Na+/K+ pump is key to SGLT1 function; exchanges Na+ for /K+ to maintain the gradient; uses energy (i.e., ATPase); this allows Na+ to drag the monosaccharide (glucose) along through the transport channel and into the enterocyte.
Summary • Complex carbs have been digested to simple sugars • Enterocyte has absorbed them and delivered them to the blood
Key Pathways in Carbohydrate Utilization • Insulin-stimulated uptake • Low circulating concentrations of insulin: GLUT4 is sequestered within cytosolic vesicles in myocytes and adipocytes • Accumulation of glucose in blood triggers insulin release from pancreatic ß-cells; Insulin-receptor signaling induces the redistribution of GLUT4 from intracellular storage sites to the plasma membrane; once incorporated into the cell membrane, GLUT4 facilitates the passive diffusion of circulating glucose down its concentration gradient • The pathway in which GLUT4 is integrated into the plasma membrane begins with insulin binding to its receptor dimer • The receptor dimer autophosphorylates (intrinsic tyrosine kinase activity) and subsequently activates insulin-responsive substrate-1 (IRS1), which initiates a series of reactions to activate protein kinase B (PKB) • Once phosphorylated, PKB is in its active form and phosphorylates other targets that stimulate GLUT4 mobilization and trafficking to the plasma membrane • Once inside cells, glucose is rapidly phosphorylated by hexokinase to form G6P; it is this form of glucose that serves as the initial substrate for glycolysis • G6P cannot diffuse back out of cells; thus G6P serves to maintain the concentration gradient for glucose to passively enter cells.
Basal State Insulin-Stimulated From Oatey et al. Biochem. J. (1997) 327, 637±642
Converting Sugars to an Energy Currency • Key concepts • Glycolysis • TCA cycle • Electron transport chain • Oxidative phosphorylation Sugar ATP
Glycolysis • G6P is trapped in cell cytosol; glycolysis occurs in cytosol
ATP is consumed at RXN • 1 and 3 • 6C molecule split into two • 3C molecules • 3C molecule drives • production of NADH • and ATP • No net ATP until PEP is • converted to pyruvate • Under aerobic conditions, • NADH will be further • utilized in mitochondria to • power the electron trans- • port chain (generate ATP) • 6. Under “anaerobic” conditions, • lactate is produced
Glycolysis Summary • C6H12O6 + 2 NAD+ + 2 ADP + 2 P -----> 2 pyruvic acid, CH3(C=O)COOH + 2 ATP + 2 NADH + 2 H+ Steps 1 and 3 = - 2ATPSteps 6 and 9 = + 4 ATPNet "visible" ATP produced = 2
With insufficient oxygen, or in cells lacking mitochondria (RBC), pyruvate will be reduced at the expense of NADH, to lactate by lactate dehydrogenase.
TCA (Citric Acid) Cycle • Now, we are shifting from the cytosol, to the mitochondria • Using C-containing molecules that originated with dietary carbohydrate and metabolized to tri-carboxylic acids , we are generating ATP and reducing power that will flow into the electron transport chain • “anything containing C and H that can be reduced to CO2 and H2O contains energy”…..oxygen serves as the terminal e- acceptor in the electron transport chain
Key Features, TCA Cycle • The citric acid cycle begins with Acetyl-CoA (ACoA) transferring its two-carbon acetyl group to the four-carbon acceptor compound (oxaloacetate) to form a six-carbon compound (citrate) • The citrate then goes through a series of chemical transformations, losing first one, then a second carboxyl group as CO2. The carbons lost as CO2 originate from what was oxaloacetate, not directly from acetyl-CoA. The carbons donated by acetyl-CoA become part of the oxaloacetate carbon backbone after the first turn of the citric acid cycle. Loss of the ACoA-donated carbons as CO2 requires several turns of the citric acid cycle. However, because of the role of the citric acid cycle in anabolism, they may not be lost since many TCA cycle intermediates are also used as precursors for the biosynthesis of other molecules. • Most of the energy made available by the oxidative steps of the cycle is transferred as energy-rich electrons to NAD+, forming NADH. For each acetyl group that enters the citric acid cycle, three molecules of NADH are produced
Key Features, TCA Cycle • Electrons are also transferred to the electron acceptor Q, forming QH2. • At the end of each cycle, the four-carbon oxaloacetate has been regenerated, and the cycle continues • Mitochondria in animals including humans possess two succinyl-CoA synthetases, one that produces GTP from GDP, and another that produces ATP from ADP • Products of the first turn of the cycle are: one GTP (or ATP), three NADH, one QH2, two CO2 • Because two ACoA molecules are produced from each glucose molecule, two cycles are required per glucose molecule. Therefore, at the end of both cycles, the products are: two GTP (or ATP), six NADH, two QH2, and four CO2
The Respiratory Chain • We have generated reducing power: per turn • (1) FADH2 • (3) NADH • (1) QH2 But…… the energy currency we are after is ATP • The respiratory (e-) transport chain allows us to finish extracting energy from what was our dietary carbs by oxidation of the reducing power, coupled to phosphorylation of ADP • “Oxidative Phosphorylation” ATP
ATP Yield • Oxidation of NADH to NAD+ pumps 3 protons which charges the electrochemical gradient with enough potential to generate 3 ATP • Oxidation of FADH2 to FAD+ pumps 2 protons which charges the electrochemical gradient with enough potential to generate 2 ATP.
Respiratory Chain • Protons are translocated across the membrane, from the matrix to the intermembrane space • Electrons are transported along the membrane, through a series of protein carriers • Oxygen is the terminal electron acceptor, combining with e- and H+ ions to produce water • As NADH and FADH2deliver more H+ and electrons into the ETC, the proton gradient increases, with H+ building up outside the inner mitochondrial membrane, and OH- inside the membrane • It is the energy derived from this proton (electrochemical) gradient that is used to drive phosphorylation of ADP (synthesis of ATP)
NADH and FADH2 carry protons (H+) and e- to the electron transport chain located in the membrane. The energy from the transfer of electrons along the chain transports protons across the membrane and creates an electrochemical gradient. As the accumulating protons follow the electrochemical gradient back across the membrane through an ATP synthase complex, the energy of the gradient is transferred by the ATP synthase system from the gradient to ADP in the synthesis of ATP. At the end of the electron transport chain, two protons, two electrons, and half of an oxygen molecule combine to form water. Since oxygen is the final electron acceptor, the process is called aerobic respiration.
Excess Energy from CH2O can be stored as fat • De novo lipogenesis • Acetyl CoA is the key
Lipogenesis: Key Points • Occurs in the cytosol; encompasses • fatty acid synthesis and triacylglycerol • synthesis • Catalyzed by acetyl CoA carboxylase • Requires biotin for carboxylation rxn • Product is malonyl CoA This is the rate limiting step……. Full activation requires excess energy as is reflected in (1) adequate cellular ATP and low AMP, (2) an accumulation of citrate in the TCA cycle that ultimately exits the mitochondria via an elaborate shuttle to return to the cytosol
Lipogenesis: Key Points • Sequential additions of 2C units (acetyl CoA) results in the formation of palmitate (C16:0) 8 acetyl CoA + 7 ATP + 14 NADPH + 6 H ---> palmitate + 14 NADP + 8 CoA + 7 ADP + 7 Pi + 6 H2O • Catalyzed by Fatty Acid Synthase • an enzyme complex with 7 catalytic activities
Lipogenesis: Key Points • Palmitate is esterified on a glycerol backbone to form “triacyl” glycerol
Lipogenesis: Key Points • Avians and humans: Liver (hepatocyte) is primary site of de novo lipogenesis • Pig: about 100% in the adipocyte itself • Beef: adipocyte • Rodents: adipocyte and hepatocyte, about 50:50 • Dog/Cat: Mixed, but probably favors hepatocyte
Overall Summary • Digested, absorbed and metabolized CH2O to convert the energy contained in the hydrocarbons to a usable currency, ATP • Captured excess energy from these CH2O molecules in an energy dense form, suitable for storage (triacylglycerol), which can be mobilized to meet deficits that might arise later