Human Sulfotransferase Assays With PAPS Production in situ

Yanan Sun; Lukas Corbinian Harps; Matthias Bureik; Maria Kristina Parr

doi:10.3389/fmolb.2022.827638

Human Sulfotransferase Assays With PAPS Production in situ

Front Mol Biosci. 2022 Feb 28:9:827638. doi: 10.3389/fmolb.2022.827638. eCollection 2022.

Authors

Yanan Sun^{1

2}, Lukas Corbinian Harps¹, Matthias Bureik², Maria Kristina Parr¹

Affiliations

¹ Pharmaceutical and Medicinal Chemistry (Pharmaceutical Analyses), Institute of Pharmacy, Freie Universitaet Berlin, Berlin, Germany.
² School of Pharmaceutical Science and Technology, Health Sciences Platform, Tianjin University, Tianjin, China.

Abstract

For in vitro investigations on human sulfotransferase (SULT) catalyzed phase II metabolism, the costly cofactor 3'-phosphoadenosine-5'-phosphosulfate (PAPS) is generally needed. In the present study, we developed and optimized a new approach that combines SULT-dependent biotransformation using recombinant and permeabilized fission yeast cells (enzyme bags) with PAPS production in situ applying quality by design principles. In the initial application of the procedure, yeast cells expressing human SULT1A3 were used for the production of 4'-hydroxypropranolol-4-O-sulfate from 4-hydroxypropranolol. The optimized protocol was then successfully transferred to other sulfonation reactions catalyzed by SULT2A1, SULT1E1, or SULT1B1. The concomitant degradation of some sulfoconjugates was investigated, and further optimization of the reaction conditions was performed in order to reduce product loss. Also, the production of stable isotope labelled sulfoconjugates was demonstrated utilizing isotopically labelled substrates or ³⁴S-sulfate. Overall, this new approach results in higher space-time yields while at the same time reducing experimental cost.

Keywords: PAPS; SULT; fission yeast; in vitro metabolism; isotopic labelling; method optimization; quality by design; sulfonation.