Biosynthesis of Fatty Acids

Transcription

1 Biosynthesis of Fatty Acids CH 511 ource b-oxidation of FA Glycolysis of glucose n H CoA Acetyl CoA CoA Malonyl CoA Carboxylation of acetyl CoA H 3 C CH 2 CH 2 n-1 CoA aturated thioesters aturated fatty acids Dr. olomon Derese 38

2 CH 511 The processes of fatty acid biosynthesis are known to be catalyzed by a multienzyme complex known as Fatty Acid ynthase (FA). Fatty acid synthase is arranged around a central Acyl Carrier Protein (ACP), which contains a protein bound pantetheine chain [Condensing Enzyme (HE condensing )] similar to the long chain of Coenzyme A, with six distinct catalytic centers. Dr. olomon Derese 39

3 H N H N H H - P er FA ACP CH 511 NH 2 H N N - P - P N N N N H H H Coenzyme A - P H Dr. olomon Derese 40 -

4 CH 511 Dr. olomon Derese 41

5 Fatty Acid ynthase (FA) TE H AT CE MT ACP KR H CH 511 pantetheine chain ER DH AT= Acetyl transferase MT= Malonyl transferase CE= Condensing enzyme ACP= Acyl Carrier Protein KR= Keto Reductase ER= Enoyl Reductase DH= DeHydratase TE= ThioEsterase Dr. olomon Derese 42

6 CH 511 Before acetyl CoA and malonyl CoA can be used in biosynthetic reactions, they have to be attached to the fatty acid synthase. In this reaction the CoA group of acetyl CoA and malonyl CoA are substituted by the thiol groups (H) of HE condensing and ACP, respectively, of FA. The thiol groups are behaving as nucleophiles and the CoA group as a leaving group in a nucleophilic acyl substitution reaction. Dr. olomon Derese 43

7 CH 511 nce they are anchored to the fatty acid synthase, a carbon-carbon bond formation reaction occurs. Acetyl-CoA and malonyl-coa themselves are not involved in the condensation step: they are converted into enzyme-bound thioesters, Acetyl-E condensing and Malonyl -ACP, respectively. Dr. olomon Derese 44

8 CH 511 The intermediates in fatty acid synthesis are covalently linked to the Acyl Carrier Protein (ACP). Dr. olomon Derese 45

9 Biosynthesis of Fatty Acids CH 511 CoA H Decarboxylative Claisen condensation H H H AT CE ACP CE ACP CE ACP Fatty Acid ynthase (FA) molecular machine REDUCTIN 1) KR 2) DH 3) ER n H TRANLCATIN H TE H CE ACP CE ACP n H CE ACP AT= Acetyl transferase MT= Malonyl transferase CE= Condensing enzyme ACP= Acyl Carrier Protein KR = Keto Reductase ER = Enoyl Reductase DH= Dehydratase TE= Thioesterase Dr. olomon Derese 46

10 teps in the Biosynthesis of Fatty Acids tep I CH 511 Transfer of the acetyl group of acetyl CoA to the Condensing Enzyme (H-E condensing ). CoA + H-Econdensing E condensing + CoAH tep II Transformation of Malonyl CoA into Malonyl ACP. H CoA + H-ACP H ACP Malonyl CoA Malonyl ACP + CoA-H Dr. olomon Derese 47

11 tep III CH 511 Condensation of acetyl E condensing with Malonyl ACP. E condensing H ACP ACP + HE condensing Dr. olomon Derese 48

12 CH 511 It has been shown that the carboxylation of acetyl CoA goes with retention of configuration but the condensation with Malonyl ACP goes with inversion of configuration. A H H B H C CoA HC - 3 ATP Biotin H-ACP (Retention of configuration) Dr. olomon Derese 49 H H H B H H C ACP ACP Ecodensing C B (Inversion of configuration)

13 tep IV CH 511 tereospecific reduction mediated by NADPH producing 3-(R)-hydroxy intermediate exclusively. H + H ACP ACP H H s H N R NADPH C NH2 C NH2 N R NADP Dr. olomon Derese 50

14 tep V CH 511 yn elimination produces a 2-(E-enoyl)-ACP derivative, a trans-alkene. H H H ACP - H 2 ACP 2-(E-enoyl)-ACP Dr. olomon Derese 51

15 tep VI CH 511 The cycle is completed by further reduction mediated by NADPH to produce a saturated acyl- ACP. NADPH ACP ACP H Dr. olomon Derese 52

16 Ketone Methylene - Reduction CH 511 Achieved in 3 steps: H ACP KR H ACP H H H R -H 2 syn-elimination DH C NH2 N ACP R ER ACP H ACP Dr. olomon Derese 53

17 CH 511 Repetition of this cycle (steps II to VI) utilizing the newly formed acyl intermediate in place of acetyl CoA, leads to the lengthening of the carbon chain by two carbon atoms every cycle, to yield acyl-species of the general formula Me(CH 2 CH 2 ) n CH 2 CACP. This process terminates when the chain reaches C 16 or C 18, yielding palmitic or stearic acid, or their thiol esters. Dr. olomon Derese 54

18 CH 511 It is probable that as the chain length approaches C 16 - C 18, the active site thiol of the condensing enzyme has greater affinity for an acetyl-acp species. That is, steric or electronic effects hinder access acyl substrates bigger than C 16 - C 18 to the active site, and termination of the chain results. Dr. olomon Derese 55

19 CH 511 Me(CH 2 CH 2 ) n CH 2 CE condensing H 2 Me(CH 2 CH 2 ) n CH 2 C 2 H Extension of the chain beyond C 18 does occur but the ultimate chain length is rarely greater than C 22 -C 24, except in higher plants where the chain of up to C 30 are encountered. Dr. olomon Derese 56

20 CH 511 The combination of one acetate starter unit with seven malonates would give the C 16 fatty acid, palmitic acid, and with eight malonates the C 18 fatty acid, stearic acid. Note that the two carbons at the head of the chain (methyl end) are provided by acetate, not malonate, whilst the remainder are derived from malonate, which itself is produced by carboxylation of acetate. This means that all carbons in the fatty acid originate from acetate, but malonate will only provide the C 2 chain extension units and not the C 2 starter group. Dr. olomon Derese 57

21 tep I HC 3 ATP CoA Biotin H-E condensing H H-ACP CoA tep II CH 511 E condensing H ACP Malonyl-ACP tep III ACP NADPH + H + tep IV tep VI NADP H ACP ACP NADP NADPH + H + tep V Dr. olomon Derese 58 ACP

22 CH 511 The pathway can be visualized as a cyclic process in which the acetyl primer undergoes a series of Claisen condensation reactions with seven malonyl extender molecules and, following each condensation, the [b-carbon of the b-3-ketoacyl moiety formed is completely reduced by a threestep ketoreduction-dehydration-enoyl-reduction process. The saturated acyl chain product of one cycle becomes the primer substrate for the following cycle, so that two saturated carbon atoms are added to the primer with each turn of the cycle. Dr. olomon Derese 59

23 CH 511 dd numbered fatty acids The linear combination of acetate C 2 units explains why the common fatty acids are straight chained and possess an even number of carbon atoms. ther acyl CoA moieties (e.g. proinoyl CA, with three carbons) may also function as starter units in place of the usual starter acetyl CoA. Dr. olomon Derese 60

24 CH 511 The biosynthesis of fatty acids that possesses an odd number of carbon atoms is essentially similar to the biosynthesis of their even numbered counter parts. The synthesis of odd numbered fatty acids involve the use of the same extender unit, malonyl ACP, with an odd numbered starter unit. Dr. olomon Derese 61

25 E condensing tarter unit CH 511 Proinoyl E condensing H ACP Malonyl ACP Reduction ACP Extender unit ACP It would appear that it is the availability of unusual starters that determines whether normal or abnormal fatty acids are produced, rather than a requirement for special enzymes. Dr. olomon Derese 62

26 CH 511 Hydnocarpus wightiana (Flacourtiaceae) Dr. olomon Derese 63

27 Branched-chain Fatty acids CH 511 While straight chain fatty acids are the most common, branched chain fatty acids have been found to occur in mammalian systems. Branched fatty acids are formed either I. By priming the reaction with a branched starter (e.g 3-Methylbutyric CoA or 2- methylbutyl CoA) CoA CoA 2-Methylbutyric acid 3-Methylbutyric acid Dr. olomon Derese 64

28 CH 511 II. By using an alkylated malonyl CoA extender unit. Dr. olomon Derese 65

29 Unsaturated fatty acids CH 511 The majority of naturally occurring unsaturated fatty acids are in the C18 series. Acids shorter than C14, or higher than C22, are rare. ome representative examples are: H C 2 H Palmitoleic acid Dr. olomon Derese 66

30 H CH 511 C 2 H leic acid H (>80% of oil in olive oil) H Linoleic acid g-linoleic acid Dr. olomon Derese 67

31 Geometry of double bonds CH 511 It will be noted that most of the double bonds in the examples given possess the cis (Z) - stereochemistry while trans double bonds are found in fatty acids, they occur more rarely. The cis double bond has an important biological significance: by introducing a bend in the alkyl chain it prevents the hydrophobic chains of the acyl groups in fats, phosphoglycerides form compact aggregates maintaining the fluidity of fat depot and cell membranes. Dr. olomon Derese 68

32 CH 511 aturated fatty acid Trans unsaturated fatty acid Cis unsaturated fatty acid Dr. olomon Derese 69

33 CH 511 The typical pattern in polyunsaturated fatty acids is to have methylene groups flanked by two double bonds a methylene interrupted pattern of unsaturation. Dr. olomon Derese 70

34 Biosynthesis of unsaturated fatty acids CH 511 There are two common routes towards the biosynthesis of unsaturated fatty acids depending on the organism: I. Anaerobic route (occurs in some bacteria) - Proceed in the absence of oxygen II. Aerobic route (in animals and plants) - An absolute requirement for oxygen Dr. olomon Derese 71

35 CH 511 The aerobic process is by far the most common, and operates in yeasts, certain bacteria, algae, higher plants and vertebrates. The aerobic route directly introduces a double bond into a fatty acid precursor by a process known as oxidative desaturation, which is regulated by desaturase enzymes. The anaerobic pathway is mainly confined to anaerobic bacteria. Dr. olomon Derese 72

36 I. Anaerobic route CH 511 The fact that unsaturated fatty acids are synthesized in the absence of oxygen is apparent from their wide spread occurrence in anaerobic bacteria. However, only monounsaturated fatty acids are synthesized. Dehydrogenation occurs during chain elongation. Dr. olomon Derese 73

37 Anaerobic route to oleic acid CH 511 Desaturase H ACP cis-double bond leic acid Dr. olomon Derese 74

38 CH 511 This is an anaerobic route as no oxidation is required (the double bond is already there it just has to be moved) and is used by prokaryotes such as bacteria. Dr. olomon Derese 75

39 II. Aerobic route CH 511 Production of unsaturated fatty acids (insertion of double bonds) requires molecular oxygen. In an oxidation step, hydrogen is removed and combined with 2 to form water. Dehydration occurs after the required chain length of fatty acid is formed leading to mono- and poly-unsaturated fatty acids. Dr. olomon Derese 76

40 CH 511 This is apparently accomplished by syn elimination of a vicinal pair of pro-r hydrogen atoms, resulting in the formation of a cis-double bond exclusively. The double bond is usually introduced between C-9 and C-10. Dr. olomon Derese 77

41 CH 511 Example Dr. olomon Derese 78

42 CH 511 Most organisms possess a 9-desaturase enzyme that introduces a cis double bond into a saturated fatty acid at C-9. The position of further desaturation then depends very much on the organism. Non-mamalian enzymes tend to introduce additional double bonds between the existing double bond and the methyl terminus. Animals always introduce new double bonds towards the carboxyl group. Dr. olomon Derese 79

43 CH 511 In plants 12 9 xidative Desaturation 9 H In animals 6 H Linoleic acid g-linoleic acid Dr. olomon Derese 80

44 Higher plants CH 511 Lower plants C 2 H Animals Insects Dr. olomon Derese 81

45 CH 511 The first double bond introduced into a saturated acyl chain is generally in the 9 position so that substrates for further desaturation contain either a 9 double bond or one derived from the 9 position by chain elongation. Just like 9 desaturation that inserts the first double bond, further desaturation is an oxidative process requiring molecular oxygen. Dr. olomon Derese 82

46 CH 511 Animal systems cannot introduce double bonds beyond the C-9 position. Thus, second and subsequent double bonds are always inserted between an existing bond and the carboxyl end of the acyl chain, never on the methyl side. Plants, on the other hand, introduce second and third double bonds between the existing double bond and the terminal methyl group. Dr. olomon Derese 83

47 CH 511 Consequently, double bonds are found at the C-9, C-6, and C-5 positions as a result of desaturation in animals, at the C-9, C-12 and C-15 positions in plants, and at the C-5, C-6, C-9, C-12 and C-15 positions in insects and other invertebrates. Plants: Further unsaturation occurs primarily in this region H 3 C n(h 2 C) (CH 2 ) m H Dr. olomon Derese 84 9 Animals: Further unsaturation occurs primarily in this region

48 Fatty Acid Modifications CH 511 The biosynthesis of fatty acids mainly yield straight chain saturated fatty acids with sixteen and/or eighteen carbons. ther fatty acids are obtained by structural modifications of these straight chain saturated acids. These modifications include: I. Chain elongation to give longer fatty acids. II. Desaturation, giving unsaturated fatty acids. Dr. olomon Derese 85

49 ACP CH 511 ACP TEARIC ACID All rganisms ACP Animals LEIC ACID 18:1 (9c) Desaturation towards carboxyl terminus ACP LINLEIC ACID 18:2 (9c, 12c) ACP Plants Fungi 18:2 (9c, 6c) -LINLENIC ACID 18:2 (9c, 12c, 15c) Dr. olomon Derese 86

50 CH 511 The most common mono unsaturated FA in animals are oleic acid (18:1(9c)) and palmitoleic acid (16:1(9c)). Fatty Acid Desaturase enzymes in animals can only introduce double bond up to C-9. Thus the important unsaturated fatty acids Linoleic (with C-9 and C-12 double bonds) and Linolenic acids (with C-9, C-12 and C-15 double bonds) can not be biosynthesized in animals including humans. Dr. olomon Derese 87

51 CH 511 They must be obtained from the diet. Plants have the enzymes necessary to synthesize these acids. uch fatty acids are described as Essential Fatty Acids (EFA). Linolenic acid, an omega-3 fatty acid, and linoleic acid, an omega-6 fatty acid are both found in soybean oil and other types of plant oils. Dr. olomon Derese 88

52 CH 511 Dr. olomon Derese 89

53 CH 511 Animals can metabolize these two fatty acids obtained from the diet to form longer and more unsaturated PUFAs to meet their metabolic needs. ince these two fatty acids must be obtained from the diet, they are considered to be essential fatty acids. Linoleic acid is the starting material from which the body makes arachidonic acid, the precursor for prostaglandins, the hormone like substance that regulates a wide range of body functions, including growth, wound healing and epidermal health. Dr. olomon Derese 90

54 CH 511 ANIMAL w 6 ACP ACP LINLEIC ACID 18:2 (9c, 12c) g-linleic ACID 18:3 (6c, 9c, 12c) Chain extension by reaction with malonate ANIMAL ACP ARACHIDNIC ACID 20:4 (5c, 8c, 11c, 14c) ACP DIHM g-linleic ACID 20:3 (8c, 11c, 14c) Dr. olomon Derese 91

55 CH 511 w 3 ACP ANIMAL 18:3 (9c, 12c, 15c) DEATURAE TEARIDNIC ACID ACP 18:4 (6c, 9c, 12c, 15c) -LINLENIC ACID Chain extension by reaction with malonate DEATURAE ACP EICAPENTAENIC ACID (EPA) 20:5 (5c, 8c, 11c, 14c, 17c) EICATETRAENIC ACID ACP 20:4 (8c, 11c, 14c, 17c) Dr. olomon Derese 92

56 Further tructure Modification CH 511 Ad H 3 C AM R leoyl CoA Electrophilic addition reaction CoA H CH 2 Dihydrosterculic acid NADPH CoA Desaturation CoA CoA Tuberculosteraic acid terculic acid CoA Dr. olomon Derese 93

57 Prostaglandins CH 511 The prostaglandins are a group of modified C20 fatty acids first isolated from human semen and it was recognized that they were synthesized in the prostate gland (hence the name). They are now known to occur widely in animal tissues, but only in tiny amounts, and they have been found to exert a wide variety of pharmacological effects on humans and animals. Dr. olomon Derese 94

58 CH 511 It is thought that they are moderators of hormone activity in the body, a theory that explains their far reaching biological effects. They are active at very low, hormone-like concentrations and can regulate blood pressure, contractions of smooth muscle, gastric secretion, and platelet aggregation. Imbalances in prostaglandins can lead to nausea, diarrhea, inflammation, pain, fever, menstrual disorders, asthma, ulcers, hypertension, drowsiness or blood clots. Dr. olomon Derese 95

59 CH 511 The basic prostaglandin skeleton is that of a cyclized C20 fatty acid containing a cyclopentane ring, a C7 side-chain with the carboxyl function, and a C8 sidechain with the methyl terminus H H 20 C7 11 H H Basic skeleton of Prostaglandin C8 Dr. olomon Derese 96

60 CH 511 Naturally occurring prostaglandins contain a cyclopentane ring, a trans double bond between C- 13 and C-14, and a hydroxyl group at C-15. The prostaglandins are produced by most mammalian cells. Prostaglandins are biosynthesized from three fatty acids, dihomo-g-linolenic (20:3 (8c,11c,14c)), arachidonic (20:4 (5c,8c,11c,14c)), and eicosapentaenoic (20:5 (5c,8c,11c,14c,17c)) acid, which yield prostaglandins of the 1-, 2-, and 3-series, respectively. Dr. olomon Derese 97

61 CH 511 These three fatty acids are obtained from the essential fatty acids linoleic and linolenic acid. H DIHM g-linleic ACID H ARACHIDNIC ACID EICATETRAENIC ACID H H H H H H H PGE 1 H H H PGE 2 PGE 3 The numerical subscripts indicate the number of carbon-carbon double bonds in the side chains. Dr. olomon Derese 98

62 Biosynthesis of Prostaglandins CH 511 ARACHIDNIC ACID H Biallylic hydrogen xygen is a diradical H H H Biallylic free radical H Resonance tabilized H Dr. olomon Derese 99

63 CH 511 A methylene flanked by double bonds on both sides is susceptible to free radical oxidation; free radical reaction allows addition of and formation of peroxide 2 radical. Dr. olomon Derese 100

64 H PGG 2 CH 511 H H H Peroxidase H H PGH 2 H Dr. olomon Derese 101

65 H CH 511 H PGH 2 Radical H H H PGE 2 cleavage of cyclic epoxide H +H,- H H H H H H PGD 2 H H H PGF 2 Dr. olomon Derese 102

66 CH 511 Prostaglandins are named in accordance with the format PGX, where X designates the functional groups of the five-membered ring. PGAs, PGBs, and PGCs all contain a carbonyl group and a double bond in the five-membered ring. The location of the double bond determines whether a prostaglandin is a PGA, PGB, or PGC. R 1 R R 1 1 R 2 PGA PGB PGC R 2 R 2 Dr. olomon Derese 103

67 CH 511 The letters following the abbreviation PG indicate the nature and location of the oxygencontaining substituents present in the cyclopentane ring. PGDs and PGEs are b-hydroxyketones, and PGFs are 1,3-diols. A subscript indicates the total number of double bonds in the side chains, and and b indicate the configuration of the two H groups in a PGF: indicates a cis diol and b indicates a trans diol. Dr. olomon Derese 104

68 CH 511 H R 1 H H PGD R 2 H H PGE1 H H PGE 2 H H H H H H H H PGF 2 PGF 1 Dr. olomon Derese 105