Communication
Thiosuccinyl Peptides as Sirt5-Specific Inhibitors
Bin He, Jintang Du,† and Hening Lin*
Department of Chemistry and Chemical Biology, Cornell University, Ithaca, New York 14853, United States
S
* Supporting Information
ABSTRACT: Sirtuins, a class of enzymes known as
nicotinamide adenine dinucleotide-dependent deacety-
lases, have been shown to regulate a variety of biological
processes, including aging, transcription, and metabolism.
Sirtuins are considered promising targets for treating
several human diseases. There are seven sirtuins in humans
(Sirt1−7). Small molecules that can target a particular
human sirtuin are important for drug development and
fundamental studies of sirtuin biology. Here we demon-
strate that thiosuccinyl peptides are potent and selective
Sirt5 inhibitors. The design of these inhibitors is based on
our recent discovery that Sirt5 prefers to catalyze the
hydrolysis of malonyl and succinyl groups, rather than an
acetyl group, from lysine residues. Furthermore, among
the seven human sirtuins, Sirt5 is the only one that has this
Figure 1. NAD-dependent demalonylation and desuccinylation
reactions catalyzed by Sirt5.
unique acyl group preference. This study demonstrates
that the different acyl group preferences of different
sirtuins can be conveniently utilized to develop small
molecules that selectively target different sirtuins.
III sirtuins.13 Several mitochondrial proteins are malonylated
and succinylated on lysine residues.13 Sirt5 serves to remove
some of the malonyl and succinyl groups from the proteins,
possibly as a mechanism for reversible regulation of protein
activity in mitochondria.13 Independently, Zhao and co-workers
also identified lysine succinylation in Escherchia coli as a new
post-translational modification.14
The discovery of the robust enzymatic activity now provides
a reliable assay for the development of Sirt5 inhibitors.13 In
addition, since Sirt5’s acyl group preference is unique among all
the human sirtuins,13 we reasoned that we could take advantage
of this to develop Sirt5-specific inhibitors. Such inhibitors
would be valuable tools to study the biological function of Sirt5
in cells and to evaluate whether Sirt5 would be a good target for
treating human diseases. Herein we report that thiosuccinyl
peptides can be used as Sirt5-specific inhibitors.
Thioacetyl peptides can inhibit sirtuins with deacetylase
activities by forming a stalled covalent intermediate (Figure
2).19−21 Because Sirt5 uses the same mechanism as the
deacetylases to remove malonyl and succinyl groups,13 we
reasoned that thiosuccinyl or thiomalonyl peptides would be
mechanism-based inhibitors for Sirt5. Because other sirtuins do
not recognize malonyl and succinyl lysine peptides,13 we
predicted that thiomalonyl and thiosuccinyl peptides should be
Sirt5-specific inhibitors. To test this hypothesis, we synthesized
a histone H3 lysine 9 (H3K9) thiosuccinyl peptide (H3K9TSu,
Figure 3). We chose to use the thiosuccinyl group because
irtuins are a class of enzymes known as nicotinamide
S
adenine dinucleotide (NAD)-dependent deacetylases.1
Humans have seven sirtuins, Sirt1−7, which are known to
play important roles in many biological processes, such as the
regulation of life span, transcription, and metabolism.2 Small
molecules that can regulate sirtuin activity have been shown to
have potential in treating several human diseases,3 such as
cancer,4−7 diabetes,8,9 and Parkinson’s disease.10 Although
there are controversies regarding the link between sirtuins and
longer life span11 and the effectiveness of Sirt1 activators,12 it is
clear that sirtuins have important biological functions.
Inhibitors that are specific for a particular sirtuin will be very
useful for investigating the biological function and therapeutic
potential of the specific sirtuin. The major obstacle in the
development of small molecules that can regulate sirtuin
activity is the fact that four of the seven human sirtuins (Sirt4−
7) have either very weak or no deacetylase activity.1,2 This
poses two difficulties for the development of small molecules
that can regulate sirtuin activity. First, it is hard to develop
inhibitors or activators that target these sirtuins because no
robust activity assay is available. Second, it is hard to tell
whether the inhibitors/activators that target Sirt1, Sirt2, and
Sirt3 can also target Sirt4−7.
Recently, our laboratory discovered that human Sirt5, a
mitochondrial sirtuin with weak deacetylase activity, is an
efficient demalonylase and desuccinylase (Figure 1).13 The
specificity for negatively charged malonyl and succinyl groups is
determined by a conserved Arg and Tyr residue in most class-
Received: September 26, 2011
Published: January 12, 2012
© 2012 American Chemical Society
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dx.doi.org/10.1021/ja2090417 | J. Am. Chem.Soc. 2012, 134, 1922−1925