Purification of bacterially-expressed chick NSD3-SET that is active in an in vitro methyltransferase assay

Abstract

The lysine methyltransferase NSD3, which belongs to a family of histone H3 lysine 36 methyltransferases that are conserved among vertebrates and overexpressed in many cancers, is independently required for both neural crest cell specification and migration in the developing chick embryo. Since cytoplasmic protein lysine methylation is also required for neural crest cell migration in chick, one possibility is that NSD3 methylates non-histone substrates. To test this hypothesis, it is necessary to purify NSD3 protein that is active in a methyltransferase assay, in order to evaluate candidate substrates individually and on protein arrays. In this thesis, I describe a detailed method for high-yield purification of a chick NSD3 catalytic fragment, followed by a chick NSD3 methyltransferase assay with which to test for activity on polynucleosomes and/or candidate non-histone substrates.