Preparation of enzymatically active recombinant class III protein deacetylases

Brian J. North; Bjoern Schwer; Nidhi Ahuja; Brett Marshall; Eric Verdin

doi:10.1016/j.ymeth.2005.03.004

Preparation of enzymatically active recombinant class III protein deacetylases

Brian J. North, Bjoern Schwer, Nidhi Ahuja, Brett Marshall, Eric Verdin

Research output: Contribution to journal › Article › peer-review

37 Scopus citations

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.

Original language	English (US)
Pages (from-to)	338-345
Number of pages	8
Journal	Methods
Volume	36
Issue number	4
DOIs	https://doi.org/10.1016/j.ymeth.2005.03.004
State	Published - Aug 2005
Externally published	Yes

All Science Journal Classification (ASJC) codes

Molecular Biology
Biochemistry, Genetics and Molecular Biology(all)

Access to Document

10.1016/j.ymeth.2005.03.004

Cite this

@article{213c0c2b0ce34e8982f308de2a769f9c,

title = "Preparation of enzymatically active recombinant class III protein deacetylases",

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.",

author = "North, {Brian J.} and Bjoern Schwer and Nidhi Ahuja and Brett Marshall and Eric Verdin",

note = "Funding Information: We thank Maki Hirao and Julian Simon (Fred Hutchinson Cancer Center) for providing us with the GST-SIRT1 construct. We thank Sarah Sande for help with the preparation of the manuscript, John Carroll and Jack Hull for graphics, and Stephen Ordway and Gary Howard for editorial assistance. We apologize to colleagues whose work could not be cited due to space constraints. Our research is supported by the NIH, The J. David Gladstone Institutes, and The Sandler Foundation. Eric Verdin is a Senior Scholar of the Ellison Medical Foundation.",

year = "2005",

month = aug,

doi = "10.1016/j.ymeth.2005.03.004",

language = "English (US)",

volume = "36",

pages = "338--345",

journal = "ImmunoMethods",

issn = "1046-2023",

publisher = "Academic Press Inc.",

number = "4",

}

TY - JOUR

T1 - Preparation of enzymatically active recombinant class III protein deacetylases

AU - North, Brian J.

AU - Schwer, Bjoern

AU - Ahuja, Nidhi

AU - Marshall, Brett

AU - Verdin, Eric

N1 - Funding Information: We thank Maki Hirao and Julian Simon (Fred Hutchinson Cancer Center) for providing us with the GST-SIRT1 construct. We thank Sarah Sande for help with the preparation of the manuscript, John Carroll and Jack Hull for graphics, and Stephen Ordway and Gary Howard for editorial assistance. We apologize to colleagues whose work could not be cited due to space constraints. Our research is supported by the NIH, The J. David Gladstone Institutes, and The Sandler Foundation. Eric Verdin is a Senior Scholar of the Ellison Medical Foundation.

PY - 2005/8

Y1 - 2005/8

N2 - 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.

AB - 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.

UR - http://www.scopus.com/inward/record.url?scp=23944452336&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=23944452336&partnerID=8YFLogxK

U2 - 10.1016/j.ymeth.2005.03.004

DO - 10.1016/j.ymeth.2005.03.004

M3 - Article

C2 - 16091304

AN - SCOPUS:23944452336

VL - 36

SP - 338

EP - 345

JO - ImmunoMethods

JF - ImmunoMethods

SN - 1046-2023

IS - 4

ER -

Preparation of enzymatically active recombinant class III protein deacetylases

Abstract

All Science Journal Classification (ASJC) codes

Access to Document

Other files and links

Fingerprint

Cite this