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submitted on 23.09.2020 and posted on 23.09.2020by Felix Kaspar, Peter Neubauer, Anke Kurreck
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 Thermus thermophilus 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 106 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.
A. K. is CEO of the biotech company BioNukleo GmbH. F. K. is a scientist at BioNukleo GmbH and P. N. is a member of the advisory board. These affiliations constitute no conflict of interest with the results presented and discussed in this report.