High-level bacterial expression and purification of human SirT2 protein for NMR studies.
Silent information regulator 2 (Sir2) proteins are a class of protein deacetylase enzymes that play key roles in transcriptional gene silencing, DNA repair, and aging. Here, we describe the high-level bacterial expression and purification of a human SirT2 construct that yields high resolution NMR spectra. By removing the N-terminal helix alpha0 and using Thioredoxin as a fusion partner, greater than 10 mg/L of purified protein can be obtained from minimal media. The protein is fully functional and enables NMR-based screening and structural studies of this important protein.
Sun
Chaohong
C
Global Pharmaceutical Discovery Division, Abbott Laboratories, Abbott Park, IL, USA.
Song
Danying
D
Marcotte
Patrick A
PA
Richardson
Paul L
PL
Hajduk
Philip J
PJ
D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D019906:Nuclear Magnetic Resonance, Biomolecular; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification*; D056565:Sirtuin 2; D037761:Sirtuins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification*; D013879:Thioredoxins / Q000378:metabolism
D011993:Recombinant Fusion Proteins; D013879:Thioredoxins; C474199:SIRT2 protein, human; D056565:Sirtuin 2; D037761:Sirtuins