Abstract 1278 Mitochondrial Hsp40 chaperone, Tid1, is a novel mtDNA replication factor

O-linked beta-N-acetylglucosamine (O-GlcNAc) transferase (OGT) is the only human enzyme that transfers the O-GlcNAc sugar from UDP-GlcNAc to nuclear and cytoplasmic proteins.OGT can O-GlcNAcylate over one thousand substrates that participate in almost every cellular process, and misregulation of O-GlcNAc levels can lead to a variety of diseases.Despite its large scope of substrates, how OGT chooses its many targets is poorly understood.OGT has two domains: glycosyltransferase (catalytic) domain and tetratricopeptide (TPR) domain.Even though the catalytic domain is where the sugar transfer occurs, it is not where selection happens.It has recently been proposed that adaptor proteins drive selection by interacting with the TPR domain to direct OGT to subsets of substrates.Recently, CARM1 (Coactivator Associated Arginine Methyltransferase 1) was shown to interact with OGT and cause an increase in glycosylation of rat brain lysates, suggesting a role as an OGT adaptor.However, the exact binding site of CARM1 along OGT's structure is unknown.We hypothesize that adaptors bind along the solvent-exposed surface of OGT's TPR domain and that CARM1 would as well.To test this hypothesis, we used an OGT protein library where we incorporated a photoactivatable unnatural amino acid at 26 locations along the solvent-exposed surface of the TPR domain to covalently map CARM1's interaction.Using this library, we found that CARM1 does interact along the TPR domain at two main locations of CARM1 on OGT TPR 1 (F27, W38) as well as TPRs 11-13 (V346, M372, T409).Surprisingly, we found that the interaction along TPRs 11-13 showed crosslinked adducts with a molecular weight equal to approximately two OGTs and one CARM1, suggesting that CARM1 is able to interact with an OGT dimer complex.These results provide insights into how CARM1 interacts with OGT and will help guide future func