State of the art in (semi-)synthesis of Ubiquitin- and Ubiquitin-like tools

Ubi-tagging: A ubiquitin-based modular platform for site-directed antibody conjugation

“…7). ( 5) The established full chemical synthesis of ubiquitin [45][46][47] further expands its potential applications as an antibody conjugation tag. This facilitates the attachment of chemical modi cations such as small molecules, uorophores, tags, or chemical warheads at one or more positions in a de ned manner (Supplementary Fig.…”

A new dawn beyond lysine ubiquitination

“…What has enabled breakthroughs in the field of canonical ubiquitination is the capacity to prepare defined Ub chains and ubiquitinated substrates using chemical rather than enzymatic methods. These have then been used to investigate the effects of ubiquitination on protein structure and function, and to determine the DUBs that process them (Figure 6) 94 . Access to homogenous Ub conjugates is a challenge because the enzymes that carry out the modification are often unknown, are not amenable to large scale reconstitution, or modify multiple sites in vitro.…”

“…To achieve site-specific attachment, esterification of a defined threonine side chain was achieved using orthogonal side chain protection. However, in the field of chemical lysine ubiquitination, amino acid derivatives that undergo chemoselective and traceless isopeptide bond formation have provided much-needed versatility, enabling convergent chemical synthesis of more complex conjugates and even chemoselective ubiquitination of recombinant proteins 94 . Application of these principles to non-lysine ubiquitination would enable access to defined non-lysine ubiquitinated substrates whose study will shed further light on the function of this modification.…”

In‐Plate Chemical Synthesis of Isopeptide‐Linked SUMOylated Peptide Fluorescence Polarization Reagents for High‐Throughput Screening of SENP Preferences

“…In contrast to the extensive knowledge on deubiquitinating enzymes (DUBs) we are very limited in our understanding of SENPs substrate specificity. Biochemical analysis and crystal structures have been instrumental in the discovery of linkage specificity of several DUBs, [22–25] and significant contributions to profile DUBs have come from the development of assay reagents/probes [26] . For example, in previous work by Geurink et al.…”

“…Biochemical analysis and crystal structures have been instrumental in the discovery of linkage specificity of several DUBs,[ 22 , 23 , 24 , 25 ] and significant contributions to profile DUBs have come from the development of assay reagents/probes. [26] For example, in previous work by Geurink et al. DUB specificity was investigated by using isopeptide‐linked fluorescent assay reagents.…”