Characterization of the Enzymatic Activity of SETDB1 and Its 1:1 Complex with ATF7IP

Biochemistry
Aravind BasavapathruniThomas V Riera

Abstract

The protein methyltransferase (PMT) SETDB1 is a strong candidate oncogene in melanoma and lung carcinomas. SETDB1 methylates lysine 9 of histone 3 (H3K9), utilizing S-adenosylmethionine (SAM) as the methyl donor and its catalytic activity, has been reported to be regulated by a partner protein ATF7IP. Here, we examine the contribution of ATF7IP to the in vitro activity and substrate specificity of SETDB1. SETDB1 and ATF7IP were co-expressed and 1:1 stoichiometric complexes were purified for comparison against SETDB1 enzyme alone. We employed both radiometric flashplate-based and SAMDI mass spectrometry assays to follow methylation on histone H3 15-mer peptides, where lysine 9 was either unmodified, monomethylated, or dimethylated. Results show that SETDB1 and the SETDB1:ATF7IP complex efficiently catalyze both monomethylation and dimethylation of H3K9 peptide substrates. The activity of the binary complex was 4-fold lower than SETDB1 alone. This difference was due to a decrease in the value of kcat as the substrate KM values were comparable between SETDB1 and the SETDB1:ATF7IP complex. H3K9 methylation by SETDB1 occurred in a distributive manner, and this too was unaffected by the presence of ATF7IP. This finding is important a...Continue Reading

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Citations

Dec 8, 2016·Nature Reviews. Molecular Cell Biology·Paul B Talbert, Steven Henikoff
Oct 6, 2019·SLAS Discovery·Shane M BukerP Ann Boriack-Sjodin
Oct 3, 2019·EMBO Reports·Takeshi TsusakaYoichi Shinkai
Mar 9, 2018·Advances in Urology·Joshua L PierceJames F Amatruda
May 4, 2021·Frontiers in Cell and Developmental Biology·Chao WanRocky S Tuan

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