title: Expression, purification, and S-nitrosylation of recombinant histone deacetylase 8 in Escherichia coli.
abstract: Histone deacetylase (HDAC) 8 is a zinc ion dependent enzyme involved in removing the acetyl group from the core histones and other proteins which belong to Class I HDACs. It was reported that nitric oxide (NO) is a key regulator of HDAC function and S-nitrosylation of HDAC2 induces chromatin remodelling in neurons. This work reports the successful recombinant expression of human HDAC8 in Escherichia coli with two plasmids and the purification and S-nitrosylation in vitro. It was found that HDAC8 can be S-nitrosylated by the NO donor S-nitrosoglutathione (GSNO) in vitro, and the activity of HDAC8 was significantly inhibited when incubated with GSNO and S-nitrosocysteine in a time- and dosage-dependent manner, but sodium nitroprusside (SNP), and dithiothreitol cannot reverse this inhibition. These observations support and extend the concept that NO may regulate HDAC8 function by S-nitrosylation.
authors.affiliation: Institute of Medicinal Chemistry, School of Pharmaceutical Sciences, Shandong University, Ji'nan, China.
authors.forename: J H
authors.initials: JH
authors.lastname: Feng
authors.forename: F B
authors.initials: FB
authors.lastname: Jing
authors.forename: H
authors.initials: H
authors.lastname: Fang
authors.forename: L C
authors.initials: LC
authors.lastname: Gu
authors.forename: W F
authors.initials: WF
authors.lastname: Xu
mesh_terms: D015153:Blotting, Western; D003545:Cysteine / Q000031:analogs & derivatives / Q000138:chemical synthesis / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli; D006655:Histone Deacetylases / Q000302:isolation & purification* / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D009569:Nitric Oxide / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism*; D012097:Repressor Proteins / Q000302:isolation & purification* / Q000378:metabolism*; D026422:S-Nitrosoglutathione / Q000138:chemical synthesis / Q000378:metabolism*; D026403:S-Nitrosothiols / Q000138:chemical synthesis / Q000378:metabolism
chemical_list: D017931:DNA Primers; D011994:Recombinant Proteins; D012097:Repressor Proteins; D026403:S-Nitrosothiols; D009569:Nitric Oxide; D026422:S-Nitrosoglutathione; C021315:S-nitrosocysteine; C409470:HDAC8 protein, human; D006655:Histone Deacetylases; D003545:Cysteine