Fat mobilization in adipocytes b R Note insulin

Fat mobilization in adipocytes b. R Note: insulin inhibits TAG mobilization PKA P Hormone-sensitive lipase TAG Hormone-sensitive lipase DAG FFA MAG FFA glycerol FFA See Fig 16. 7 Horton

Fatty acid oxidation 1. Activation • Acyl Co. A synthetase (thiokinase) 2. Transport into mitochondria • acylcarnitine transferase/acylcarnitine translocase 3. b oxidation cycle • • acyl Co. A dehydrogenase enoyl Co. A hydratase L-3 -hydroxy Co. A dehydrogenase Thiolase

Acyl Co. A synthetase R-COO- + Co. A-SH + ATP O R-C- S-Co. A + AMP + PPi Note: 4 different enzymes specific for FA of differing chain length. location: outer membrane of mitochondria, ER membranes

Transport of fatty acyl Co. A into mitochondria Carnitine Fatty Acyl. Co. A + carnitine Carnitine acyltransferase I Malonyl Co. A acylcarnitine + Co. A translocase Mito matrix Fatty Acyl. Co. A + carnitine acylcarnitine + Co. A Carnitine acyltransferase II

Net yield of ATP C 16 FA + Co. A + ATP C 16 acyl Co. A + AMP + PP 1 C 16 acyl Co. A + 7 NAD + 7 FAD 8 Ac. Co. A 31 NADH 15 FADH 2 8 GTP 8 Ac. Co. A + 7 NADH + 7 FADH 2 24 NADH + 8 FADH 2 + 8 GTP + 16 CO 2 77. 5 ATP 22. 5 ATP 8 ATP 108 ATP - 2 ATP Net = 106 ATP

b-oxidation of unsaturated fatty acids • 2, 4 dieonyl-Co. A reductase: converts cis to trans double bond • Enoyl-Co. A isomerase: converts diene to single double bond

b-oxidation of unsaturated fatty acids e. g. Linoleic acid: C 18 cis, cis-D 9, 12 3 H C 18 C C 16 C C 14 C C 12 C C 10 C C 8 C C 6 C C 4 3 rounds of b-oxidation 3 Ac-Co. A C 12 cis, cis-D 3, 6 3 H C 18 C C 16 C C 14 C C 12 C g b C 10 C a C 8 CO-S-Co. A Enoyl-Co. A isomerase C 12 trans, cis-D 2, 6 3 H C 12 C C 10 C C 8 C C 6 C g b a C 4 C C 2 CO-S-Co. A 1 round of b-oxidation Ac-Co. A C 10 cis-D 4 3 H C 12 C C 8 C C 6 C C 4 C C 2 CO-S-Co. A

C 10 cis-D 4 3 H C C C 8 12 C C 6 C C 4 C CO-S-Co. A C 2 Acyl-Co. A dehydrogenase C 10 trans, cis-D 2, 4 3 H C C C 8 12 C C 6 C C 4 C CO-S-Co. A C 2 2, 4 dieonyl-Co. A reductase NADP C 10 trans-D 3 3 H C C 12 C 8 C C 6 C C 4 C C 2 CO-S-Co. A Enoyl-Co. A isomerase C 10 cis-D 2 3 H C 12 C C 8 C C 6 C C 4 C C 2 CO-S-Co. A continued b-oxidation

b-oxidation of odd numbered fatty acids Requires: • Propionyl Co. A carboxylase (biotin) - adds CO 2 • methyl malony Co. A racemase - converts D isomer of methyl malonyl Co. A to L isomer • methyl malonyl Co. A mutase (adenosylcobalamin) - rearranges MMCo. A to yield succinyl Co. A

Oxidation of odd-numbered fatty acids O CH 3 -CH 2 -C-S-Co. A CO 2 Proprionyl Co. A Biotin, ATP CH 3 O -OOC-CH C-S-Co. A D-methylmalonyl Co. A racemase O -OOC-CH C-S-Co. A L-methylmalonyl Co. A CH 3 adenosylcobalamin mutase -OOC-CH 2 O C-S-Co. A succinyl Co. A

Cobalamin B 12 methylcobalamin R = CH 3 - transfer of methyl groups adenosylcobalamin R = 5’-deoxyadenosinyl - intramolecular rearrangements See fig 7. 24 Horton

Fatty acid biosynthesis Where: cytoplasm liver, fat cells When: good energy charge, insulin Process: 1. Transfer of Ac. Co. A from mito to cyto 2. Acetyl Co. A carboxylase 3. Fatty acid synthase

1. Transfer of Ac. Co. A from mito to cyto Citrate lyase Citrate + ATP + + Co. A NADH NADPH malate OAA + Ac. Co. A + ADP + Pi Citrate lyase OAA Pyruvate Ac. Co. A citrate cyto mito H+ Antiport with pyr or Pi OAA citrate Pyruvate Ac. Co. A

Question: Incubation of tissue using the above pathway with only one of succinate-2, 3 -14 C or succinate-1, 4 -14 C will result in the production of radiolabeled fatty acid. Identify which substrate will yield 14 C-fatty acids.

Acetyl-Co. A carboxylase 1 (ACC 1) Ac. Co. A + CO 2 + ATP biotin -OOC malonyl Co. A + ADP + Pi CH 2 C - SCo. A O Regulation: • Hormonal control - AMP kinase - inhibited by PKA mediated phosphorylation • glucagon (liver) • adrenalin (adipocytes) - enhanced by insulin • • Allosteric regulation - citrate activates phospho form - inhibited by palmitoyl Co. A Nutritional status

Regulation of FA metabolism by phosphorylation Adrenalin Glucagon activates inhibits c. AMP ACT FA synthesis AMPactivated kinase PKA Malonyl Co. A ACC(active) PP 2 A ACC-PO 4(inactive) TAG lipase (inactive) phosphatase TAG lipase-PO 4 (active) Note: insulin activates ACC by stimulating the dephosphorylation reaction

Steps in FA biosynthesis 1. Loading: transfer to ACP and ketoacyl-ACPsynthase i. CH 3 C - SCo. A + ACP-SH CH 3 O ii. CH 3 O C - ACP + S-Synthase O iii. -OOC- C – SACP + Co. ASH CH 3 C – S-Synthase + ACP-SH O CH 2 C - SCo. A O -OOC- ACP-SH Co. ASH CH 2 C – S-ACP O

2. Condensation: -OOC- CH 2 C – S-ACP + CH 3 O C – S-Synthase O HS-Synthase CO 2 CH 3 C – CH 2 C – S-ACP O O

3. reduction CH 3 H C – CH 2 C – S-ACP O O CH 3 C CH 2 C – S-ACP OH NADPH O NADP 4. dehydration H CH 3 H C – CH 2 C – S-ACP OH O CH 3 C CH C – S-ACP O H 2 O 5. reduction H CH 3 C CH C – S-ACP CH 3 O CH 2 CH C – S-ACP O NADPH NADP

Subsequent rounds of synthesis - transfer of growing FA to S-Synthase - addition of 2 carbon units from malonyl-S-ACP CH 3 CH 2 C – S-ACP O Synthase-SH ACP-SH CH 3 CH 2 C – S-synthase O -OOC- CH 2 C-SACP CO 2 + Synthase-SH O CH 3 CH 2 C O CH C – S-ACP O

• Chain elongation • introduction of double bonds • linoleic and linolenic acids • synthesis of arachidonic acid – an important precursor of several biologically active molecules