Expanding the Library of Covalent Cysteine Cathepsin Probes Featuring Sulfoxonium Ylide Electrophiles

Overview

Journal ACS Omega

Publisher American Chemical Society

Specialty Chemistry

Date 2024 Nov 4

PMID 39494001

Authors

Bangyan Xu

Simon J Mountford

Philip E Thompson

Laura E Edgington-Mitchell

Affiliations

Soon will be listed here.

Abstract

Covalent activity-based probes are invaluable tools to monitor protease activity in vitro and in vivo. We recently discovered that dimethyl sulfoxonium ylides (SYs) bind selectively to cysteine cathepsin proteases in a mechanism-dependent manner. Herein, we present the synthetic routes and characterization of an expanded library of SY probes with a greater diversity in recognition sequences. The probes exhibit a range of potency and selectivity for the cathepsin family members. We also investigated the impact of fluorophore positioning on probes bearing P1 lysine. When sulfonated cyanine 5 was attached via the lysine side chain, the resulting probe was selective for cathepsin S. When attached to the α-amine, with the side chain amine either free or Boc-protected, the probes reacted with both cathepsin S and X. Bulk in the P1 position is thus well tolerated by cathepsin S but not cathepsin X. We examined the impact of Cy5 sulfonation on probe properties, demonstrating that unsulfonated probes exhibit greater cellular uptake, which affects their relative selectivity. Finally, we demonstrated that SY probes exhibit minimal labeling of cathepsin S in freshly prepared lysates, but this increases during the prolonged incubation of lysates. This work extends our understanding of SY probes and informs future probe development.

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