In the 30 years since the discovery of nucleocytoplasmic glycosylation, O-GlcNAc has been demonstrated to regulate protein function by modulating characteristics that include protein folding, localization, degradation, activity, post-translational modifications, and interactions. The cell coordinates these molecular events, on thousands of cellular proteins, in concert with environmental and physiological cues to fine-tune epigenetics, transcription, translation, signal transduction, the cell cycle, and metabolism. The cellular stress response is no exception: diverse forms of injury result in dynamic changes to the O-GlcNAc sub-proteome that promote survival. To date, the majority of studies have focused on identifying proteins and pathways regulated by O-GlcNAc that combat cytotoxicity. Currently, little is known about the regulation of the two enzymes that write and erase the O-GlcNAc-modification, the O-GlcNAc transferase (OGT) and the O-GlcNAcase (OGA). To provide insight into the regulation of OGA, the enzyme that catalyzes the removal of O-GlcNAc, proximity biotin ligation (BioID) in combination with Stable Isoptope Labeling of Amino Acids in Cell Culture (SILAC) was used to define the basal and stress-induced interactome of OGA. This analysis revealed 90 interaction partners of OGA, many of which exhibited increased binding to OGA upon stress. The associations of OGA with fatty acid synthase, OGT, heat shock cognate 70 kDa protein (HSC70), filamin-A, and heat shock protein 27 (HSP27) were confirmed by co-immuno-precipitation. Furthermore, the pool of OGA bound to FAS demonstrated a substantial reduction in specific activity (~85%), suggesting that FAS is a novel protein inhibitor of OGA. Consistent with these observations, overexpression of FAS augmented O-GlcNAcylation of a subset of cellular proteins. Collectively, these data support a model in which protein interactors of OGA, such as FAS, regulate its local activity and thus lead to remodeling of the O-GlcNAc-sub proteome in a manner consistent with cell survival.