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1、杨荣武杨荣武生物化生物化学原理学原理第二版第二版Chapter29 FFA metabolismOutlineFatty acid oxidation1.-oxidation2.-oxidation3.-oxidationFormation and utilization of ketone bodiesFatty acid synthesisRegulation of FFA metabolismBeta Oxidation of Fatty AcidsKnoop showed that fatty acids must be degraded by removal of 2-C units
2、 $Albert Lehninger showed that this occurred in the mitochondria $F. Lynen and E. Reichart showed that the 2-C unit released is acetyl-CoA, not free acetate $The process begins with oxidation of the carbon that is beta to the carboxyl carbon, so the process is calledbeta-oxidationFranz Knoops classi
3、c experiment indicating that fatty acids are metabolically oxidized at their -carbon atomCoA activates FAs for oxidationAcyl-CoA synthetase condenses fatty acids with CoA, with simultaneous hydrolysis of ATP to AMP and PPi $Formation of a CoA ester is expensive energetically $Reaction just barely br
4、eaks even with ATP hydrolysis $But subsequent hydrolysis of PPi drives the reaction strongly forward $Note the acyl-adenylate intermediate in the mechanism!Connecting the FFA to CoA-SHCarnitine as a CarrierCarnitine carries fatty acyl groups across the inner mitochondrial membrane $Short chain fatty
5、 acids are carried directly into the mitochondrial matrix $Long-chain fatty acids cannot be directly transported into the matrix $Long-chain FAs are converted to acyl carnitines and are then transported in the cell $Acyl-CoA esters are formed inside the inner membrane in this way Transport of acyl-c
6、arnitine into the matrixrate-limiting step for oxidation of FAs -Oxidation of Fatty AcidsA Repeated Sequence of 4 Reactions $Strategy: create a carbonyl group on the -C $First 3 reactions do that; fourth cleaves the -keto ester$Products: an acetyl-CoA and a fatty acid two carbons shorter, FADH2, NAD
7、H $The first three reactions are crucial and classic - we will see them again and again in other pathways oxidation - Fatty acids are degraded by repeated cycles of oxidation at the b-carbon and cleavage of the C-C bond to yield acetate unitsOxidation of fatty acids: three steps, one newThe -oxidati
8、on of saturated fatty acidsAcyl-CoA DehydrogenaseOxidation of the C-C bond $A family of three soluble matrix enzymes $Mechanism involves proton abstraction, followed by double bond formation and hydride removal by FAD $Electrons are passed to an electron transfer flavoprotein, and then to the ETC$En
9、zyme is inhibited by a metabolite of hypoglycin (from akee fruit)The acyl-CoA dehydrogenase reactionFourth reaction: thiolaseaka -ketothiolase $Cysteine thiolate on enzyme attacks the -carbonyl group $Thiol group of a new CoA attacks the shortened chain, forming a new, shorter acyl-CoA $This is the
10、reverse of a Claisen condensation: attack of the enolate of acetyl-CoA on a thioester $Even though it forms a new thioester, the reaction is favorable and drives other three Mechanism of action of ketoacyl-CoA thiolaseSummary of -OxidationRepetition of the cycle yields a succession of acetate units
11、$Thus, palmitic acid yields eight acetyl-CoAs $Complete -oxidation of one palmitic acid yields 106 molecules of ATP $Large energy yield is the consequence of the highly reduced state of the carbon and compact storage (no hydration) of fatty acidsYield of ATP during the Complete Oxidation of One Mole
12、cule of Palmitoyl-CoA - oxidation challengesmonounsaturated FApolyunsaturated FAodd-carbon FA(-C containing CH3 group)Unsaturated Fatty AcidsConsider monounsaturated fatty acids:$Oleic acid, palmitoleic acid $Normal -oxidation for three cycles $cis-3 acyl-CoA cannot be utilized by acyl-CoA dehydroge
13、nase $Enoyl-CoA isomerase converts this to trans- 2 acyl CoA -oxidation continues from this point -oxidation of a mono unsaturated fatty acidPolyunsaturated Fatty AcidsSlightly more complicated Same as for oleic acid, but only up to a point:3 cycles of -oxidation enoyl-CoA isomerase 1 more round of
14、-oxidation trans- 2, cis- 4 structure is a problem!2,4-Dienoyl-CoA reductase to the rescue!Odd-Carbon Fatty Acids -Oxidation yields propionyl-CoA $Odd-carbon fatty acids are metabolized normally, until the last three-C fragment - propionyl-CoA - is reached $Three reactions convert propionyl-CoA to s
15、uccinyl-CoA $The involvement of biotin and B12 What about 3-C leftovers?the coenzyme B12 reactionBranched-Chain Fatty AcidsAn alternative to -oxidation is required $Branched chain FAs with branches at odd-number carbons are not good substrates for -oxidation $-oxidation is an alternative $Phytanic a
16、cid -oxidase decarboxylates with oxidation at the alpha position $-oxidation occurs past the branch The -oxidation pathway for phytanic acid oxidationPeroxisomal -Oxidation Peroxisomes - organelles that carry out flavin-dependent oxidations, regenerating oxidized flavins by reaction with O2 to produ
17、ce H2O2 $Similar to mitochondrial -oxidation, but initial double bond formation is by acyl-CoA oxidase $Electrons go to O2 rather than e- transport $Fewer ATPs result Comparison of Mitochondrial and Peroxisomal fatty acid oxidationKetone Bodies A special source of fuel and energy for certain tissues
18、 $Some of the acetyl-CoA produced by fatty acid oxidation in liver mitochondria is converted to acetone, acetoacetate and -hydroxybutyrate $These are called ketone bodies$Source of fuel for brain, heart and muscle $Major energy source for brain during starvation $They are transportable forms of fatt
19、y acids!Ketone Bodies - IIInteresting Aspects of Their Synthesis $Occurs only in the mitochondrial matrix $First step is reverse thiolase $Second reaction makes HMG-CoA $These reactions are mitochondrial analogues of the (cytosolic) first two steps of cholesterol synthesis$Third step - HMG-CoA lyase
20、 - is similar to the reverse of citrate synthase Formation of ketone bodiesUtilization of ketone bodies in extra-hepatic tissuesKetone Bodies and DiabetesStarvation of cells in the midst of plenty$Glucose is abundant in blood, but uptake by cells in muscle, liver, and adipose cells is low $Cells, me
21、tabolically starved, turn to gluconeogenesis and fat/protein catabolism $In type I diabetics, OAA is low, due to excess gluconeogenesis, so Ac-CoA from fat/protein catabolism does not go to TCA, but rather to ketone body production $Acetone can be detected on breath of type I diabeticsFatty Acid Bio
22、synthesis The Biosynthesis and Degradation Pathways are DifferentAs in cases of glycolysis/gluconeogenesis and glycogen synthesis/breakdown, fatty acid synthesis and degradation go by different routes There are four major differences between fatty acid breakdown and biosynthesis The DifferencesBetwe
23、en fatty acid biosynthesis and breakdown Intermediates in synthesis are linked to -SH groups of acyl carrier proteins (as compared to -SH groups of CoASynthesis in cytosol; breakdown in mitochondriaEnzymes of synthesis are one polypeptideBiosynthesis uses NADPH/NADP+; breakdown uses NADH/NAD+ and FA
24、D/FADH2Comparison of fatty acid oxidation and fatty acid biosynthesisGeneral Features of FFA biosynthesisCellular location-cytosolPrimers- acetyl-CoAActivated 2-C units - malonyl-CoADecarboxylation of malonyl-CoA and reducing power of NADPH drive chain growthChain grows to 16-carbonsOther enzymes ad
25、d double bonds and more CsChallenge: Ac-CoA in CytosolWhat are the sources?Amino acid degradation produces cytosolic acetyl-CoAFAA oxidation produces mitochondrial acetyl-CoAGlycolysis yields cytosolic pyruvate which is converted to acetyl-CoA in mitochondriaCitrate-malate-pyruvate shuttle provides
26、cytosolic acetate units and reducing equivalents for fatty acid synthesisThe citrate-malate-pyruvate shuttleAcetyl-CoA CarboxylaseThe ACC enzyme commits acetate to FA synthesisCarboxylation of acetyl-CoA to form malonyl-CoA is the irreversible, committed step in fatty acid biosynthesisACC uses bicar
27、bonate and ATP (and biotin)E.coli enzyme has three subunitsAnimal enzyme is one polypeptide with all three functions - biotin carboxyl carrier, biotin carboxylase and transcarboxylaseMammalian Acetyl-CoA Carboxylase Has Two Major Isoforms. There are two major isoforms of ACC. ACC1 occurs in adipose
28、tissue and ACC2 occurs in tissues that oxidize but do not synthesize fatty acids, such as heart muscle.Acetyl-CoA carboxylase -biotin transfers carboxyl groupsAcetyl-CoA carboxylase reactionThe Acyl Carrier Protein (ACP)Carrier of intermediates in fatty acid synthesisa 77 residue protein in E.coli -
29、 with a phosphopantetheine containing -SHIn terms of function, its a large CoAComparison of ACP and CoAAcyl carrier protein and coenzyme AFatty Acid Synthesis in AnimalsFatty Acid Synthase - a multienzyme complexDimer of 250 kD multifunctional polypeptidesthe roles of active site serines on AT & MTS
30、teps 3-6 repeat to elongate the chainMammalian FA Synthase DimerFirst three steps of FA synthesisStep45 of FA SynthesisStep 7 and Repeats of FA SynthesisElongation, beyond the 16-C length of palmitate, occurs in mitochondria. Endoplasmic reticulum enzymes also yield longer fatty acids. Fatty acid el
31、ongation within mitochondria occurs via b b-oxidation running in reverse, except that NADPH serves as electron donor for the final reduction step. Desaturation: Mammalian cells have limited ability to introduce double bonds in fatty acids. Some unsaturated fatty acids are dietary essentials, e.g., l
32、inoleic acid, 18:2 cis D D9,12. Mammalian cells cannot create double bonds at certain locations (e.g., D D12).Fatty Acid ModificationElongation and Desaturation of FARegulation of FFA OxidationRate-limiting enzymes is carnitine palmitoyl transferase I (CPT I)It is inhibited by malonyl-CoA when fatty
33、 acid synthesis is stimulated. This inhibition keeps the newly synthesized fatty acids out of the mitochondria and thus away from the -oxidation system. In heart muscle, an oxidative tissue that does not carry out fatty acid biosynthesis, contains ACC2, whose sole function appears to be the synthesi
34、s of malonyl-CoA to regulate fatty acid oxidation.Rate-limiting enzymes is ACC1ACC1 forms long, active filamentous polymers from inactive protomers Palmitoyl-CoA (product) favors monomersCitrate favors the active polymeric formPhosphorylation modulates citrate activation and palmitoyl-CoA inhibitionAMP-dependent protein kinase (AMPK), which phosphorylates (inactivates)ACCRegulation of FFA SynthesisRegulation of ACC1
