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

ABSTRACT: Small ubiquitin-like modifiers (SUMOs) are conjugated to protein substrates in cells to regulate their function. The attachment of SUMO family members SUMO1-3 to substrate proteins is reversed by specific isopeptidases called SENPs (sentrin-specific protease). Whereas SENPs are SUMO-isoform or linkage type specific, comprehensive analysis is missing. Furthermore, the underlying mechanism of SENP linkage specificity remains unclear. We present a high-throughput synthesis of 83 isopeptide-linked SUMO-based fluorescence polarization reagents to study enzyme preferences. The assay reagents were synthesized via a native chemical ligation-desulfurization protocol between 11-mer peptides containing a γ-thiolysine and a SUMO3 thioester. Subsequently, five recombinantly expressed SENPs were screene

SUBMITTER: Huppelschoten Y 

PROVIDER: S-EPMC10107784 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature