine + alpha-D-2,3-dideoxyribose 1-phosphate
-
-
-
-
?
2'-deoxyadenosine + phosphate
adenine + 2-deoxy-alpha-D-ribose 1-phosphate
-
12% of the activity with inosine
-
-
?
2'-deoxyguanosine + phosphate
guanine + 2-deoxy-alpha-D-ribose 1-phosphate
-
20% of the activity with inosine
-
-
?
2,6-diaminopurine ribonucleoside + phosphate
2,6-diaminopurine + alpha-D-ribose 1-phosphate
-
-
-
-
?
2-chloroadenosine + phosphate
2-chloroadenine + alpha-D-ribose 1-phosphate
-
-
-
-
?
3-(beta-D-ribofuranosyl)hyopxanthine + alpha-D-ribose 1-phosphate
?
-
-
-
-
?
3-deazainosine + phosphate
alpha-D-ribose 1-phosphate + 3-deazahypoxanthine
-
-
-
-
?
6-(2-thienyl)-9-(beta-D-ribofuranosyl)purine + phosphate
6-(2-thienyl)purine + alpha-D-ribose 1-phosphate
-
-
-
-
?
6-cyclopropyl-9-(beta-D-ribofuranosyl)purine + phosphate
6-cyclopropylpurine + alpha-D-ribose 1-phosphate
-
-
-
-
?
6-ethyl-9-(beta-D-ribofuranosyl)purine + phosphate
6-ethylpurine + alpha-D-ribose 1-phosphate
-
-
-
-
?
6-phenyl-9-(beta-D-ribofuranosyl)purine + phosphate
6-phenylpurine + alpha-D-ribose 1-phosphate
-
-
-
-
?
7-methyladenosine + phosphate
7-methyladenine + alpha-D-ribose 1-phosphate
-
-
-
-
?
7-methylguanosine + phosphate
7-methylguanine + alpha-D-ribose 1-phosphate
-
-
-
-
?
7-methylinosine + phosphate
7-methylhypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
-
?
9-(6-deoxy-5-C-methyl-beta-D-ribo-hexofuranosyl)-6-methylpurine + phosphate
?
-
-
-
-
r
9-[beta-D-arabinofuranosyl]-2-fluoro-adenine + phosphate
?
-
a toxic prodrug
-
-
?
adenosine + phosphate
alpha-D-ribose 1-phosphate + adenine
activity with mutant enzyme N239D, wild-type and mutant enzyme Y191L show no activity
-
-
?
deoxyadenosine + phosphate
alpha-D-ribose 1-phosphate + adenine
-
61% of the activity with deoxyguanosine
-
-
?
deoxyguanosine + phosphate
guanine + alpha-D-deoxyribose 1-phosphate
-
-
-
-
?
N2,3-etheno-O6-methylguanine + alpha-D-ribose 1-phosphate
N2,3-etheno-O6-methylguanosine + phosphate
-
-
-
r
N2,3-ethenoguanine + alpha-D-ribose 1-phosphate
N2,3-ethenoguanosine + phosphate
-
-
-
r
nicotinamide riboside + phosphate
nicotinamide + alpha-D-ribose 1-phosphate
-
-
-
r
O6-methyl-N2,3-ethenoguanine + alpha-D-ribose 1-phosphate
O6-methyl-N2,3-ethenoguanosine + phosphate
-
-
-
r
purine deoxyribonucleoside + phosphate
purine + 2-deoxy-alpha-D-ribose 1-phosphate
-
-
-
-
?
purine ribonucleoside + phosphate
purine + alpha-D-ribose 1-phosphate
-
-
-
-
?
xanthosine + phosphate
xanthine + alpha-D-ribose 1-phosphate
-
-
-
r
7-methylguanosine + phosphate
7-methylguanine + alpha-D-ribose 1-phosphate
-
-
-
?
7-methylguanosine + phosphate
7-methylguanine + alpha-D-ribose 1-phosphate
-
-
-
-
?
7-methylguanosine + phosphate
7-methylguanine + alpha-D-ribose 1-phosphate
-
-
-
?
8-azaguanine + alpha-D-ribose 1-phosphate
8-azaguanosine + phosphate
-
-
-
r
8-azaguanine + alpha-D-ribose 1-phosphate
8-azaguanosine + phosphate
-
-
-
r
adenosine + phosphate
adenine + alpha-D-ribose 1-phosphate
-
-
-
?
adenosine + phosphate
adenine + alpha-D-ribose 1-phosphate
-
-
-
?
adenosine + phosphate
adenine + alpha-D-ribose 1-phosphate
-
-
-
r
adenosine + phosphate
adenine + alpha-D-ribose 1-phosphate
59.9% of the activity with 2'-deoxyinosine
-
-
?
guanosine + phosphate
guanine + alpha-D-ribose 1-phosphate
-
-
-
?
guanosine + phosphate
guanine + alpha-D-ribose 1-phosphate
-
-
-
r
guanosine + phosphate
guanine + alpha-D-ribose 1-phosphate
58.7% of the activity with 2'-deoxyinosine
-
-
?
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
?
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
?
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
r
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
r
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
highest activity
-
-
r
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
83% of the activity with 2'-deoxyinosine
-
-
?
2'-deoxyinosine + phosphate
hypoxanthine + 2-deoxy-alpha-D-ribose 1-phosphate
-
-
-
-
?
2'-deoxyinosine + phosphate
hypoxanthine + 2-deoxy-alpha-D-ribose 1-phosphate
-
22% of the activity with inosine
-
-
?
9-[2-deoxy-beta-D-ribofuranosyl]-6-methylpurine + phosphate
?
-
a toxic prodrug
-
-
?
9-[6-deoxy-beta-L-talofuranosyl]-2-fluoro-adenine + phosphate
?
-
a toxic prodrug, substrate of enzyme mutant M64V, the drug product has anti-tumor activity
-
-
?
9-[6-deoxy-beta-L-talofuranosyl]-2-fluoro-adenine + phosphate
?
-
a toxic prodrug, substrate of enzyme mutant M64V
-
-
?
9-[6-deoxy-beta-L-talofuranosyl]-6-methylpurine + phosphate
?
-
a toxic prodrug, substrate of enzyme mutant M64V, the drug product has anti-tumor activity
-
-
?
9-[6-deoxy-beta-L-talofuranosyl]-6-methylpurine + phosphate
?
-
a toxic prodrug, substrate of enzyme mutant M64V, liberation of methylpurine
-
-
?
9-[beta-L-lyxofuranosyl]-2-fluoro-adenine + phosphate
?
-
a toxic prodrug, substrate of enzyme mutant M64V, the drug product has anti-tumor activity
-
-
?
9-[beta-L-lyxofuranosyl]-2-fluoro-adenine + phosphate
?
-
a toxic prodrug, substrate of enzyme mutant M64V
-
-
?
adenosine + phosphate
adenine + alpha-D-ribose 1-phosphate
-
no activity
-
-
?
adenosine + phosphate
adenine + alpha-D-ribose 1-phosphate
-
61% of the activity with deoxyguanosine
-
-
?
adenosine + phosphate
adenine + alpha-D-ribose 1-phosphate
-
41% of the activity with inosine
-
-
?
deoxyinosine + phosphate
hypoxanthine + alpha-D-deoxyribose 1-phosphate
-
-
-
-
r
deoxyinosine + phosphate
hypoxanthine + alpha-D-deoxyribose 1-phosphate
-
2'-deoxyinosine and 5'-deoxyinosine
-
-
r
deoxyinosine + phosphate
hypoxanthine + alpha-D-deoxyribose 1-phosphate
-
88% of the activity with deoxyguanosine
-
-
r
guanosine + phosphate
guanine + alpha-D-ribose 1-phosphate
-
-
-
r
guanosine + phosphate
guanine + alpha-D-ribose 1-phosphate
-
48% of the activity with deoxyguanosine
-
-
?
guanosine + phosphate
guanine + alpha-D-ribose 1-phosphate
no activity with mutant enzyme N239D
-
-
?
guanosine + phosphate
guanine + alpha-D-ribose 1-phosphate
-
66% of the activity with inosine
-
-
?
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
-
?
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
-
r
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
?
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
r
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
46% of the activity with deoxyguanosine
-
r
xanthosine + phosphate
alpha-D-ribose 1-phosphate + xanthine
no activity with mutant enzymes Y191L and N239D
-
-
?
additional information
?
-
catalysis involves protonation of the purine base at position N7 by residue Asp204, which is triggered by Arg217
-
-
?
additional information
?
-
-
catalysis involves protonation of the purine base at position N7 by residue Asp204, which is triggered by Arg217
-
-
?
additional information
?
-
no substrate: xanthosine, uridine, 2'-deoxythymidine, cytidine
-
-
?
additional information
?
-
-
no substrate: xanthosine, uridine, 2'-deoxythymidine, cytidine
-
-
?
additional information
?
-
N1-methylated guanosine, inosine, and adenosine derivatives are selective substrates for hexameric PNP from Escherichia coli. Hexameric PNPs accept as substrates many nucleoside analogues with various modifications in the purine ring, with an important exception of 7-deaza nucleosides. N7-Methylated guanosine, inosine and adenosine are unusual fluorescent substrates of both trimeric and hexameric PNPs. Escherichia coli PNP converts pro-drugs, which are relative nontoxic purine nucleosides (for example 6-methyl purine 2'-deoxynucleoside or fludarabine), to their respective purine analogs (here, to 6-methylpurine and 2-fluoroadenine, respectively), which are very potent, toxic drugs. Enzymatic ribosylation of 8-azapurines, overview. Isoadenosine (3-beta-D-ribosyl-adenine), is, unlike the parent adenosine, a quite good substrate for mammalian and bacterial PNP. Ribosylation of tri-cyclic nucleobase analogues and phosphorolysis
-
-
-
additional information
?
-
N1-methylated guanosine, inosine, and adenosine derivatives are selective substrates for hexameric PNP from Escherichia coli. Hexameric PNPs accept as substrates many nucleoside analogues with various modifications in the purine ring, with an important exception of 7-deaza nucleosides. N7-Methylated guanosine, inosine and adenosine are unusual fluorescent substrates of both trimeric and hexameric PNPs. Escherichia coli PNP converts pro-drugs, which are relative nontoxic purine nucleosides (for example 6-methyl purine 2'-deoxynucleoside or fludarabine), to their respective purine analogs (here, to 6-methylpurine and 2-fluoroadenine, respectively), which are very potent, toxic drugs. Enzymatic ribosylation of 8-azapurines, overview. Isoadenosine (3-beta-D-ribosyl-adenine), is, unlike the parent adenosine, a quite good substrate for mammalian and bacterial PNP. Ribosylation of tri-cyclic nucleobase analogues and phosphorolysis
-
-
-
additional information
?
-
N7-D-ribosyl-2,6-diamino-8-azapurine is a good fluorogenic substrate for mammalian forms of PNP, including human blood PNP, while the N8-riboside is selective to the Escherichia coli enzyme. Wild-type calf PNP gives N7- and N8-ribosides, while the calf enzyme N243D PNP mutant directs the ribosyl substitution at N9- and N7-positions. The Escherichia coli enzyme ribosylates adenine and 8-azaadenine. In both cases, the N9-ribosides are the only detectable ribosylation products for the wild-type enzyme, while the D204N mutant reacting with the latter substrate does produce the N8-riboside. The altered specificity of the PNP mutants towards some of the purine analogues can be related to differences in binding modes of 8-azapurine bases in the PNP active site, including binding in the upside-down position (rotation by 180 degree when compared with standard binding mode). Ribosylation of 2,6-diamino-8-azapurine leads to at least three fluorescent ribosides. The N9-riboside is very highly fluorescent, but its fluorescence is generally similar, in terms of lambdamax and decay time, to that of the free base. By contrast, fluorescence of N7- and N8-ribosides is red-shifted by about 70 nm and thanks to this shift their cleavage via phosphorolysis (or hydrolysis) leads to a high fluorogenic effect at about 360 nm, which can be analytically useful
-
-
-
additional information
?
-
-
N7-D-ribosyl-2,6-diamino-8-azapurine is a good fluorogenic substrate for mammalian forms of PNP, including human blood PNP, while the N8-riboside is selective to the Escherichia coli enzyme. Wild-type calf PNP gives N7- and N8-ribosides, while the calf enzyme N243D PNP mutant directs the ribosyl substitution at N9- and N7-positions. The Escherichia coli enzyme ribosylates adenine and 8-azaadenine. In both cases, the N9-ribosides are the only detectable ribosylation products for the wild-type enzyme, while the D204N mutant reacting with the latter substrate does produce the N8-riboside. The altered specificity of the PNP mutants towards some of the purine analogues can be related to differences in binding modes of 8-azapurine bases in the PNP active site, including binding in the upside-down position (rotation by 180 degree when compared with standard binding mode). Ribosylation of 2,6-diamino-8-azapurine leads to at least three fluorescent ribosides. The N9-riboside is very highly fluorescent, but its fluorescence is generally similar, in terms of lambdamax and decay time, to that of the free base. By contrast, fluorescence of N7- and N8-ribosides is red-shifted by about 70 nm and thanks to this shift their cleavage via phosphorolysis (or hydrolysis) leads to a high fluorogenic effect at about 360 nm, which can be analytically useful
-
-
-
additional information
?
-
-
enzyme shows a high tolerance to the steric and hydrophobic environment at the 6-position of the synthesized purine ribonucleosides
-
-
?
additional information
?
-
etheno-derivatives of guanine, O6-methylguanine, and isoguanine are prepared and examined as potential substrates of purine-nucleoside phosphorylase in the reverse (synthetic) pathway. 1,N2-ethenoguanine and 1,N6-etheno-isoguanine are excellent substrates for purine-nucleoside phosphorylase (PNP) from Escherichia coli, while O6-methyl-N2,3-ethenoguanine exhibits moderate activity with this enzyme. The ribosylation reaction is fully reversible by the addition of a phosphate buffer up to a concentration of about 5 mM. The inability of the enzyme to ribosylate N2,3-ethenoguanine is not due to geometric hindrance, but rather should be ascribed to unfavorable energetic factors, in accordance with previous reports about the relative instability of the glycosidic bond in this compound. N2,3-etheno-O6-methylguanine is a poor substrate, it competitively competes with guanine in the ribosylation process, acting as quasi inhibitor of the Escherichia coli PNP. 6-Aminopurine ribosides, not possessing a mobile proton at the N1 position, do not react with trimeric forms of PNP
-
-
-
additional information
?
-
-
etheno-derivatives of guanine, O6-methylguanine, and isoguanine are prepared and examined as potential substrates of purine-nucleoside phosphorylase in the reverse (synthetic) pathway. 1,N2-ethenoguanine and 1,N6-etheno-isoguanine are excellent substrates for purine-nucleoside phosphorylase (PNP) from Escherichia coli, while O6-methyl-N2,3-ethenoguanine exhibits moderate activity with this enzyme. The ribosylation reaction is fully reversible by the addition of a phosphate buffer up to a concentration of about 5 mM. The inability of the enzyme to ribosylate N2,3-ethenoguanine is not due to geometric hindrance, but rather should be ascribed to unfavorable energetic factors, in accordance with previous reports about the relative instability of the glycosidic bond in this compound. N2,3-etheno-O6-methylguanine is a poor substrate, it competitively competes with guanine in the ribosylation process, acting as quasi inhibitor of the Escherichia coli PNP. 6-Aminopurine ribosides, not possessing a mobile proton at the N1 position, do not react with trimeric forms of PNP
-
-
-
additional information
?
-
hexameric PNPs accept as substrates many nucleoside analogues with various modifications in the purine ring, with an important exception of 7-deaza nucleosides. N7-Methylated guanosine, inosine and adenosine are unusual fluorescent substrates of both trimeric and hexameric PNPs
-
-
-
additional information
?
-
hexameric PNPs accept as substrates many nucleoside analogues with various modifications in the purine ring, with an important exception of 7-deaza nucleosides. N7-Methylated guanosine, inosine and adenosine are unusual fluorescent substrates of both trimeric and hexameric PNPs
-
-
-
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
1-methyladenosine + phosphate
1-methyladenine + alpha-D-ribose 1-phosphate
-
-
-
r
1-methylguanosine + phosphate
1-methylguanine + alpha-D-ribose 1-phosphate
-
-
-
r
6-mercaptopurine-2'-deoxyriboside + phosphate
6-mercaptopurine + 2-deoxy-alpha-D-ribose 1-phosphate
-
-
-
r
8-azaguanine + alpha-D-ribose 1-phosphate
8-azaguanosine + phosphate
-
-
-
r
8-azaguanosine + phosphate
8-azaguanine + alpha-D-ribose 1-phosphate
-
-
-
r
9-beta-D-arabinosyl-2-fluoroadenine + phosphate
2-fluoroadenine + beta-D-arabinose 1-phosphate
i.e. fludarabine
-
-
r
9-[2-deoxy-beta-D-ribofuranosyl]-6-methylpurine + phosphate
?
-
a toxic prodrug
-
-
?
9-[6-deoxy-beta-L-talofuranosyl]-2-fluoro-adenine + phosphate
?
-
a toxic prodrug, substrate of enzyme mutant M64V, the drug product has anti-tumor activity
-
-
?
9-[6-deoxy-beta-L-talofuranosyl]-6-methylpurine + phosphate
?
-
a toxic prodrug, substrate of enzyme mutant M64V, the drug product has anti-tumor activity
-
-
?
9-[beta-D-arabinofuranosyl]-2-fluoro-adenine + phosphate
?
-
a toxic prodrug
-
-
?
9-[beta-L-lyxofuranosyl]-2-fluoro-adenine + phosphate
?
-
a toxic prodrug, substrate of enzyme mutant M64V, the drug product has anti-tumor activity
-
-
?
guanosine + phosphate
guanine + alpha-D-ribose 1-phosphate
-
-
-
r
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
r
nicotinamide riboside + phosphate
nicotinamide + alpha-D-ribose 1-phosphate
-
-
-
r
purine deoxyribonucleoside + phosphate
purine + 2-deoxy-alpha-D-ribose 1-phosphate
-
-
-
-
?
purine ribonucleoside + phosphate
purine + alpha-D-ribose 1-phosphate
-
-
-
-
?
xanthosine + phosphate
xanthine + alpha-D-ribose 1-phosphate
-
-
-
r
adenosine + phosphate
adenine + alpha-D-ribose 1-phosphate
-
-
-
r
guanosine + phosphate
guanine + alpha-D-ribose 1-phosphate
-
-
-
r
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
r
inosine + phosphate
hypoxanthine + alpha-D-ribose 1-phosphate
-
-
-
r
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
acyclovir
ACV, an acyclic derivative of the PNP substrate guanosine, an acyclic analogue of 2'-deoxyguanosine, that is used as an antiviral drug for the treatment of some human viral infections. The ACV molecule occupies the nucleoside-binding pocket in the active-site cavity, where it adopts two conformations. Sulfate ions are located in both the nucleoside-binding and phosphate-binding pockets of the enzyme. Binding structure, overview
N(6)-Methylformycin A
-
competitive with respect to inosine, 7-methylguanosine and 7-methyladenosine
N2,3-etheno-O6-methylguanine
is a poor substrate, it competitively competes with guanine in the ribosylation process, acting as quasi inhibitor of the Escherichia coli PNP
N2,3-ethenoguanine
exhibits moderate, possibly competitive inhibition of Escherichia coli PNP
Formycin A
a potent inhibitor of hexameric PNPs. Inhibitor-enzyme interaction and kinetic analysis with wild-type and mutant PNPs, detailed overview. With the wild-type enzyme, in the P8.3 at 10°C a model of binding one molecule per enzyme hexamer gives the best fit and at 25°C all types of fits are comparable. The strongest association is observed in the phosphate buffer pH 7.0. On the other hand, results show that the presence of phosphate at pH 8.3 is responsible for a strong binding impairment (Kd increase). The behaviour of the Kapp calculated for the PNPY-FA complexes corresponds to the Kapp of the PNPWT-FA. Their values suggest stronger than in the PNPWT-FA complexes association in the P7 and P8.3 at 10°C and in H8.3, P7 and P8.3 at 25°C
additional information
formycins are 9-deaza-8-aza-nucleosides and selective inhibitors of hexameric PNPs. 8-Aza-9-deazapurine derivatives as enzyme inhibitors, overview
-
additional information
formycins are 9-deaza-8-aza-nucleosides and selective inhibitors of hexameric PNPs. 8-Aza-9-deazapurine derivatives as enzyme inhibitors, overview
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.2
2,6-diamino-8-azapurine
above, pH 6.5, 25°C, recombinant wild-type enzyme and mutant D204N
0.126
9-beta-D-(2-deoxyribofuranosyl)-6-methylthiopurine
pH 7.4, 25°C
0.08
N7-D-ribosyl-2,6-diamino-8-azapurine
about, pH 6.5, 25°C, recombinant wild-type enzyme
0.007
N8-D-ribosyl-2,6-diamino-8-azapurine
pH 6.5, 25°C, recombinant wild-type enzyme
0.02
N9-D-ribosyl-2,6-diamino-8-azapurine
about, pH 6.5, 25°C, recombinant wild-type enzyme
additional information
additional information
steady-state kinetics and kinetic analysis
-
additional information
additional information
-
steady-state kinetics and kinetic analysis
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0003
6-methylformycin A
pH and temperature not specified in the publication
0.005
formycin B
pH and temperature not specified in the publication
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
0.07
0.09
0.36
0.015
-
wild-type enzyme, substrate 9-[6-deoxy-beta-L-talofuranosyl]-6-methylpurine
0.0167
-
mutant M64V, substrate 9-[beta-D-arabinofuranosyl]-2-fluoro-adenine
0.02
-
wild-type enzyme, substrate 9-[beta-D-arabinofuranosyl]-2-fluoro-adenine
0.13
-
wild-type enzyme, substrate 9-[6-deoxy-beta-L-talofuranosyl]-2-fluoro-adenine
0.567
-
wild-type enzyme, substrate 9-[2-deoxy-beta-D-ribofuranosyl]-6-methylpurine
1.43
-
mutant M64V, substrate 9-[6-deoxy-beta-L-talofuranosyl]-6-methylpurine
23.3
-
mutant M64V, substrate 9-[2-deoxy-beta-D-ribofuranosyl]-6-methylpurine
4
-
mutant M64V, substrate 9-[6-deoxy-beta-L-talofuranosyl]-2-fluoro-adenine
8.83
-
wild-type enzyme, substrate 9-[2-deoxy-beta-D-ribofuranosyl]-6-methylpurine
9.83
-
mutant M64V, substrate 9-[2-deoxy-beta-D-ribofuranosyl]-6-methylpurine
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6.8
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
25 - 70
-
25°C: about 40% of maximal activity, 70°C: about 60% of maximal activity
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
evolution
malfunction
substrate specificity of trimeric and hexameric PNPs may be changed by mutations of the crucial active site amino acids, namely Asp in hexameric PNPs and Asn in trimeric PNPs
physiological function
evolution
in some organisms, like Escherichia coli, two distinct forms of PNP exist, with markedly different structure and substrate specificity. The second form, the so-called Escherichia coli PNP-II, is sometimes referred to as xanthosine phosphorylase, since it is inducible by this nucleoside, but its specificity is not limited to this compound, and includes guanosine, inosine and nicotinamide riboside
malfunction
substrate specificity of trimeric and hexameric PNPs may be changed by mutations of the crucial active site amino acids, namely Asp in hexameric PNPs and Asn in trimeric PNPs
additional information
evolution
in some organisms, like Escherichia coli, two distinct forms of PNP exist, with markedly different structure and substrate specificity. The second form, the so-called E. coli PNP-II, is sometimes referred to as xanthosine phosphorylase, since it is inducible by this nucleoside, but its specificity is not limited to this compound, and includes guanosine, inosine and nicotinamide riboside
evolution
the enzyme belongs to the type I PNP family of hexameric enzymes
physiological function
Escherichia coli PNP converts pro-drugs, which are relative nontoxic purine nucleosides (for example 6-methyl purine 2'-deoxynucleoside or fludarabine), to their respective purine analogues (here, to 6-methylpurine and 2-fluoroadenine, respectively), which are very potent, toxic drugs
physiological function
PNP catalyzes a reversible phosphorolysis of the N-glycosidic bond in natural purine nucleosides, as well as inmany purine analogs, some of them displaying marked biological and pharmacological activity
physiological function
purine nucleoside phosphorylases (PNPs) catalyze the reversible phosphorolysis of nucleosides and are key enzymes involved in nucleotide metabolism. They are essential for normal cell function and can catalyze the transglycosylation
physiological function
the enzyme catalyzes the reversible phosphorolysis of purine ribonucleosides
additional information
the enzyme-catalyzed reaction involves the proton transfer from the carboxyl group of Asp204 to the nitrogen atom N7 of the purine base. This process is accompanied by conformational changes. The C-terminal alpha-helix (214-236) is divided by a gamma-turn into two helices (214-219 and 223-236), resulting in the residue Arg217 coming closer to Asp204, and the active site adopts a closed conformation. Upon phosphate binding, the position of the residue Arg24 is fixed via a hydrogen bond with Arg217 (NE_Arg2-O_Arg217). The key role of the residues Asp204, Arg217, and Arg24 in the catalytic activity is confirmed by the replacement of these residues with alanine. These replacements lead to a substantial decrease in the activity toward natural substrates
additional information
possible tautomeric forms of 1,N6-etheno-isoguanine. The main reason for slow ribosylation and the resistance of the parent N2,3-ethenoguanine to the enzymatic ribosylation is probably unfavorable energetics
additional information
-
possible tautomeric forms of 1,N6-etheno-isoguanine. The main reason for slow ribosylation and the resistance of the parent N2,3-ethenoguanine to the enzymatic ribosylation is probably unfavorable energetics
PDB
SCOP
CATH
UNIPROT
ORGANISM
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
homohexamer
hexamer
homohexamer
homohexamer
analysis of amino-acid sequence of PNP with the secondary-structure elements of the monomers, enzyme crystals are used for the determination of the three-dimensional structure of Escherichia coli PNP. Each of the six active sites of the enzyme is located in a deep cleft between the subunits of the dimer and is formed with the participation of the residues of the adjacent subunit. The cleft is bounded by two beta-ribbons (residues 84-93 and 177-179) and residues of irregular loops (loops 18-22, 155-164, 179-182, and 202-214) of one subunit and fragments of loops (1-79 and 40-47) of the adjacent subunit of the dimer, the side groups of the residues Arg43* and His4* of the latter subunit being located in the active site cavity. Comparison of subunits
homohexamer
the hexameric molecule can be considered as a trimer of dimers, where each dimer contains two complete active sites. Each monomer consists of a central core comprising mixed beta-sheet surrounded by alpha-helices with an extended C-terminal alpha7 helix. The active sites are located within the deep cavity between the dimer subunits and comprise amino-acid residues from both subunits. The active-site cavity is predominantly lined by hydrophobic amino acids from strands beta5, beta7, beta8, beta9, and beta10 and helix alpha7. The adjacent dimer subunit contributes the side chains of His4* and Arg43* to the active site
hexamer
-
irrespective of protein concentration, ionic strength, temperature, presence of substrates, or destabilizing factors, the only active enzyme form is a hexamer
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
crystal structures of the wild type in complexes with phosphate and sulfate, respectively, and of the R24A mutant in complex with phosphate/sulfate, to about 2 A resolution. The structural data show that previously observed conformational change is a result of the phosphate binding and its interaction with residue Arg24
hanging drop method, crystal structure of the ternary complex of the hexameric enzyme with formycin A derivatives and phosphate or sulfate ions is determined at 2.0 A resolution
hanging drop vapor diffusion method. Enzyme in complex with nucleoside analogs: 2-fluoroadenosine, 9-beta-D-ribofuranosyl-6-methylthiopurine, 2-fluoro-2'-deoxyadenosine, 9-beta-D-(2-deoxyribofuranosyl)-6-methylthiopurine, adenosine, formycin B, 7-deazaadenosine, 9-beta-D-arabinofuranosyl-adenine, 9-beta-D-xylofuranosyl-adenine or inosine
purified enzyme PNP in complex with inhibitor ACV, liquid diffusion method, mixing of 0.0035 ml of 32 mg/ml protein in 0.02 M Tris-HCl, pH 7.5, with 0.0035 ml of reservoir solution containing 25% ammonium sulfate, 0.05 M sodium citrate, pH 4.9, 0.04% sodium azide, and 5 mM acycloguanosine, and equilbration against 0.18 ml of reservoir solution, X-ray diffraction structure determination at 2.32 A resolution using the molecular replacement method. The ACV molecule is observed in two conformations and sulfate ions are located in both the nucleoside-binding and phosphate-binding pockets of the enzyme
purified recombinant enzyme, crystals are grown in microgravity by the capillary counterdiffusion method through a gel layer, X-ray diffraction structure determination and analysis at 0.99 A resolution, molecular replacement using the structure of the hexameric Escherichia coli enzyme, PDB ID 1ECP, as template
ternary complex of enzyme with a phosphate ion and formycin A, to 0.975 A resolution. The structure reveals, in some active sites, an unexpected binding site for phosphate and exhibits a stoichiometry of two phosphate molecules per enzyme subunit. In these active sites, the phosphate and nucleoside molecules are found not to be in direct contact but being bridged by three water molecules
complex of enzyme with hypoxanthine at 2.15 A resolution, structural data from crystal structure may be useful in designing prodrugs that can be activated by Escherichia coli enzyme but not the human enzyme
-
crystal structures of the wild-type and a C-terminal KH/S1 domain-truncated mutant of PNPase at resolutions of 2.6 A and 2.8 A, respectively
-
crystals for X-ray analysis are obtained by vapor-diffusion equilibration of droplets hanging from siliconized coverslips inverted on Linbro plates
-
two crystal forms in presence of guanine and phosphate and a third crystal form in presence of xanthine and phosphate, crystals are grown at pH values between 8 and 9, crystals can not be grown at physiologic pH, no good crystals can be obtained in presence of xanthosine, guanosine or ribose 1-phosphate
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
D204A
0.1% of wild-type activity towards inosine, adenosine and guanosine
D204A/R217A
about 0.2% of wild-type activity towards inosine, adenosine and guanosine
D204N
F159A
site-directed mutagenesis, the PNPF159A-FA complexes show a weak association of formycin A to the mutant's active center
F159Y
site-directed mutagenesis, a prominent quenching of the PNPF159Y emission indicates a complex formation, with the strongest association in the phosphate buffer, pH 7.0, relative to the wild-type enzyme. On the other hand, results testify to a deterioration of the interactions in the wild-type PNP/PNPF159Y mutant and formycin A complexes in the presence of the phosphate, pH 8.3
R117E/K121E/D139R
the mutant shows greater stability compared with the wild type dimer and changes in its structures compared with the wild type dimer in a hexamer
R117E/K121E/D139R/F120D
the mutant shows greater stability compared with the wild type dimer and changes in its structures compared with the wild type dimer in a hexamer
R117E/K121E/D139R/F120D/I128S/F131G
the mutant shows greater stability compared with the wild type dimer and changes in its structures compared with the wild type dimer in a hexamer
R117E/K121E/D139R/Y173S
the mutant shows greater stability compared with the wild type dimer and changes in its structures compared with the wild type dimer in a hexamer
R217A
about 0.5% of wild-type activity towards inosine and guanosine, 10.8% of activity towards adenosine
R24A
absence of a conformational change upon binding of phosphate, about 0.2 to 0.6% of wild-type activity towards inosine, adenosine and guanosine
D204N
site-directed mutagenesis, the enzymatic ribosylation of 1,N6-etheno-isoguanine using Escherichia coli PNP wild-type and D204N mutant enzymes gives different products, which are identified on the basis of NMR analysis and comparison with the product of the isoguanosine reaction with chloroacetic aldehyde, which gives an essentially single compound, identified unequivocally as N9-riboside
M64V
N239D
mutant enzyme shows no activity with the wild-type substrates inosine, xanthosine and guanosine. Unlike the wild-type enzyme, the mutant enzyme shows activity with adenosine
Y191L
mutant enzyme shows no activity with the wild-type substrate xanthosine. The ratio of Vmax to Km for inosine as substrate is 1.7fold lower than the ratio for wild-type enzyme
additional information
-
the C-terminal KH/S1 domain-truncated mutant binds and cleaves RNA less efficiently with an eightfold reduced binding affinity. The mutant forms a less stable trimer than the full-length PNPase. The crystal structure of DeltaKH/S1 is more expanded, containing a slightly wider central channel than that of the wild-type PNPase, suggesting that the KH/S1 domain helps PNPase to assemble into a more compact trimer, and it regulates the channel size allosterically
D204N
less than about 4% of wild-type activity towards inosine, adenosine and guanosine
D204N
site-directed mutagenesis, the mutant exhibits altered substrate specificity compared to wild-type
M64V
-
mutant enzyme is able to cleave numerous 5-modified nucleoside analogs with much greater efficiency than the wild-type enzyme
M64V
-
naturally occuring mutant, the mutant enzyme is able to cleave numerous 5-modified nucleoside analogs with much greater efficiency than the wild-type enzyme
M64V
-
the mutant is able to cleave numerous 5'-modified nucleoside analogs with much greater efficiency than the wild type enzyme but no activity with adenosine or inosine
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
45
-
half-life at pH 6-7: 8-9 min. Half-life at pH 7-8: 5-7 min. At pH 6.5 in presence of 5 mg/ml bovine serum albumin half-life is 17 min
50
50
-
5 min, 75-85% loss of activity, 1 mM hypoxanthine significantly protects against thermal denaturation
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant enzyme from Escherichia coli strain BL21(DE3)/pERPUPHOI cell extract supernatant by anion exchange chromatography, ultrafiltration, and gel filtration
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression in Escherichia coli
gene pnp, sequence comparisons of Escherichia coli enzyme with human and Bacillus subtilis enzymes, recombinant expression in Escherichia coli strain BL21(DE3)/pERPUPHOI
expression of the Escherichia coli purine nucleoside phosphorylase/9-(2-deoxy-beta-dribofuranosyl)-6-methylpurine suicide gene system in human hepatocarcinoma cells
-
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
Escherichia coli PNP-II (xanthosine phosphorylase) is inducible by xanthosine
APPLICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
drug development
pharmacology
substrate 6-mercaptopurine-2'-deoxyriboside is of special interest, because, in contrast to a nucleoside, its parent purine is highly cytotoxic and is known as one of the first compounds applied as anti-cancer drugs
synthesis
drug development
differences in specificity between homotrimeric (including human enzyme) and homohexameric PNPs, including various pathogenic organisms, make them interesting potential drug targets
medicine
synthesis
important application of PNP is in chemo-enzymatic synthesis of bioactive nucleoside analogues, utilizing various types of PNP among others. This application may be extended to tricyclic nucleobase analogues, particularly to adenine, isoguanine, and guanine derivatives. In particular, N9-D-riboside of can be obtained quantitatively from 1,N2-ethenoguanine, and N9-beta-D- and N7-beta-D-ribosides of 1,N6-etheno-isoguanine as a mixture, using the Escherichia coli PNP as a biocatalyst
drug development
differences in specificity between homotrimeric (including human enzyme) and homohexameric PNPs, including various pathogenic organisms, make them interesting potential drug targets
drug development
owing to their key role in the purine salvage pathway, PNPs are attractive targets for drug design against some pathogens
synthesis
use of agar from four different polymer carriers as a suitable matrix for whole recombinant cell entrapment. Although the immobilization process reduces the substrate affinity and catalytic efficiency of recombinant cells, it can significantly enhance the stability and reusability of these cells. The immobilized whole cell biocatalyst is applied to produce ribavirin, as a model nucleoside synthesis reaction showing relative high productivity of rabavirin and quick reaction time
synthesis
enzymatic ribosylation of fluorescent 8-azapurine derivatives, like 8-azaguanine and 2,6-diamino-8-azapurine, with purine-nucleoside phosphorylase (PNP) as a catalyst, leads to N9, N8, and N7-ribosides. The final proportion of the products may be modulated by point mutations in the enzyme active site. As an example, ribosylation of the latter substrate by wild-type calf PNP gives N7- and N8-ribosides, while the N243D mutant directs the ribosyl substitution at N9- and N7-positions. The same mutant allows synthesis of the fluorescent N7-D-ribosyl-8-azaguanine. The mutated form of the Escherichia coli PNP, D204N, can be utilized to obtain non-typical ribosides of 8-azaadenine and 2,6-diamino-8-azapurine as well. The N7- and N8-ribosides of the 8-azapurines can be analytically useful, as illustrated by N7-D-ribosyl-2,6-diamino-8-azapurine, which is a good fluorogenic substrate for mammalian forms of PNP, including human blood PNP, while the N8-riboside is selective to the Escherichia coli enzyme
medicine
-
structural data from crystal structure may be useful in designing prodrugs that can be activated by E. coli enzyme but not the human enzyme
medicine
-
delivery of replication-competent retrovirus expressing Escherichia coli purine nucleoside phosphorylase increases the metabolism of the prodrug, fludarabine phosphate and suppresses the growth of bladder tumor xenografts. Replication-competent retroviral vectors are a potentially efficient gene delivery method and replication-competent retroviral vector mediated PNP gene transfer and fludarabine phosphate treatment might be a novel and potentially therapeutic modality for bladder cancer
medicine
-
expression of the Escherichia coli purine nucleoside phosphorylase/9-(2-deoxy-beta-dribofuranosyl)-6-methylpurine suicide gene system in human hepatocarcinoma cells together with a chimeric human alpha-fetoprotein promoter. In the presence of hypoxia, the Escherichia coli purine nucleoside phosphorylase gene directed by the human promoter is specifically expressed in two human hepatocarcinoma cell lines and, the therapy yields significant and selective cytotoxicity in both alpha-fetoprotein-positive and low-alpha-fetoprotein-generating hepatocarcinoma cells
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Parks, R.E.; Agarwal, R.P.
Purine nucleoside phosphorylase
The Enzymes, 3rd Ed. (Boyer, P. D. , ed. )
7
483-514
1972
Klebsiella aerogenes, Aeromonas hydrophila, Brevibacillus brevis, Bacillus cereus, Bacillus subtilis, Bacillus licheniformis, Bacterium cadaveris, Bos taurus, Canis lupus familiaris, Gallus gallus, Columba livia, Clavibacter michiganensis subsp. sepedonicus, Oryctolagus cuniculus, Escherichia coli, Equus caballus, Pectobacterium carotovorum, Felis catus, Homo sapiens, Lactobacillus leichmannii, Leucisus rusticus, Macaca mulatta, Micrococcus luteus, Papio hamadryas, Proteus vulgaris, Shewanella putrefaciens, Rattus norvegicus, salmon, Salmonella enterica subsp. enterica serovar Enteritidis, Sus scrofa
-
Cook, W.J.; Ealick, S.E.; Krenitsky, T.A.; Stoeckler, J.D.; Helliwell, J.R.; Bugg, C.E.
Crystallization and preliminary x-ray investigation of purine-nucleoside phosphorylase from Escherichia coli
J. Biol. Chem.
260
12968-12969
1985
Escherichia coli
Bzowska, A.; Kulikowska, E.; Shugar, D.
Formycins A and B and some analogues: selective inhibitors of bacterial (Escherichia coli) purine nucleoside phosphorylase
Biochim. Biophys. Acta
1120
239-247
1992
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Crystal structure of Escherichia coli purine nucleoside phosphorylase complexed with acyclovir
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Escherichia coli (P0ABP8), Escherichia coli
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Escherichia coli (P0ABP8), Escherichia coli
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1,N6-ethenoadenine and other fluorescent nucleobase analogs as substrates for purine-nucleoside phosphorylases Spectroscopic and kinetic studies
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Bacillus cereus, Pectobacterium carotovorum, Thermus thermophilus, Plasmodium lophurae, Helicobacter pylori (A0A518Y5Z2), Homo sapiens (P00941), Escherichia coli (P0ABP8), Escherichia coli (P45563), Bos taurus (P55859), Cellulomonas sp. (P81989), Plasmodium falciparum (Q8I3X4)
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Tri-cyclic nucleobase analogs and their ribosides as substrates of purine-nucleoside phosphorylases II guanine and isoguanine derivatives
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Escherichia coli (P45563), Escherichia coli
Stachelska-Wierzchowska, A.; Wierzchowski, J.; Bzowska, A.; Wielgus-Kutrowska, B.
Tricyclic nitrogen base 1,N6-ethenoadenine and its ribosides as substrates for purine-nucleoside phosphorylases spectroscopic and kinetic studies
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Escherichia coli (P0ABP8), Escherichia coli
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