Donu, D. | bioRxiv Science

bioRxiv2025

Differential Regulation of SIRT5 Activity by Reduced Nicotinic Acid Riboside (NARH)

SIRT5, one of the human sirtuins, catalyzes the removal of acyl substitutions from lysine residues in a NAD+-dependent manner. In addition to the deacetylase activity, SIRT5 also demonstrates strong desuccinylase, demalonylase, and deglutarylase activity. Through deacylating a broad spectrum of cellular proteins and enzymes, SIRT5 is heavily involved in the regulation of energy metabolism, reactive oxygen species (ROS) reduction, and ammonia detoxification. Accumulating evidence also suggest SIRT5 as a potential therapeutic target for the treatment of neurodegenerative diseases, metabolic disorders, and cancer. In the current study, we report the identification and characterization a SIRT5 modulator, reduced nicotinic acid riboside (NARH). It shows differential regulation of the distinct activities of SIRT5: activates desuccinylation, but mildly suppresses deacetylation. NARH binds to SIRT5 in the absence of NAD+, and demonstrates cellular target engagement and activity. The potential NARH binding site is further investigated using a suite of biochemical and computational approaches. The current study provides greatly-needed mechanistic understanding of SIRT5 regulation, as well as a novel chemical scaffold for further activator development.

biochemistry↗

bioRxiv2024

Biochemical Characterization and Inhibitor Discovery for PfSir2A -- New Tricks for An Old Enzyme

The Sir2 enzyme from Plasmodium falciparum (PfSir2A) is essential for the antigenic variation of this parasite, and its inhibition is expected to have therapeutic effects for malaria. Selective PfSir2A inhibitors are not available yet, partially due to the fact that this enzyme demonstrates extremely weak in vitro deacetylase activity, making the characterization of its inhibitors rather challenging. In the current study, we report the biochemical characterization and inhibitor discovery for this enzyme. PfSir2A exhibits greater enzymatic activity in the presence of DNA for both the peptide and histone protein substrates, suggesting that nucleosomes may be the real substrates of this enzyme. Indeed, it demonstrates robust deacetylase activity against nucleosome substrates, stemming primarily from the tight binding interactions with the nucleosome. In addition to DNA/nucleosome, free fatty acids (FFAs) are also identified as endogenous PfSir2A regulators. Myristic acid, a biologically relevant FFA, shows differential regulation of the two distinct activities of PfSir2A: activates deacetylation, but inhibits defatty-acylation. The structural basis of this differential regulation was further explored. Moreover, synthetic small molecule inhibitors of PfSir2A were discovered through the screening of a library of human sirtuin regulators. The mechanism of inhibition of the lead compounds were investigated. Collectively, the mechanistic insights and inhibitors described in this study will facilitate the future development of small molecule PfSir2A inhibitors as antimalarial agents.

biochemistry↗