Dataset Information

Sustainable Flow-Synthesis of (Bulky) Nucleoside Drugs by a Novel and Highly Stable Nucleoside Phosphorylase Immobilized on Reusable Supports.

ABSTRACT: The continuous synthesis of valuable nucleoside drugs was achieved in up to 99 % conversion by using a novel halotolerant purine nucleoside phosphorylase from Halomonas elongata (HePNP). HePNP showed an unprecedented tolerance to DMSO, usually required for substrate solubility, and could be immobilized on agarose microbeads through disulfide bonds, via a genetically fused Cystag. This covalent yet reversible binding chemistry showcased the reusability of agarose microbeads in a second round of enzyme immobilization with high reproducibility, reducing waste and increasing the sustainability of the process. Finally, the flow synthesis of a Nelarabine analogue (6-O-methyl guanosine) was optimized to full conversion on a 10 mm scale within 2 min residence time, obtaining the highest space-time yield (89 g L^-1 h^-1 ) reported to date. The cost-efficiency of the system was further enhanced by a catch-and-release strategy that allowed to recover and recirculate the excess of sugar donor from the downstream water waste.

SUBMITTER: Benitez-Mateos AI

PROVIDER: S-EPMC9298701 | biostudies-literature | 2022 Jan

REPOSITORIES: biostudies-literature

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Sustainable Flow-Synthesis of (Bulky) Nucleoside Drugs by a Novel and Highly Stable Nucleoside Phosphorylase Immobilized on Reusable Supports.

Benítez-Mateos Ana I AI Paradisi Francesca F

ChemSusChem 20211127 1

The continuous synthesis of valuable nucleoside drugs was achieved in up to 99 % conversion by using a novel halotolerant purine nucleoside phosphorylase from Halomonas elongata (HePNP). HePNP showed an unprecedented tolerance to DMSO, usually required for substrate solubility, and could be immobilized on agarose microbeads through disulfide bonds, via a genetically fused Cystag. This covalent yet reversible binding chemistry showcased the reusability of agarose microbeads in a second round of e ...[more]

PMID: 34726353

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Project description:Nucleoside phosphorylases (NPs) are pivotal enzymes in the salvage pathway, catalyzing the reversible phosphorolysis of nucleosides to produce nucleobases and α-D-ribose 1-phosphate. Due to their efficiency in catalyzing nucleoside synthesis from purine or pyrimidine bases, these enzymes hold significant industrial importance in the production of nucleoside-based drugs. Given that the thermodynamic equilibrium for purine NPs (PNPs) is favorable for nucleoside synthesis-unlike pyrimidine NPs (PyNPs, UP, and TP)-multi-enzymatic systems combining PNPs with PyNPs, UPs, or TPs are commonly employed in the synthesis of nucleoside analogs. In this study, we report the first development of two engineered bifunctional fusion enzymes, created through the genetic fusion of purine nucleoside phosphorylase I (PNP I) and thymidine phosphorylase (TP) from Thermus thermophilus. These fusion constructs, PNP I/TP-His and TP/PNP I-His, provide an innovative one-pot, single-step alternative to traditional multi-enzymatic synthesis approaches. Interestingly, both fusion enzymes retain phosphorolytic activity for both purine and pyrimidine nucleosides, demonstrating significant activity at elevated temperatures (60-90 °C) and within a pH range of 6-8. Additionally, both enzymes exhibit high thermal stability, maintaining approximately 80-100% of their activity when incubated at 60-80 °C over extended periods. Furthermore, the transglycosylation capabilities of the fusion enzymes were explored, demonstrating successful catalysis between purine (2'-deoxy)ribonucleosides and pyrimidine bases, and vice versa. To optimize reaction conditions, the effects of pH and temperature on transglycosylation activity were systematically examined. Finally, as a proof of concept, these fusion enzymes were successfully employed in the synthesis of various purine and pyrimidine ribonucleoside and 2'-deoxyribonucleoside analogs, underscoring their potential as versatile biocatalysts in nucleoside-based drug synthesis.

Dataset Information

Sustainable Flow-Synthesis of (Bulky) Nucleoside Drugs by a Novel and Highly Stable Nucleoside Phosphorylase Immobilized on Reusable Supports.

Publications

Sustainable Flow-Synthesis of (Bulky) Nucleoside Drugs by a Novel and Highly Stable Nucleoside Phosphorylase Immobilized on Reusable Supports.

Similar Datasets

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