Structure of a bacterial putative acetyltransferase defines the fold of the human O -GlcNAcase C-terminal domain

Abstract

The dynamic modification of proteins by O-linked N-acetylglucosamine (O-GlcNAc) is an essential posttranslational modification present in higher eukaryotes. Removal of O-GlcNAc is catalysed by O-GlcNAcase, a multi-domain enzyme that has been reported to be bifunctional, possessing both glycoside hydrolase and histone acetyltransferase (AT) activity. Insights into the mechanism, protein substrate recognition and inhibition of the hydrolase domain of human OGA (hOGA) have been obtained via the use of the structures of bacterial homologues. However, the molecular basis of AT activity of OGA, which has only been reported in vitro, is not presently understood. Here, we describe the crystal structure of a putative acetyltransferase (OgpAT) that we identified in the genome of the marine bacterium Oceanicola granulosus, showing homology to the hOGA C-terminal AT domain (hOGA-AT). The structure of OgpAT in complex with acetyl coenzyme A (AcCoA) reveals that, by homology modelling, hOGA-AT adopts a variant AT fold with a unique loop creating a deep tunnel. The structures, together with mutagenesis and surface plasmon resonance data, reveal that while the bacterial OgpAT binds AcCoA, the hOGA-AT does not, as explained by the lack of key residues normally required to bind AcCoA. Thus, the C-terminal domain of hOGA is a catalytically incompetent ‘pseudo’-AT.

Nutrient regulation of gene expression by O-GlcNAcylation of chromatin

“…Hayakawa et al over-expressed OGA in emybryonic stem cells (ESC) and observed a decreased HAT activity toward H3 K14 and H4 K8 upon Thiamet-G treatment or with the expression of OGA D175A , which also showed a significant decrease in O-GlcNAcase activity [17], suggesting that O-GlcNAcase activity is required for HAT activity in vivo . Conversely, a structural study of the HAT domain of human OGA has shown that it shouldn’t be able to bind Acetyl-CoA [18]. Since recombinant OGA purified from bacteria acetylated H3 K14 and H4 K8 but only when pre-incubated with mammalian cell lysates [16], it is then possible that the observed HAT activity could be due to interaction and activation with a third HAT.…”

O-GlcNAcylation, an Epigenetic Mark. Focus on the Histone Code, TET Family Proteins, and Polycomb Group Proteins

“…Therefore, O -GlcNAcylation would be able to regulate the acetylation of the histones tails, but this function remains controversial (35). Whisenhunt et al demonstrated that OGA/NCOAT, OGT, the co-repressor Sin3A (Switch-independent 3A), and HDAC1 (Histone Deacetylase 1) co-exist in a complex that was named O -GlcNAczyme (36).…”

O-GlcNAc and the Epigenetic Regulation of Gene Expression

“…One of these contains a domain with similarities to a histone acetyltransferase domain but lacking the critical residues for catalytic activity. This has been termed a pseudoHAT domain (41). The other major O-GlcNAcase isoform has a 14-amino extension that serves to target it to lipid droplets.…”