There are nine protein arginine methyltransferases (PRMTs 1–9) expressed in humans that vary in both subcellular localization and substrate specificity. The variation in substrate specificity between isozymes leads to competing effects that result in either activation or repression of tumor suppressor genes. Current methods used to study substrate specificity for these enzymes utilize radioisotopic labeling of substrates, mass spectrometry analysis of complex samples, or coupled assays that monitor cofactor degradation. Herein, we report the development of a rapid, nonradioactive, and sensitive method for screening multiple peptides in parallel to gain insight into the substrate specificity of PRMT enzymes. Our assay provides a major advantage over other high-throughput screening assays (e.g., ELISA, AlphaScreen chemiluminescence) by eliminating the need for purification of individual peptides and provides a timesaving, cost-effective alternative to the traditional PRMT assays. A one-bead one-compound (OBOC) peptide library was synthesized and subsequently screened against PRMT1 in a 96-well plate. This screen resulted in identification of a novel PRMT1 substrate with kinetic parameters similar to histone H4–21 (e.g., the best-known PRMT1 peptide substrate).Keywords: arginine; high-throughput; histone; one-bead one-compound; protein arginine methyltransferase; substrate screening

Agmatine deiminases (AgDs) catalyze the hydrolytic conversion of agmatine (decarboxylated arginine) to N-carbamoylputrescine with concomitant release of ammonia. These enzymes, which are encoded by some pathogenic bacterial species, confer a competitive survival advantage by virtue of energy production and acid tolerance through agmatine catabolism. Herein we report the development of a clickable activity-based protein profiling (ABPP) probe that targets the AgD encoded by Streptococcus mutans with high selectivity and sensitivity.