Protein modification by ADP-ribose via acid-labile linkages

Area of Science:

• Biochemistry
• Molecular Biology
• Cell Biology

Background:

• Nicotinamide adenine dinucleotide (NAD) serves as the ADP-ribose donor for protein mono-ADP-ribosyltransferases.
• ADP-ribosylation is a post-translational modification crucial for regulating signaling, differentiation, and protein transport.
• Previous studies focused on specific linkages, leaving potential new modification types unexplored.

Purpose of the Study:

• To synthesize and characterize model conjugates of ADP-ribose with acetal linkages.
• To investigate the chemical stability of these novel linkages.
• To determine if similar acid-labile ADP-ribose modifications occur in vivo in animal proteins.

Main Methods:

• Preparation of low molecular weight ADP-ribose conjugates with acetal linkages.
• Structural elucidation using Nuclear Magnetic Resonance (NMR) spectroscopy.
• Chemical stability assays in formic acid, comparing to known linkages.
• In vivo analysis of rat liver proteins for ADP-ribose modification.

Main Results:

• Successfully synthesized and characterized ADP-ribose acetal model conjugates.
• Demonstrated rapid release of ADP-ribose from acetal models in formic acid, distinguishing them from other linkages.
• Confirmed in vivo modification of rat liver proteins via acid-labile ADP-ribose linkages.
• Quantified modification at ~16 pmol ADP-ribose/mg protein, found across subcellular fractions.

Conclusions:

• Established a new class of endogenous ADP-ribose modification in animal cells characterized by acid-labile linkages.
• The broad distribution across subcellular fractions suggests a significant role in cellular regulation.
• This finding opens new avenues for understanding ADP-ribosylation's impact on cellular functions.