#K-NP-0002

The Nucleoside Phosphorylase Screening Kit was designed to identify the most suitable nucleoside phosphorylases for the production of nucleoside analogues in a transglycosylation reaction. The Kit contains 4 pyrimidine and 3 purine nucleoside phosphorylases from thermophilic organisms with different temperature optima and substrate spectra. The Nucleoside Phosphorylase Screening Kit is designed for screening in small scale: 40 reactions in 100 µl scale can be performed. The conversion of appr. 10 mg of nucleoside is possible with the Nucleoside Phosphorylase Screening Kit.

Kit components:

Enzymes:   4 pyrimidine nucleoside phosphorylases (Y01-Y04), 3 purine   nucleoside phosphorylases (N01-N03) - 1 mL each of 0.2 mg/mL

Buffers:  6 mL Phosporolysis buffer, 6 mL Transglycosylation buffer

Positive Controls:  3 mg Adenosine, 3 mg Uridine, 3 mg Adenine, 3 mg Uracil

Description:

Nucleoside analogues are a potential class of cytotoxic, antiviral and antiprotozoal drugs. Nucleoside analogs have been in clinical use for almost 50 years and are cornerstones of treatment for patients with cancer or viral infections. Modified nucleosides are currently produced through a long and tedious chemical process that limits the production of new compounds. In contrast, the enzymatic synthesis of nucleosides by the transglycosylation reaction proceeds with strict stereo- and regioselectivity and requires organic solvents (ethanol, acetonitrile) only for the isolation of the individual desired compound. The production process becomes sustainable and products with a purity >99% are achieved. Production time and costs are drastically reduced.

Biochemical action of nucleoside phosphorylases:

Nucleoside phosphorylases catalyze the following reaction:

Nucleoside + Pi ↔ Sugar-1-phosphate + base

Unit definition:

One unit of pyrimidine nucleoside phosphorylase will cause the phosphorolysis of 1.0 μmole of uridine to uracil and ribose 1-phosphate per min at pH 7.4 at optimum temperature. One unit of purine nucleoside phosphorylase will cause the phosphorolysis of 1.0 μmole of adenosine to adenine and ribose 1-phosphate per min at pH 7.4 at optimum temperature.

Transglycosylationreaction:

Temperature optima and preferences of BioNukleos nucleoside phosphorylases:

Product use limitations:

This product is developed and sold exclusively for research purposes and in vitro use only. The product was not tested for use in diagnostics or drug development, nor is suitable for administration to humans or animals.

#K-NP-0003

The Pyrimidine nucleoside phosphorylase screening kit incl. D-Ribose-1-phosphate is designed for a fast screening for optimum production conditions for pyrimidine nucleoside analogs. The Kit systems allows for a direct synthesis of pyrimidine nucleoside analogs starting from Ribose-1-phosphate. Compared to transglycosylation reactions much higher product yields are obtained.

 The Kit contains 4 pyrimidine nucleoside phosphorylases from thermophilic organisms with different temperature optima and substrate spectra.

 Pyrimidine nucleoside phosphorylase screening kit incl. D-Ribose-1-phosphate is designed for screening in small scale: 40 reactions in 100 µl scale can be performed. The conversion of appr. 10 mg of nucleoside is possible with the enzyme in the kit.

Kit components:

Enzymes:   4 pyrimidine nucleoside phosphorylases (Y01-Y04) - 1 mL each of   0.2 mg/mL

Buffers:  6 mL production buffer

Positive Controls:  3 mg uracil

Description:

Nucleoside analogues are a potential class of cytotoxic, antiviral and antiprotozoal drugs. Nucleoside analogs have been in clinical use for almost 50 years and are cornerstones of treatment for patients with cancer or viral infections. Modified nucleosides are currently produced through a long and tedious chemical process that limits the production of new compounds. In contrast, the enzymatic synthesis of nucleosides proceeds with strict stereo- and regioselectivity and requires organic solvents (ethanol, acetonitrile) only for the isolation of the individual desired compound. The production process becomes sustainable and products with a purity >99% are achieved. Production time and costs are drastically reduced.

 So far, nucleoside analogs are produced in a transglycosylation reaction: the transfer of a sugar residue from one nucleoside to another. However, with increasing numbers of modifications the yield of the reaction can be low. In contrast, the direct reaction starting from a sugar-1-phosphate leads to high yields.

Biochemical action of nucleoside phosphorylases:

Pyrimdine nucleoside phosphorylases catalyze the following reaction:

Pyrimidine nucleoside + Pi ↔ Sugar-1-phosphate + base

Unit definition:

One unit of pyrimidine nucleoside phosphorylase will cause the phosphorolysis of 1.0 μmole of uridine to uracil and ribose 1-phosphate per min at pH 7.4 at optimum temperature.

Direct reactions for the production of pyrimidine nucleosides starting from D-Ribose-1-phosphate:

Temperature optima and preferences of pyrimidine nucleoside phosphorylases:

Product use limitations: This product is developed and sold exclusively for research purposes and in vitro use only. The product was not tested for use in diagnostics or drug development, nor is suitable for administration to humans or animals.

#K-NP-0004

The Pyrimidine nucleoside phosphorylase screening kit incl. 2-Deoxyribose-1-phosphate is designed for a fast screening for optimum production conditions for pyrimidine nucleoside analogs. The Kit systems allows for a direct synthesis of pyrimidine nucleoside analogs starting from 2-Deoxyribose-1-phosphate. Compared to transglycosylation reactions much higher product yields are obtained.

The Kit contains 4 pyrimidine nucleoside phosphorylases from thermophilic organisms with different temperature optima and substrate spectra.

Pyrimidine nucleoside phosphorylase screening kit incl. 2-Deoxyribose-1-phosphate is designed for screening in small scale: 40 reactions in 100 µl scale can be performed. The conversion of appr. 10 mg of nucleoside is possible with the enzyme in the kit.

Kit components:

Enzymes:   4 pyrimidine nucleoside phosphorylases (Y01-Y04) - 1 mL each of   0.2 mg/mL

Buffers:  6 mL Production buffer

Positive Controls:  3 mg Uracil

Nucleoside analogues are a potential class of cytotoxic, antiviral and antiprotozoal drugs. Nucleoside analogs have been in clinical use for almost 50 years and are cornerstones of treatment for patients with cancer or viral infections. Modified nucleosides are currently produced through a long and tedious chemical process that limits the production of new compounds. In contrast, the enzymatic synthesis of nucleosides proceeds with strict stereo- and regioselectivity and requires organic solvents (ethanol, acetonitrile) only for the isolation of the individual desired compound. The production process becomes sustainable and products with a purity >99% are achieved. Production time and costs are drastically reduced.

So far, nucleoside analogs are produced in a transglycosylation reaction: the transfer of a sugar residue from one nucleoside to another. However, with increasing numbers of modifications the yield of the reaction can be low. In contrast, the direct reaction starting from a sugar-1-phosphate leads to high yields.

Biochemical action of nucleoside phosphorylases:

Pyrimdine nucleoside phosphorylases catalyze the following reaction:

Pyrimidine nucleoside + Pi ↔ Sugar-1-phosphate + base

Direct reactions for the production of pyrimidine nucleosides starting from D-Ribose-1-phosphate:

Unit definition:

One unit of pyrimidine nucleoside phosphorylase will cause the phosphorolysis of 1.0 μmole of uridine to uracil and ribose 1-phosphate per min at pH 7.4 at optimum temperature.

Temperature optima and preferences of pyrimidine nucleoside phosphorylases:

#K-NP-0005

The Purine nucleoside phosphorylase screening kit incl. D-Ribose-1-phosphate is designed for a fast screening for optimum production conditions for purine nucleoside analogs. The Kit systems allows for a direct synthesis of purine nucleoside analogs starting from Ribose-1-phosphate. Compared to transglycosylation reactions much higher product yields are obtained.

The Kit contains 3 purine nucleoside phosphorylases from thermophilic organisms with different temperature optima and substrate spectra.

Purine nucleoside phosphorylase screening kit incl. D-Ribose-1-phosphate is designed for screening in small scale: 40 reactions in 100 µl scale can be performed. The conversion of appr. 10 mg of nucleoside is possible with the enzyme in the Kit.

Kit components:

Enzymes: 3 purine nucleoside phosphorylases (N01-N03) - 1 mL each of 0.2 mg/mL

Buffers:  6 mL production buffer

Positive Controls:  3 mg adenine

Description:

Nucleoside analogues are a potential class of cytotoxic, antiviral and antiprotozoal drugs. Nucleoside analogs have been in clinical use for almost 50 years and are cornerstones of treatment for patients with cancer or viral infections. Modified nucleosides are currently produced through a long and tedious chemical process that limits the production of new compounds. In contrast, the enzymatic synthesis of nucleosides proceeds with strict stereo- and regioselectivity and requires organic solvents (ethanol, acetonitrile) only for the isolation of the individual desired compound. The production process becomes sustainable and products with a purity >99% are achieved. Production time and costs are drastically reduced.

So far, nucleoside analogs are produced in a transglycosylation reaction: the transfer of a sugar residue from one nucleoside to another. However, with increasing numbers of modifications the yield of the reaction can be low. In contrast, the direct reaction starting from a sugar-1-phosphate leads to high yields.

Biochemical action of nucleoside phosphorylases:

Purine nucleoside phosphorylases catalyze the following reaction:

Purine nucleoside + Pi ↔ Sugar-1-phosphate + base

Unit definition:

One unit of purine nucleoside phosphorylase will cause the phosphorolysis of 1.0 μmole of adenosine to adenine and ribose 1-phosphate per min at pH 7.4 at optimum temperature.

Direct reactions for the production of purine nucleosides starting from D-Ribose-1-phosphate:

Temperature optima and preferences of pyrimidine nucleoside phosphorylases:

Product use limitations:

 This product is developed and sold exclusively for research purposes and in vitro use only. The product was not tested for use in diagnostics or drug development, nor is suitable for administration to humans or animals.

#K-KI-0011

The Nucleoside Kinase Kit + Phosphate Regeneration System was designed to enzymatically phosphorylate a wide range of purine and pyrimidine nucleoside analogs using enzyme NK-12. The use of the phosphate donor regeneration system offers several advantages:

a) Simplified analytics and purification. Regeneration of the phosphate donor will - in case of 100 % conversion - lead to a two-compound reaction solution only containing the synthesized phosphorylated product of interest and ATP, whereas without regeneration also ADP and AMP will be present.

b) Free choice of substrate to phosphate donor ratio. Some enzymatic phosphorylation reactions require a low phosphate donor concentration. Ratios of substrate to phosphate donor lower than 1:1 will lead to a loss of yield if the phosphate donor is not regenerated during the reaction.

Biochemical action of nucleoside kinases:

Nucleoside kinases catalyze the following reaction:

Nucleoside + ATP ↔ NMP + ADP

Note: ATP is the preferred phosphate donor. However, if desired, other phosphate donors can be used, e.g. CTP. The included regeneration system works with all natural nucleoside triphosphates.

Unit definition: One unit of nucleoside kinase will cause the conversion of 1.0 μmol of substrate per min at pH 7.6 at optimum 37°C. ATP is the phosphate donor and is converted to ADP during the reaction. One unit of regeneration kinase will cause the conversion of 1.0 μmol of ADP to ATP per min at pH 7.6 at 37°C.

Product use limitations: This product is developed and sold exclusively for research purposes and in vitro use only. The product was not tested for use in diagnostics or drug development, nor is suitable for administration to humans or animals.

#K-KI-0021

Description:

 The NMP Kinase Kit + Phosphate Regeneration System was designed to enzymatically phosphorylate a wide range of purine and pyrimidine nucleoside monophosphates using enzymes NMPK-21 and -22.  Nucleoside diphosphates and to some extent nucleoside triphosphates are formed as products. The use of the phosphate donor regeneration system offers several advantages:

a) Simplified analytics and purification. Regeneration of the phosphate donor will - in case of 100 % conversion - lead to a two-compound reaction solution only containing the synthesized phosphorylated substrate and ATP, whereas without regeneration also ADP and AMP will be present possibly interfering with HPLC-DAD analysis and/or purification.

b) Free choice of substrate-phosphate donor ratio. Some enzymatic phosphorylation reactions require a low phosphate donor concentration. Ratios of substrate to phosphate donor lower than 1:1 will lead to a loss of yield if the phosphate donor is not regenerated during the reaction.

Biochemical action of nucleoside monophosphate kinases:

Nucleoside monophosphate kinases catalyze the following reaction: NMP + ATP ↔ NDP (+ NTP) + ADP

Unit definition:

One unit of nucleoside monophosphate kinase will cause the conversion of 1.0 μmol each of nucleoside and ATP to 5’- phosphorylated nucleoside and ADP per min at pH 7.6 at optimum 37°C. One unit of regeneration kinase will cause the conversion of 1.0 μmol each of ADP and regeneration phosphate donor to regeneration donor product and ATP per min at pH 7.6 at 37°C. Product use limitations: This product is developed and sold exclusively for research purposes and in vitro use only. The product was not tested for use in diagnostics or drug development, nor is suitable for administration to humans or animals.

Kit components:

Enzymes:

NMP Kinase NMPK-21 E-KI-0021 (2 ml) – 0.1 mg/ml

NMP Kinase NMPK-22 E-KI-0022 (2 ml) – 0.1 mg/ml

Regeneration Kinase RK E-RE-0001 (100 µl) – 1 kU/ml

Buffers: 10x Phosphorylation & Regeneration Buffer (10xB P+R) B-KI-0001 (1 ml)

Other: Regeneration Phosphate Donor D-RE-0001 (2 ml) – 50 mM; Protocol

#K-KI-0031

The NDP Kinase Kit + Phosphate Regeneration System was designed to enzymatically phosphorylate a wide range of purine and pyrimidine nucleoside diphosphates using enzyme NDPK-32. To some extent nucleoside monophosphates are also accepted as substrates. The use of the phosphate donor regeneration system offers several advantages:

a) Simplified analytics and purification. Regeneration of the phosphate donor will - in case of 100 % conversion - lead to a two-compound reaction solution only containing the synthesized phosphorylated substrate and ATP, whereas without regeneration also ADP and AMP will be present possibly interfering with HPLC-DAD analysis and/or purification.

b) Free choice of substrate-phosphate donor ratio. Some enzymatic phosphorylation reactions require a low phosphate donor concentration. Ratios of substrate to phosphate donor lower than 1:1 will lead to a loss of yield if the phosphate donor is not regenerated during the reaction.

Biochemical action of nucleoside diphosphate kinases:

Nucleoside diphosphate kinases catalyze the following reaction: NDP + ATP ↔ NTP + ADP

Unit definition: One unit of nucleoside kinase will cause the conversion of 1.0 μmol each of NDP and ATP to NTP and ADP per min at pH 7.6 at optimum 37°C. One unit of regeneration kinase will cause the conversion of 1.0 μmol each of ADP and regeneration phosphate donor to regeneration donor product and ATP per min at pH 7.6 at 37°C. Product use limitations: This product is developed and sold exclusively for research purposes and in vitro use only. The product was not tested for use in diagnostics or drug development, nor is suitable for administration to humans or animals.

#K-KI-0001

The Nucleoside/NMP/NDP Kinase Kit + Phosphate Regeneration System was designed to enzymatically phosphorylate a wide range of purine and pyrimidine nucleoside analogs using enzymes NK-12, NMPK-21, NMPK-22 and NDPK-32. The use of the phosphate donor regeneration system offers several advantages:

a) Simplified analytics and purification. Regeneration of the phosphate donor will - in case of 100 % conversion - lead to a two-compound reaction solution only containing the synthesized phosphorylated product of interest and ATP, whereas without regeneration also ADP and AMP will be present.

b) Free choice of substrate to phosphate donor ratio. Some enzymatic phosphorylation reactions require a low phosphate donor concentration. Ratios of substrate to phosphate donor lower than 1:1 will lead to a loss of yield if the phosphate donor is not regenerated during the reaction.

Biochemical action of kinase mixtures:

Mixtures of nucleoside/NMP/NDP kinases catalyze the following reaction: Nucleoside + 3*ATP ↔ (NMP + NDP +) NTP + 3*ADP

Unit definition: One unit of nucleoside/nucleotide kinase will cause the conversion of 1.0 μmol of substrate per min at pH 7.6 at optimum 37°C. ATP is the phosphate donor and is converted to ADP during the reaction. One unit of regeneration kinase will cause the conversion of 1.0 μmol of ADP to ATP per min at pH 7.6 at 37°C.

Kit components:

Enzymes:

Nucleoside Kinase NK-12 E-KI-0012 (2 ml) – 0.1 mg/ml

NMP Kinase NMPK-21 E-KI-0021 (2 ml) – 0.1 mg/ml

NMP Kinase NMPK-22 E-KI-0022 (2 ml) – 0.1 mg/ml

NDP Kinase NDPK-32 E-KI-0032 (2 ml) – 0.1 mg/ml

Regeneration Kinase RK E-RE-0001 (100 µl) – 1 kU/ml

Buffers: 10x Phosphorylation & Regeneration Buffer (10xB P+R) B-KI-0001 (1 ml)

Other: Regeneration Phosphate Donor D-RE-0001 (2 ml) – 50 mM; Protocol

#E-PNP-0001;  #E-PNP-0002; #E-PNP-0003

Recombinant enzyme, expressed in E.coli

≥ 50 Units/ml

Protein concentration: 0.2 mg/mL

Suitable for the conversion of appr. 10 mg of purine nucleoside

Purine nucleoside phosphorylase has the ability to perform both phosphorolysis and synthesis of purine nucleoside analogues. The enzyme is useful for the production of modified nucleosides in combination with a pyrimidine nucleoside phosphorylase in a transglycosylation reaction . 

Biochemical action of purine nucleoside phosphorylases: 

Purine nucleoside phosphorylases catalyze the following reaction:

Purine nucleoside + Pi ↔ Sugar-1-phosphate + base

Unit definition: 

One unit of purine nucleoside phosphorylase will cause the phosphorolysis of 1.0 μmole of adenosine to adenine and ribose 1-phosphate per min at pH 7.4 at optimum temperature.

Temperature optima and preferences of pyrimidine nucleoside phosphorylases:

Product use limitations:

This product is developed and sold exclusively for research purposes and in vitro use only. The product was not tested for use in diagnostics or drug development, nor is suitable for administration to humans or animals.

#E-PyNP-0001; #E-PyNP-0002; #E-PyNP-0003; #E-PyNP-0004

Recombinant enzyme, expressed in E.coli

≥ 50 Units/ml

Protein concentration: 0.2 mg/mL

Suitable for the conversion of appr. 10 mg of pyrimidine nucleoside

Pyrimidine nucleoside phosphorylase has the ability to perform both phosphorolysis and synthesis of pyrimidine nucleoside analogues. The enzyme is useful for the production of modified nucleosides in combination with a purine nucleoside phosphorylase in a transglycosylation reaction. 

Biochemical action of pyrimidine nucleoside phosphorylases: 

Pyrimidine nucleoside phosphorylases catalyze the following reaction:

Pyrimidine nucleoside + Pi ↔ Sugar-1-phosphate + base

Unit definition: 

One unit of pyrimidine nucleoside phosphorylase will cause the phosphorolysis of 1.0 μmole of uridine to uracil and ribose 1-phosphate per min at pH 7.4 at optimum temperature.

Temperature optima and preferences of pyrimidine nucleoside phosphorylases:

Product use limitations:

This product is developed and sold exclusively for research purposes and in vitro use only. The product was not tested for use in diagnostics or drug development, nor is suitable for administration to humans or animals.

#E-KI-0012

Nucleoside Kinase NK-12 has the ability to monophosphorylate a wide range of purine and pyrimidine nucleosides and nucleoside analogs at the 5’-position.

Biochemical action of nucleoside kinases:

Nucleoside kinases catalyze the following reaction: Nucleoside + ATP  ↔  5’-NMP + ADP

Note: ATP is the preferred phosphate donor. However if desired, other phosphate donors can be used, e.g. CTP.

 Amount: 2ml; Concentration: 0.1 mg/ml

#E-KI-0021; E-KI-0022

Nucleoside Monophosphate Kinases have the ability to phosphorylate mainly pyrimidine nucleoside monophosphates at the 5’-position to yield nucleoside diphosphates and to some extent nucleoside triphosphates.

Biochemical action of nucleoside monophosphate kinases:

Nucleoside monophosphate kinases catalyze the following reversible reaction: NMP + ATP ↔ NDP (+ NTP) + ADP

Amount: 2ml; Concentration: 0.1 mg/ml

Product use limitations:

This product is developed and sold exclusively for research purposes and in vitro use only. The product was not tested for use in diagnostics or drug development, nor is suitable for administration to humans or animals.

#E-KI-0032

 Nucleoside Diphosphate Kinase NDPK-32 has the ability to phosphorylate purine nucleoside diphosphates at the 5’-position to yield nucleoside triphosphates. To some extent, also pyrimidine nucleoside diphosphates and nucleoside monophosphates are accepted as substrates.

Biochemical action of nucleoside monophosphate kinases:

Nucleoside diphosphate kinases catalyze the following reversible reaction: (NMP +) NDP + ATP ↔ NTP + ADP

Physical properties:

Isoelectric point: 7.15

Molecular weight [kDa]: 16.6

Length [amino acids]: 150

Optimum temperature: 37°C

Amount: 2ml; Concentration: 0.1 mg/ml

Product use limitations:

This product is developed and sold exclusively for research purposes and in vitro use only. The product was not tested for use in diagnostics or drug development, nor is suitable for administration to humans or animals.

#SP-0001 : D-Ribose-1-phosphate Ba salt

Sugar- phosphates are synthesized enzymatically and are currently rare and expensive. They are used in a number of different applications, i.e. direct nucleoside synthesis.

#SP-0002: 2-Deoxyribose-1-phosphate Ba salt

Amount: 5 mg/50 mg