Nature communicationsOct 6, 2025

A streamlined method to produce pseudouridine using a custom-designed process

mRNA Technology Weekly Brief ↗PubMed ↗DOI ↗OA ↗

Updated Jun 27, 2026

Abstract

Essence

A designer enzymatic pathway in E. coli produced gram-scale with a simplified purification strategy and favorable techno-economic modeling.

Evidence

This biomanufacturing platform experiment optimized UMP nucleosidase, pseudouridine monophosphate glycosidase, and phosphatase components, reaching 44.8 g/L and 45.3 g/L titers and 71% purification recovery.

Caveat

The evidence is a production-platform demonstration and economic analysis, so broader industrial performance and nucleoside-molecule generalizability remain to be validated.

Simplified

mRNA vaccines, featured by incorporated (Ψ), represent a milestone in combating diseases, thus highlighting Ψ importance in drug development. However, economic and environmental challenges have persisted in sustainable Ψ production. Here, we formulate a streamlined designer Ψ pathway, comprising UMP nucleosidase, ΨMP glycosidase, and ΨMP phosphatase, and realize its gram-scale production by targeted discovery of a prominent UMP-preferred nucleosidase (NmYgdH). The optimized pathway, containing NmYgdH, RjPsuG (ΨMP glycosidase), and HDHD1 (ΨMP-specific phosphatase) is cloned into E. coli and systematic evaluation of multiple strategies achieves a Ψ titer of 44.8 g·L. Moreover, a thyA-dependent, tunable, and eco-friendly strategy for sustainable Ψ production is demonstrated in a 5 L bioreactor achieving titer of 45.3 g·L. Finally, we establish a simplified-strategy for rapid Ψ purification with a recovery-rate of 71%, and techno-economic analysis is employed to validate the feasibility and advantages of this fermentation platform for Ψ biomanufacturing. Therefore, this study provides a blueprint for industrial-production of nucleoside-related molecules. -1 -1

Key numbers

45.3 g·L

Ψ Production Titer

Achieved in a 5 L bioreactor using a streamlined enzyme pathway.

71%

Recovery Rate

Achieved through a rapid purification process.

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Funding

Competing interests

Competing interests: W.C., L.Y., R.C., Z.D. and Zheng.W. have submitted two patent applications filed by Wuhan University with the China National Intellectual Property Administration (application numbers CN202510665627.7 and CN202510665631.3), as well as one related PCT patent application (international application number PCT/CN2025/105204), all of which pertain to the NmYgdH application and pseudouridine production described in this work. The remaining authors declare no competing interests.

PubMed

A streamlined method to produce pseudouridine using a custom-designed process

Abstract

Key numbers

Full Text

Funding

Competing interests

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