Facile Preparation of UFMylation Activity-Based Probes by Chemoselective Installation of Electrophiles at the C-Terminus of Recombinant UFM1

Overview

Journal ACS Cent Sci

Specialty Chemistry

Date 2022 Jun 27

PMID 35756382

Authors

Kateryna A Tolmachova

Jakob Farnung

Jin Rui Liang

Jacob E Corn

Jeffrey W Bode

Affiliations

Soon will be listed here.

Abstract

Aberrations in protein modification with ubiquitin-fold modifier (UFM1) are associated with a range of diseases, but the biological function and regulation of this post-translational modification, known as UFMylation, remain enigmatic. To provide activity-based probes for UFMylation, we have developed a new method for the installation of electrophilic warheads at the C-terminus of recombinant UFM1. A C-terminal UFM1 acyl hydrazide was readily produced by selective intein cleavage and chemoselectively acylated by a variety of carboxylic acid anhydrides at pH 3, without detriment to the folded protein or reactions at unprotected amino acid side chains. The resulting UFM1 activity-based probes show a range of tunable reactivity and high selectivity for proteins involved in UFMylation processes; structurally related E1s, E2s, and proteases associated with Ub or other Ubls were unreactive. The UFM1 probes were active both in cell lysates and in living cells. A previously inaccessible α-chloroacetyl probe was remarkably selective for covalent modification of the active-site cysteine of de-UFMylase UFSP2 .

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