Fatty Acid Handling Beta-oxidation FA transport Integration of metabolic signaling

Fatty Acid/b-oxidation Cycle 1x FADH2 1x NADH Acetyl-CoA 3x NADH+ 1xFADH2 Acyl(n)-CoA + NAD+ + FAD Acyl(n-2)-CoA + Acetyl-CoA + NADH +FADH2 Carnitine palmitoyltransferase Fatty acid elongation Acyl-CoA Acyl-CoA synthase FAD Acyl-CoA dehydrogenase Acyl-CoA FADH2 acetyl-CoA acyltransferase Acetyl-CoA Didehydroacyl-CoA Acyl-CoA hydrase CoA-SH 3-hydroxyacyl-CoA dehydrogenase Hydroxyacyl-CoA Oxoacyl-CoA NADH NAD+

Fatty acid/carbohydrate oxidation Oxygen CnH2n + 3/2 n O2  n CO2+ n H2O CnH2nOn +n O2 n CO2 + n H2O Respiratory Quotient CO2/O2 0.67 Fatty acids 1.00 Carbohydrates Adenine electron transporters 6-C glucose6 NADH + 2 FADH2 (3:1) 16-C FA  32 NADH + 16 FADH2 (2:1)

Reactive oxygen FADH2 oxidative stress Succinate; saturated FA FADH2 + Fe3+  FADH • + H+ + Fe2+ Fe2+ + H2O2Fe3+ + OH- + OH• FADH2 more completely reduces UQ than does NADH Acyl-CoA Acyl-CoA FAD FADH2 UQ O2 Acyl-CoA dehydrogenase Acyl-CoA oxidase ETF:QO oxidoreductase FADH2 FAD UQH2 H2O2 Didehydroacyl-CoA Didehydroacyl-CoA

Lipogenesis De novo synthesis of fatty acid Malonyl-CoA Mostly liver (human; diet) Cytoplasmic – ACC expression Malonyl-CoA Carboxylation of Acetyl-CoA Working substrate for FAS Fatty acid synthase Sequentially transfers 2x C of Malonyl-CoA to fatty acid chain 16-C palmitoyl-CoA Acetyl-CoA Acetyl-CoA Carboxylase Malonyl-CoA Fatty Acid Synthase Fatty acid Fatty Acid Synthase

Free fatty acids from triglycerides FFA cleavage from circulating lipoproteins Protein/cholesterol carriers: Lipoprotein Density inversely correlates with lipid Correlates with cholesterol/FA (except HDL) VLDL & LDL to IDL Lipoprotein lipase (LPL) HDL scavenges cholesterol & facilitates IDL breakdown Triglycerides are retained in intracellular droplets Don’t fit in membrane (no phosphate) Not water soluble

Mitochondrial import of fatty acids FAAcyl-CoA Acyl-Carnitine Acyl-CoA Cytoplasm Intermembrane Matrix Working substrate Boron & Boulpaep

Mitochondrial Transport Carrier protein (FABP) Long chain acyl-CoA synthetase (LCAS) Cross outer membrane via porin Convert to acylcarnitine in intermembrane Cross inner membrane via carnitine:acylcarnitine transferase Convert back to acyl-CoA in matrix

Regulation of lipid metabolism Substrate/Allosteric Palmitate inhibits insulin signaling Malonyl-CoA inhibits FA transport to mt Esp muscle, where ACC is mitochondrial Citrate activates acetyl-CoA carboxylase Palmitoyl-CoA inhibits ACC Phosphorylation AMP dependent kinase inhibits ACC FAT/CD36 translocation

Allosteric regulation of FA metabolism Carnitine Palmitoyltransferase Fatty Acid Synthetase ACC Malonyl-CoA Palmitoyl-CoA Citrate B-oxidation NADH Citric Acid Cycle Acetyl-CoA Citric acid cycle products promote FA synthesis FA synthesis intermediaries inhibit FA import

Metabolic substrate selection Fatty acids Energy dense (37 kJ/g) Abundant (95%) O2 delivery limit Diffusion limit CHO Low density (17 kJ/g) Limited supply (5%) Hydrated (67 wt%) Lower O2 requirement Readily available Substrate selection is an issue for fed/fasted state and for overall activity Lipolysis rate (%VO2) Glcolysis rate (%VO2) Total metabolic rate (%VO2 max) McClelland, 2004

Signaling integration Insulin/IGF-1 GLUT4 translocation InsRIRS-1PI3KPDKPKB --|GSK--|GS mTORp70S6k/4EBP1 Fatty acids Inhibit pyruvate dehydrogenase Inhibit insulin signaling FADAGPKC--|IRS-1 Activate glycogen synthase

AMP kinase Allosterically activated by AMP PFKglycolysis Adenylate kinase: 2 ADP  AMP + ATP ADP levels insensitive to energy state PFKglycolysis --|GSGlyconeogenesis --|ACCMalonyl CoA--|CPTFA oxidation --|ACClipogenesis TSC2--|mTOR…protein synthesis --|HMGCoAcholesterol synthesis

Hormonal Regulation Insulin Glucagon Thyroid hormone (Triiodothyronine ) Steroid T3TR ?CaMKKβ AMPK Complex III/IV proteins, PGC-1 GLUT4 Epinepherine (adrenaline) Tissue specific: muscle/liver AdbARGsACcAMPPKA GPglycogenolysis lipolysis

Peroxisome Proliferator PPAR-a/b/g (nuclear hormone, fatty acid sensor) Transcriptional complex PPAR(a/b/g), PGC-1(a/b), NRF-1/2, CREB Subunits of Complex I-V; FAT; GLUT4 Mitochondrial biogenesis Transcriptional Pathways InsulinAkt--|FOXO--|PGC-1 Ca2+CaMKCREBPGC-1 Glucagon/stressPKACREBPGC-1 AMPK?PGC-1 Post-Translational, too FADH2 O2 Peroxisome FAD H2O2 FADH2 UQ Mitochondria FAD UQH2

Visceral FA subcutaneous ↑ congestive heart failure PPAR/PGC signaling ↓ plasma NEFA Visceral FA subcutaneous ↑ insulin sensitivity ↑ mitochondria ↑ congestive heart failure ↑ fluid retention Fernandez-Marcos & Auwerx, 2011

Close relation between nutritive status and growth Insulin/IGF via PI3K Glycolysis/lipolysis Protein synthesis Differentiation AMP-Kinase IGF inhibition AcetylCoA/CoA Inhibit pyruvate dehydrogenase NADH converts oxaloacetate to malate, reduces TCA intermediaries, including citrate Growth hormone release NAD+/NADH increases with cell confluence