AMP-activate protein kinase (AMPK) is a key regulator of cellular and whole-body energy homeostasis that amends energy imbalances caused by physiological and pathological processes. During energy stress conditions (with increased cellular AMP/ATP ratio), AMPK switches on catabolic pathways and simultaneously switch-off energy-consuming anabolic pathways. The heterotrimeric AMPK αβγ complex is composed of an α-catalytic subunit and regulatory β- and γ-subunits. These subunits consist of multiple isoforms (α1/2, β1/2, γ1/2/3) in mammals those are important for regulation, function and tissue specific distribution. β1-Ser108 phosphorylation stabilises an allosteric drug site, which also likely allows regulation by an unknown natural metabolite. In this current study, to aimed to investigate the cellular fate of β1-Ser108 phosphorylation. We performed a stable isotope dimethyl labelled-based quantitative proteomic and phosphoproteomic analysis using β1/2-dKO iMEFs, transduced with either β1 mutant Ser108Ala or Ser108Glu (which previously shown to be an effective phosphomimetic). Following 1 h phenformin (2 mM) treatment, β1 Ser108Ala and Ser108Glu transduced iMEF lysates were compared to trace out the changes in proteome and phosphoproteome. At least 15 cellular phosphoproteins showed signiﬁcant changes in phosphorylation between Ser108Ala and Ser108Glu-expressing cells in our study with no detectable differences in AMPK expression or phenformin-induced Thr172 phosphorylation. We identiﬁed increased phosphorylation of p21-activated kinase 2 (PAK2) on Ser141/Ser152, located in the kinase inhibitory domain, in phenformin-treated iMEFs-expressing β1-Ser108Glu, whereas global proteome data showed no changes in PAK2 protein level. Immunoblot analysis with a phosphospeciﬁc antibody conﬁrmed signiﬁcant increase in pPAK2-Ser141 in Ser108Glu-expressing cells, validating our representative phosphoproteomic data. Pathway and network analysis using ingenuity pathway analysis (IPA) identiﬁed ‘Cell cycle, connective tissue development and function, cellular movement’ as one of the top networks associated with these perturbed cellular phosphoproteins that hints at the cellular fate of β1-Ser108 phosphorylation.