Volume 18, Issue 10 p. 931-940
Full Paper

Studies of the Binding of Modest Modulators of the Human Enzyme, Sirtuin 6, by STD NMR

Beatriz E. Bolívar

Beatriz E. Bolívar

Department of Chemistry, University at Albany, 1400 Washington Avenue, Albany, NY, 12205 USA

Search for more papers by this author
Prof. John T. Welch

Corresponding Author

Prof. John T. Welch

Department of Chemistry, University at Albany, 1400 Washington Avenue, Albany, NY, 12205 USA

Search for more papers by this author
First published: 21 February 2017
Citations: 11

Graphical Abstract

Sirtuin success: Binding of NAD+, NAM, and NAM analogues to human enzyme sirtuin 6 (SIRT6) was followed by STD NMR methodology, which confirmed the mixed inhibition observed in binding kinetics experiments. This method also afforded qualitative insight into the nature of binding, suggesting the geometry of the ligand-binding epitopes in solution with the apo- and holoenzyme.

Abstract

Pyrazinamide (PZA), an essential constituent of short-course tuberculosis chemotherapy, binds weakly but selectively to Sirtuin 6 (SIRT6). Despite the structural similarities between nicotinamide (NAM), PZA, and pyrazinoic acid (POA), these inhibitors modulate SIRT6 by different mechanisms and through different binding sites, as suggested by saturation transfer difference (STD) NMR. Available experimental evidence, such as that derived from crystal structures and kinetic experiments, has been of only limited utility in elucidation of the mechanistic details of sirtuin inhibition by NAM or other inhibitors. For instance, crystallographic structural analysis of sirtuin binding sites does not help us understand important differences in binding affinities among sirtuins or capture details of such dynamic process. Hence, STD NMR was utilized throughout this study. Our results not only agreed with the binding kinetics experiments but also gave a qualitative insight into the binding process. The data presented herein suggested some details about the geometry of the binding epitopes of the ligands in solution with the apo- and holoenzyme. Recognition that SIRT6 is affected selectively by PZA, an established clinical agent, suggests that the rational development of more potent and selective NAM surrogates might be possible. These derivatives might be accessible by employing the malleability of this scaffold to assist in the identification by STD NMR of the motifs that interact with the apo- and holoenzymes in solution.

Conflict of interest

The authors declare no conflict of interest.