Preparation of enzymatically active recombinant class III protein deacetylases.
Methods (San Diego, Calif.)
confidence
Key findings
Review of protocols for purification of enzymatically active human sirtuins 1, 2, and 3 and their activity on histone and nonhistone substrates; no clinical/biological endpoints.
View source on PubMed (PMID 16091304) ↗
- Sample size
- Not reported
- Population
- Not applicable (in vitro enzymatic activity measurement)
- Dosing
- Not reported
- Duration
- Not reported
- Route
- Not reported
- Blinding
- not_reported
- Controls
- none
- Drug class
- coenzyme
Full abstract
Class III histone deacetylases, or sirtuins, are homologous to the Saccharomyces cerevisiae transcriptional regulator SIR2. The class III enzymes are characterized by their dependence on nicotinamide adenine dinucleotide (NAD+). This cofactor serves as an acetyl-group acceptor in the deacetylation reaction generating O-acetyl-ADP-ribose. Enzymatic activity of sirtuin can be measured in vitro using recombinant proteins purified from mammalian cells after overexpression or after purification from Escherichia coli. This review discusses protocols for the purification of enzymatically active human sirtuin 1, 2, and 3 and their activities on histone and nonhistone substrates.