br AMPK structure and mechanism

AMPK: structure and mechanism of action AMPK is a metabolic master switch that regulates downstream signals based on shifts in the surrounding energy reservoir [6]. It is expressed in a number of tissues, including the kidney, the liver, the skeletal muscle, the adipose tissue, and the hypothalamus of the brain [9,10]. It is activated when adenosine triphosphate (ATP) consumption causes an increase in the adenosine monophosphate (AMP)-to-ATP ratio [6]. For example, under such conditions as hypoglycemia, exercise, hypoxia, and ischemia, in which accentuated cellular signaling cascade and intracellular stress arise, consumption of ATP is suppressed while production is spurred [11]. Regulation of AMPK activation can be achieved through either allosteric activation by AMP or stimulation by upstream kinases, including a compound consisting of 3 proteins: STE-related adaptor (STRAD), mouse protein 25, and the tumor-suppressor liver kinase B1 (LKB1) [12]. In addition, other enzymes, such as Ca++/calmodulin-dependent protein kinase kinase β (CaMKKβ) [13] and transforming growth factor (TGF) β-activated kinase [14], also participate in the cellular signaling cascade (Fig. 1). On activation, AMPK signals through its downstream substrates to achieve energy homeostasis by stimulating processes that generate ATP through such actions as fatty 4064 oxidation and glucose transport, while inhibiting those that use ATP through the opposing actions of fatty acid synthesis and protein synthesis [12]. Thus, the net effect of AMPK activation is an increased cellular energy level via the inhibition of anabolic energy-consuming pathways, as well as the stimulation of catabolic, energy-producing pathways [11]. AMPK is a heterotrimer consisting of a catalytic α subunit (α1 and α2) and regulatory β (β1 and β2) and γ subunits (γ1, γ2, and γ3). Its isoforms are tissue specific [15]. The γ subunit includes 4 distinct cystathionine beta synthase domains that perceive shifts in the AMP-to-ATP ratio, conferring the ability to switch AMPK on and off [16]. The α subunit is a catalytic domain with its activating loop at threonine-172 (Thr-172) that switches on as it gets phosphorylated by upstream AMPK [17]. Ser485 of the subunit exerts its inhibitory action via phosphorylation by Akt or protein kinase A or by autophosphorylation [18]. “ST loop,” a serine/threonine–rich insert of C-terminal domains of the α subunit, appears to contain several regulatory phosphorylation sites, which block access to upstream kinases [16]. AMPK activation can be triggered when the following conditions are fulfilled. First, increased levels of AMP binding to the γ subunit incur conformational change that exposes the active site Thr-172 on the α subunit, making it a better substrate for upstream kinases. Second, phosphorylation of the activating loop of the α subunit by an upstream kinase is required. This combination of allosteric activation and phosphorylation of the stimulatory site leads to the accentuation of AMPK activity to levels greater than 1,000-fold [15,17]. Activated AMPK then phosphorylates its main downstream targets, acetyl-CoA carboxylase (ACC) and hydroxymethylglutaryl CoA reductase (HMGCR), which are primarily involved in the rate-limiting steps of lipid homeostasis. Phosphorylation at serine 79 (Ser79) exerts inhibitory action by preventing the conversion of acetyl-CoA to malonyl-CoA, further encouraging the oxidation of fatty acids. Other downstream targets of AMPK include tuberous sclerosis complex protein-2, which is associated with cell growth and autophagy [19], and peroxisome proliferator–activated receptor-gamma coactivator-1α (PGC-1α), which hinders protein synthesis by inhibiting the mammalian target of rapamycin (mTOR) complex 1 [19], in turn inhibiting cholesterol synthesis and stimulating mitochondrial biogenesis (Fig. 1). Overall, these downstream targets of AMPK are expected to have favorable effects on DN by maintaining metabolic homeostasis.