Synthesis of 3′-α-fluoronucleosides Using Pyrimidine Nucleoside Phosphorylase of Thermus thermophilus and Purine Nucleoside Phosphorylase of Escherichia coli

2015 ◽  
pp. 191-200
Keyword(s):  
Escherichia Coli ◽  
Purine Nucleoside Phosphorylase ◽  
Thermus Thermophilus ◽  
Purine Nucleoside ◽  
Pyrimidine Nucleoside ◽  
Nucleoside Phosphorylase ◽  
Pyrimidine Nucleoside Phosphorylase

FISH-MUSCLE PURINE AND PYRIMIDINE NUCLEOSIDE PHOSPHORYLASES

1967 ◽  
Vol 45 (3) ◽  
pp. 409-419 ◽  
Author(s):  
H. L. A. Tarr ◽  
Joan E. Roy
Keyword(s):  
Enzyme Preparation ◽  
Purine Nucleoside Phosphorylase ◽  
Deae Cellulose ◽  
Fish Muscle ◽  
Purine Nucleoside ◽  
Aqueous Extracts ◽  
Pyrimidine Nucleoside ◽  
Nucleoside Phosphorylase ◽  
Pyrimidine Nucleoside Phosphorylase ◽  
Nucleoside Phosphorylases

Three purine nucleoside phosphorylase preparations (isoenzymes) were obtained by ammonium sulfate fractionation and DEAE-cellulose chromatography of aqueous extracts of lingcod muscle. Dialysis, adsorption on alumina Cγ, and elution with 0.4 M phosphate buffer yielded further purification. The most active enzyme preparation had about 120 times the activity of initial extracts. It utilized hypoxanthine, 6-mercaptopurine, guanine, 8-azaguanine, xanthine, adenine, 2,6-diaminopurine and 6-methylpurine in presence of ribose 1-phosphate or deoxyribose 1-phosphate. Several substituted purines were not utilized and did not inhibit the reaction between hypoxanthine and the pentose phosphates. The Kmwith inosine as substrate was 3.2 × 10−6 M. A pyrimidine nucleoside phosphorylase, distinct from the purine nucleoside phosphorylase, occurred in the DEAE-cellulose fraction comprising one of the purine nucleoside phosphorylases. Its activity was much lower than that of the purine nucleoside phosphorylase preparations. Uridine and thymidine were the best substrates. Deoxyuridine was a poor substrate, and neither cytidine nor deoxycytidine was utilized. The equilibrium with all preparations was about 80% in favor of nucleoside formation. The purified enzymes were all destroyed by freezing.

scholarly journals The Peculiar Case of the Hyperthermostable Pyrimidine Nucleoside Phosphorylase from Thermus Thermophilus

2020 ◽  
Author(s):  
Felix Kaspar ◽  
Peter Neubauer ◽  
Anke Kurreck
Keyword(s):  
Thermus Thermophilus ◽  
Pyrimidine Nucleoside ◽  
Reaction Conditions ◽  
Nucleoside Phosphorylase ◽  
Pyrimidine Nucleosides ◽  
Low Concentrations ◽  
Pyrimidine Nucleoside Phosphorylase ◽  
Total Turnover ◽  
Nucleoside Phosphorylases ◽  
Peculiar Case

The poor solubility of many nucleoside and nucleobases in aqueous solution demands harsh reaction conditions (base, heat, cosolvent) in nucleoside phosphorylase-catalyzed processes to facilitate substrate loading beyond the low millimolar range. This, in turn, requires enzymes which withstand these conditions. Herein we report that the pyrimidine nucleoside phosphorylase from <i>Thermus thermophilus</i> is active over an exceptionally broad pH (4-10), temperature (up to 100 °C) and cosolvent space (up to 80% (v/v) non-aqueous medium) and displays tremendous stability under harsh reaction conditions with predicted total turnover numbers of more than 10<sup>6</sup> for various pyrimidine nucleosides. However, its use as a biocatalyst for preparative applications is critically limited due to its inhibition by nucleoside substrates at low concentrations, which is unprecedented among non-specific pyrimidine nucleoside phosphorylases.<br>

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