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Updated: Jan 13, 2026

DNA-Tethered RNA Polymerase for Programmable In vitro Transcription and Molecular Computation
Published on: December 29, 2021
A universal polyphosphate kinase powers in vitro transcription
Ryusei Matsumoto1, Takayoshi Watanabe2,3, Eishin Yamazaki1
1Department of Life Science and Technology, Institute of Science Tokyo, Tokyo, Japan.
Abstract:
Polyphosphate kinases (PPKs) catalyze phosphoryl transfer between polyphosphates and nucleotides. Polyphosphates are a cost-effective source of phosphorylating power, making PPKs attractive enzymes for nucleotide production. However, at present, applications that require the simultaneous utilization of diverse nucleotides are not possible due to the restricted substrate profiles of PPKs. Here, we present a universal PPK capable of efficiently phosphorylating all eight common ribonucleotides (purines and pyrimidines, monophosphates and diphosphates) to triphosphates. Under optimal conditions, ~70% triphosphate conversion was observed for each substrate. To demonstrate the biotechnological potential of a universal PPK, we developed a one-pot assay for PPK-powered in vitro transcription. Primitive biology likely relied on enzyme promiscuity to support nascent metabolism with a compact proteome. This work highlights how applying the same principle to synthetic biology can facilitate the construction of complex in vitro reaction systems.
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