BCH/PPA 503 | Lecture Sixteen Web Notes
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Lecture Sixteen Lipids -- Fatty Acid Oxidation Purpose and Outline The purpose of todays lecture/discussion is to give an overview and some detail on specific subjects of fatty acid oxidation unique to plants and of significance in plant metabolism.
As mentioned in Lecture 15, unsaturated fatty acids are produced by desaturase-catalyzed oxidation of the more reduced saturated fatty acid molecules. Knowledge of fatty acid biosynthesis in plants has led to one of the more significant developments in the biotechnology industry. Vegetable oils (triglyceride) high in polyunsaturated fatty acids are too oxidatively unstable for cooking or spray oil applications. They therefore need to be hydrogenated. Triglycerides high in saturated or monounsaturated fatty acids are oxidatively stable, but monounsaturaded fatty acids are much healthier. The best all around cooking and spray oil is olive oil. Recently genetic engineering of soybeans for a reduction of a seed-specific ∆12 desaturase has resulted in the development of a soybean oil with the most favorable fatty acid composition on the market for cooking and spray applications as well as a number of industrial applications. See a web page of OptimumTM Quality Grains. How do you think this was achieved?
We have also already discussed fatty acid oxidation some in the synthesis of many of the monomers of wax, cutin and suberin. The recover of energy in the catabolism of fatty acids from triglycerides is mainly from ß-oxidation. This is similar in all organisms and as such is covered in general biochemistry and there is no need to cover the ß-oxidation pathway again in lecture here. I would like to emphasize a few points about ß-oxidation in plants such as the differences between plants and animals. Before β -oxidation can begin, fatty acids must 1st be released from triglycerides by a lipase and "activated" to an acyl-CoA e.g.:
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18:0 + CoA + ATP
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18:0-CoA + AMP + PPi = consumption of 2 high energy bounds or 2 ATPs
β -oxidation in plants occurs in glyoxysomes or peroxisomes, but in animals it usually occurs in mitochondria. This results in the lack of reduction of FAD to FADH in the 1st oxidation of the β-oxidation cycle in plants.
The β -oxidation of polyunsaturated fatty acids (PUFAs) such as 18:3-CoA has several differences compared with β-oxidation of 18:0. We do not have time to cover this except to point out that plants utilize PUFAs somewhat more efficiently than animals. Plants, but not animals, can convert fatty acids into glucose via β -oxidation and gluconeogenesis. In this process, acetyl-CoA resulting from β -oxidation is used to convert oxaloacetate to citrate, isocitrate and succinate in glyoxysomes: acetyl-CoA + oxaloacetate
citrate
isocitrate
succinate
The succinate is transferred to mitochondria where it is converted to oxaloacetate (OAA). The OAA is eventually converted to triose-P and finally to glucose in the cytosol as was discussed in Respiration lectures.
Unlike β-oxidation in which fatty acids are catabolized 2-C at a time, α-oxidation degrades fatty acids 1-C at a time. In mammals α-oxidation is important in the catabolism of β- methyl branched fatty acids. Although α-oxidation of fatty acids has been known to occur in plants for > 40 years, the metabolic role of this pathway is only recently beginning to be clarified. An α-dioxygenase has recently been characterized from plants that catalyzes α-oxidation (Hamberg et al., 1999). This α-dioxygenase, which is inducible by pathogens and is therefore also known as pathogen-inducible oxygenase (PIOX), catalyzes the formation of long chain aldehydes and hydroxy fatty acids as well as odd chain fatty acid catabolites (Fig. 3 Hamberg et al., 1999):
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BCH/PPA 503 | Lecture Sixteen Web Notes
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The immediate product of this α-dioxygenase is a dioxygenated fatty acid derivative with a hydroperoxy group on carbon 2. This is further metabolized to the products memtioned above and also can serve as a precursor for peroxygenase (see below) (Fig. 10 Hamberg et al., 1999):
In the remainder of this lecture I will mainly focus on the formation and metabolism of additional oxylipins or oxygenated derivatives of fatty acids.
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Oxylipin metabolism Fatty acids esterified to phospholipids, especially phosphatidylcholine, can be converted by epoxygenase or hydroxylase into epoxy or hydroxy derivatives. Fatty acids can also be converted by lipoxygenase into hydroperoxy derivatives. Most lipoxygenases are thought to act on fatty acids which are first "released" by phospholipase or lipolytic acyl hydrolase before peroxidation. Lipoxygenase can peroxidize linoleic or linolenic acids forming hydroperoxy (HOO) linoleic or linolenic acids. The primary products of lipoxygenase, the fatty acid hydroperoxides usually do not accumulate in plant tissues and can be rapidly converted into additional products by one of three known pathways. Most plant tissues appear to have the hydroperoxide lyase and allene oxide synthase branches of the lipoxygenase pathway. To date, peroxygenases have only been demonstrated in seedlings of some plants. Hydroperoxide lyase can cleave fatty acid hydroperoxides forming six and 12 carbon aldehydes. Many plants have the capacity to convert the six carbon aldehydes into alcohols. Allene oxide synthase converts the fatty acid hydroperoxides into an unstable epoxy intermediate that can be converted into a cyclic ketone by a cyclase. A series of subsequent reactions can result in formation of jasmonic acid, which can be converted into methyl jasmonate. Peroxygenases convert fatty acid hydroperoxides into epoxides, which can be converted to hydroxides by hydroxylases. This is illustrated in the following figure:
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An important aspect of fatty acid oxidation is the formation of biologically active fatty acid derivatives. This can be illustrated by the "arachidonate cascade" in animals, which generates numerous important metabolic mediators known as eicosanoids including prostaglandins (PG), thromboxanes (TX) and leukotrienes. The arachidonic cascade is initiated by release of free arachidonate by activation of phospholipase C (PLC) and diacylglycerol lipase or phospholipase A2 (PLA2) followed by oxidation by cyclooxygenase or lipoxygenase (LOX):
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The 1st committed step in eicosanoid formation involves oxidation of arachidonate to a hydroperoxide by LOX or cyclooxygenase. Higher plants have a "linolenate cascade" that generally resembles the arachidonate cascade:
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LOX is an enzyme which appears to be ubiquitous in eukaryotic organisms. LOX activity has been detected in multiple tissues of all plant species examined. Cyclooxygenase has not been detected in higher plants, but 3 enzymes in plants, LOX, allene oxide synthase and allene oxide cyclase, carry out an equivalent reaction. Interestingly, the natural plant product and 1st drug, acetylsalicylic acid or aspirin, is a potent inhibitor of both cyclooxygenase in animals and the combined reactions generating prostaglandin-like products such as jasmonic acid in plants. This is also true of other analgesic agents such as acetaminophen and ibuprofen. LOX catalyzes the peroxidation of polyunsaturated fatty acids which in higher plants and some higher fungi would be mainly linoleic (18:2) and linolenic (18:3) acids and in some cases hexadecatrienoic (16:3) acid. In animals, lower plants and lower fungi, longer chain polyunsaturated fatty acids such as arachidonic acid (20:4) (as well as 20:5 and 22:6) are more common LOX (and cyclooxygenase) substrates. The product of LOX, hydroperoxy fatty acids, are metabolized via at least four routes in higher plants to various products (see illustration below) which have biological activity and/or are important flavor and aroma compounds. These products of LOX have been referred to as octadecanoids because of their analogy to eicosanoids. Another term, oxylipins, proposed by Hamberg and Gardner, may be preferable since LOX products can be derived from 16:3. The fatty acid hydroperoxides produced by LOX can be converted into 1) keto fatty acids by other activities of LOXs; 2) hydroxy and epoxy fatty acids by cooxidative reactions of peroxygenase; 3) cleaved into two aldehydes by hydroperoxide lyase (HL) or 4) converted into an unstable allene oxide by allene oxide synthase (AOS). The biological activity of the keto fatty acids is unknown. The hydroxy and epoxy fatty acids produced by peroxygenase may have a role in cutin biogenesis and pest defense. HL products may have a role in pest defense and appear to act as pollinator and herbivore attractants especially for flowers and fruits. They are also important flavor and aroma compounds in foods, beverages and perfumes. Products of AOS, especially jasmonic acid (JA) and methyl jasmonate (MJ), have generated intense interest and appear to have numerous biological activities. These include:
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Oxylipin Formation from Linolenic Acid in Plant Tissues [IF: Isomerization factor(s)]:
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The mechanism of the LOX (lipoxygenase) catalyzed polyunsaturated fatty acid peroxidation is fairly well understood. LOX is a non-heme single iron containing dioxygenase which catalyzes the peroxidation of cis,cis -1,4 pentadiene structures:
In its oxidized state, Fe3+, LOX can abstract a proton from a PUFA such as linolenic acid. In the following example, a 13-LOX abstracts a proton from C11 of linolenic acid reducing LOX ( Fe2+) and producing a fatty acid free radical. While the substrate is still bound to LOX, the free radical shifts to C13 and the 12-13 cis double bound shifts to C11-C12 and becomes trans forming a conjugated diene structure. In the presence of O2 a peroxy radical can be formed that can be reduced to a hydroperoxide by the bound LOX enzyme. This results in the reoxidization of LOX which is then released. The hydroperoxy fatty acid can be cleaved into two aldehydes by hydroperoxide lyase also common in plant tissues:
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Of particular interest recently in plant biology has been the AOS or jasmonic acid pathway. This is increasingly being recognized as being of fundamental importance in signal transduction in response to biotic and abiotic stresses. The formation of these compounds is frequently elicited by pest attack including wounding and other stresses causing transient accumulation of intermediates of the "linolenate cascade":
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Especially interesting is the induced release of phermone-like compounds that attract predators and parasites to plants being fed upon by herbivorous insects. Koch et al. (1999) find that there are 4 groups of structurally distinct types of compounds from the linolenate cascade with different effects on release of attractants to beneficial insects: a.) 18:3 and acyclic derivatives. b.) C18 cyclopentanoid fatty acids. c.) C12 cyclopentanoid fatty acids especially epi-JA and JA. d.) Amino acid conjugates of linolenate cascade intermediates. This is illustrated in Fig. 1 of Koch et al. (1999):
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Do you see anything wrong with this figure?
Reading Assignment for Thursday, March 9 -- see the beginning of Lecture 17 web notes which will be posted by March 7 at the latest
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