Recombinant SETDB1 protein
Background
SETDB1 (SET Domain, Bifurcated 1) is a histone methyltransferase that methylates histone H3 at lysine 9. H3K9 methylation is associated with transcriptional repression and gene silencing through recruitment of HP1 proteins to methylated histones. H3K9 trimethylation is coordinated with DNA methylation, most likely through formation of a complex with MBD1 and ATF7IP that leads to transcriptional repression. SETDB1 activity is dependent on MBD1 and is heritably maintained through DNA replication by being recruited by CAF-1. SETDB1 is targeted to histone H3 by TRIM28, a factor recruited by KRAB zinc-finger proteins. Aberrant SETDB1 function is associated with the pathogenesis of Huntington Disease.
Application Notes
Recombinant SETDB1 is suitable for use in the study of enzyme kinetics, inhibitor screening, and selectivity profiling.
HMT Assay Conditions: 3 µM H3 (1-21) peptide was incubated with different concentrations of recombinant SETDB1 protein in a 10 µl reaction system containing 50 mM Tris-HCl pH 8.6, 0.02% Triton X-100, 2 mM MgCl2, 1 mM TCEP and 50 µM SAM for 1 hour, then 10 µl H3K9me2 antibody and SA-XL665 mixture (each 1:100 dilution in HTRF Detection Buffer) was added to each reaction system and incubated for 30 min. All the operations and reactions were performed at room temperature. HTRF assay was used for detection.
Protein Details
Recombinant SETDB1 protein was expressed in a baculovirus expression system as the full length protein (accession number NP_005245698.1) with an N-terminal FLAG-Tag. The molecular weight of the protein is 145 kDa.
