Abstract
Nicotinamide adenine dinucleotide (NAD+)-dependent ADP-ribosylation plays important roles in physiology and pathophysiology. It has been challenging to study this key type of enzymatic post-translational modification in particular for protein poly-ADP-ribosylation (PARylation). Here we explore chemical and chemoenzymatic synthesis of NAD+ analogues with ribose functionalized by terminal alkyne and azido groups. Our results demonstrate that azido substitution at 3′-OH of nicotinamide riboside enables enzymatic synthesis of an NAD+ analogue with high efficiency and yields. Notably, the generated 3′-azido NAD+ exhibits unexpected high activity and specificity for protein PARylation catalyzed by human poly-ADP-ribose polymerase 1 (PARP1) and PARP2. And its derived poly-ADP-ribose polymers show increased resistance to human poly(ADP-ribose) glycohydrolase-mediated degradation. These unique properties lead to enhanced labeling of protein PARylation by 3′-azido NAD+ in the cellular contexts and facilitate direct visualization and labeling of mitochondrial protein PARylation. The 3′-azido NAD+ provides an important tool for studying cellular PARylation.
Highlights
Nicotinamide adenine dinucleotide (NAD+)-dependent ADP-ribosylation plays important roles in physiology and pathophysiology
It is found that recombinant human nicotinamide riboside kinase 1 (NRK1) and nicotinamide mononucleotide adenylyltransferase 1 (NMNAT1) can efficiently catalyze formation of 3′-azido NAD+ from its nicotinamide riboside (NR) analogue precursor in the presence of adenosine triphosphate (ATP)
Inspired by NRK- and NMNAT-mediated biosynthesis of NAD+ from NR (Fig. 1a), we envisioned that ribose-functionalized NR may allow facile chemoenzymatic synthesis of NAD+ analogues (Fig. 1b)
Summary
Nicotinamide adenine dinucleotide (NAD+)-dependent ADP-ribosylation plays important roles in physiology and pathophysiology. The 3′-azido NAD+-based poly-ADP-ribose (PAR) polymers reveal improved resistance to degradation by human poly(ADP-ribose) glycohydrolase (PARG) These unique properties for the 3′-azido NAD+ collectively result in more significant labeling of protein PARylation in the cell lysates relative to NAD+ and adenine-substituted NAD+ analogues and enable direct visualization and labeling of mitochondrial protein PARylation. This ribose-functionalized NAD+ with marked activity and specificity for protein PARylation offers an important tool for studying posttranslational ADP-ribosylation and may pave the ways a
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
