Membrane-Associated and Soluble Lipoxygenase Isoforms in Tomato Pericarp (Characterization and Involvement in Membrane Alterations)

Membrane-associated and soluble lipoxygenases from green tomato (Lycopersicon esculentum Mill. cv Ailsa Craig) fruit have been identified. Microsomal lipoxygenase was localized partly in the plasma membrane and tonoplast fractions. The possibilities of glycosyl-phosphatidylinositol or transmembrane polypeptide anchors in the membrane were ruled out by differential solubilization and temperature-induced phase separation in Triton X-114. High performance liquid chromatography of reaction products combined with polarography showed that tomato lipoxygenase is capable of specific oxygenation of fatty acids esterified in phospholipids. This possibility of direct action on membrane phospholipids strengthened the hypothesis of a role for lipoxygenase in plant senescence and membrane turnover. Membrane-associated lipoxygenase is polymorphic, with two forms differing by their isoelectric points (pls) (around 4.2 and 5.1). The pl of the soluble lipoxygenase corresponds to the minor microsomal enzyme, with a pl of 5.1. The charge-differing isoforms were separated and analyzed by western blotting using anti-soybean lipoxygenase antibodies. A single polypeptide with an apparent molecular weight of 92,000 was identified in each case for the soluble and microsomal enzymes. It is suggested that a charge modification of the soluble lipoxygenase allows its association with the membrane.     

Identification and characterization of lipoxygenase isoforms in senescing carnation petals

A membrane-associated lipoxygenase and a soluble lipoxygenase have been identified in carnation (Dianthus caryophyllus L. cv Rêve) petals. Treatments of microsomal membranes by nonionic or zwitterionic detergents indicated that lipoxygenase is tightly bound to the membranes. By phase separation in Triton X-114, microsomal lipoxygenase can be identified in part as an integral membrane protein. Soluble lipoxygenase had an optimum pH range of 4.9 to 5.8, whereas microsomal lipoxygenase exhibited maximum activity at pH 6.1. Both soluble and membrane-associated lipoxygenases produced carbonyl compounds and hydroperoxides simultaneously, in the presence of oxygen. The membranous enzyme was fully inhibited by 0.1 millimolar n-propyl gallate, nordihydroguaiaretic acid, or salicylhydroxamic acid, but the effect of the three inhibitors on the soluble enzyme was much lower. The soluble lipoxygenase is polymorphic and three isoforms greatly differing by their isoelectric points were identified. Lipoxygenase activity in flowers was maximal at the beginning of withering, both in the microsomal and the soluble fractions. Substantial variations in the ratio of the two forms of lipoxygenase were noted at different sampling dates. Our results allowed us to formulate the hypothesis of a strong association of one soluble form with defined membrane constituents.     

Lipid-body lipoxygenase is expressed in cotyledons during germination prior to other lipoxygenase forms

Lipid bodies are degraded during germination. Whereas some proteins, e.g. oleosins, are synthesized during the formation of lipid bodies of maturating seeds, a new set of proteins, including a specific form of lipoxygenase (LOX; EC 1.13.11.12), is detectable in lipid bodies during the stage of fat degradation in seed germination. In cotyledons of cucumber (Cucumis sativus L.) seedlings at day 4 of germination, the most conspicuous staining with anti-LOX antibodies was observed in the cytosol. At very early stages of germination, however, the LOX form present in large amounts and synthesized preferentially was the lipid-body LOX. This was demonstrated by immunocytochemical staining of cotyledons from 1-h and 24-h-old seedlings: the immunodecoration of sections of 24-h-old seedlings with anti-LOX antiserum showed label exclusively correlated with lipid bodies of around 3 m in diameter. In accordance, the profile of LOX protein isolated from lipid bodies during various stages of germination showed a maximum at day 1. By measuring biosynthesis of the protein in vivo we demonstrated that the highest rates of synthesis of lipid-body LOX occurred at day 1 of germination. The early and selective appearance of a LOX form associated with lipid bodies at this stage of development is discussed.Abbreviations DAPI 4,6-diamidino-2-phenylindole - DIC differential interference contrast - FITC fluorescein-5-isothiocyanate - LOX lipoxygenase This work was financially supported by the Deutsche Forschungsgemeinschaft, SFB 286 (I.F., A.N., H.K.) and SFB 363 (B.H., C.W.).     

Characterization of lipases from the lipid bodies and microsomal membranes of erucic acid-free oilseed-rape (Brassica napus) cotyledons.

Lipase (triacylglycerol lipase, EC 3.1.1.3) activities have been reported previously in the lipid body and microsomal membranes of oilseed-rape (Brassica napus cv. Andor) seedlings, but conflicting data made it unclear whether there was one lipase in the lipid bodies, with the microsomal activity being attributable to fragments of lipid-body membrane, or if there were two separate lipase activities. In the present study, simultaneous characterization of the lipases under identical conditions showed they differed substantially in their pH-activity curves, kinetics and substrate specificities. (1) The kinetics of the microsomal lipase showed that the rate of lipolysis reached a plateau at concentrations above 5 mM, whereas the lipid-body lipase showed a linear increase in activity with substrate concentration up to 20 mM. (2) The pH optimum of the microsomal lipase was 7.5, whereas that of the lipid-body lipase was 9.0. The microsomal lipase was greatly inhibited at higher pH values, whereas the lipid-body lipase was much less affected. (3) Activity of the microsomal lipase was greatly diminished when substrates with longer chain length were used, and enhanced 4-fold if the substrates contained a single double bond. The lipid-body lipase was relatively unaffected by the type of fatty acid in the triacylglycerol. (4) SDS/polyacrylamide-gel electrophoresis showed little or no cross-contamination of the lipid-body and microsomal fractions. (5) The microsomal lipase activity comprised 75-80% of the total extracted.     

Physcomitrella patens has lipoxygenases for both eicosanoid and octadecanoid pathways

Mosses have substantial amounts of long chain C20 polyunsaturated fatty acids, such as arachidonic and eicosapentaenoic acid, in addition to the shorter chain C18 α-linolenic and linoleic acids, which are typical substrates of lipoxygenases in flowering plants. To identify the fatty acid substrates used by moss lipoxygenases, eight lipoxygenase genes from Physcomitrella patens were heterologously expressed in Escherichia coli, and then analyzed for lipoxygenase activity using linoleic, α-linolenic and arachidonic acids as substrates. Among the eight moss lipoxygenases, only seven were found to be enzymatically active in vitro, two of which selectively used arachidonic acid as the substrate, while the other five preferred α-linolenic acid. Based on enzyme assays using a Clark-type oxygen electrode, all of the active lipoxygenases had an optimum pH at 7.0, except for one with highest activity at pH 5.0. HPLC analyses indicated that the two arachidonic acid lipoxygenases form (12S)-hydroperoxy eicosatetraenoic acid as the main product, while the other five lipoxygenases produce mainly (13S)-hydroperoxy octadecatrienoic acid from α-linolenic acid. These results suggest that mosses may have both C20 and C18 based oxylipin pathways.     

Transient occurrence of lipoxygenase and glycoprotein gp49 in lipid bodies during fat mobilization in anise seedlings

The patterns of lipid-body proteins prepared at different stages of germination from seedlings of anise (Pimpinella ansium L.) were investigated. During the stage of fat mobilization, a set of proteins is synthesized de novo and transferred to existing lipid bodies. Analysis of the protein constituents, detected either by protein staining, concanavalin-A/peroxidase staining or immunoreaction on Western blots showed three distinct protein species: a lipoxygenase firmly integrated into the lipid bodies, a glycoprotein gp50 only extractable from the lipid bodies by treatment with sodium dodecyl sulfate and a glycoprotein gp49 solubilized by diethyl-ether treatment of lipid bodies. Both the lipoxygenase immunodetected on Western blots and gp49 stained with concanavalin-A/ peroxidase conjugate occurred transiently in the lipidbody fraction, reaching maximum concentrations between days 6 and 13 of germination. This behavior was in contrast to the decreasing level of the 18.4-kDa oleosin already present at the beginning of germination.Abbreviations gp glycoprotein - PAGE polyacrylamide gel electrophoresis - SDS sodium dodecyl sulfate We would like to thank Christiane Bernshausen and Willi Jonghaus for kind technical assistance. This research was supported by a grant from the Deutsche Forschungsgemeinschaft. I.F. is a recipient of the Friedrich Ebert-Stiftung.     

The lipoxygenases in developing soybean seeds, their characterization and synthesis in vitro

A number of lipoxygenase isoenzymes were identified in developing soybean (Glycine max L. Merrill cv Provar) seeds and two have been partially characterized. In a study of lipoxygenase level in developing soybean seeds, the enzyme content increased markedly during development. Comparisons of the lipoxygenases from mature soybean seeds and immature seeds by isoelectric focusing, chromatofocusing, sodium dodecyl sulfate polyacrylamide gel electrophoresis and peptide mapping identified two categories of isoenzyme. The isoenzymes from immature seeds were found by electron paramagnetic resonance spectroscopy to be isolated at least in part as the high spin iron(III) or active form of the enzyme in contrast to lipoxygenases from mature seeds which were isolated as electron paramagnetic resonance silent, high spin iron(II) species. The discovery of increased levels of lipoxygenases during seed development and their isolation in an active form suggests that the enzyme may play a physiological role during the maturation process. The incorporation of iron-59 from the nutrient medium into lipoxygenase during culture of immature seeds was indicative of de novo synthesis of the enzyme. The efficiency of the iron uptake was high, as indicated by the level of radioactivity found in the enzyme (one gram atom of iron per mole of lipoxygenase).     

Distinct lipoxygenase species appear in the hypocotyl/radicle of germinating soybean

Three lipoxygenase isozymes are synthesized in developing soybean (Glycine max [L.] Merr. cv Williams) embryos and are found in high levels in cotyledons of mature seeds (B Axelrod, TM Cheesbrough, S Zimmer [1981] Methods Enzymol 71: 441-451). Upon germination at least two new protein species appear which are localized mainly (on a protein basis) in the hypocotyl/radicle section. These lipoxygenase species appear also in seedlings of each of three lipoxygenase nulls (1×1, 1×2, and 1×3) deficient in one of the dormant seed lipoxygenases. The germination-associated species are distinguishable from dry seed lipoxygenase by their more acidic isoelectric points as revealed in isoelectric focusing gels. They are active from as early as 2 to at least 5 days after the start of imbibition. These germination-stimulated species qualify as lipoxygenase by their inhibition by the lipoxygenase inhibitors n-propyl gallate and salicyl hydroxamic acid and their lack of inhibition by KCN. Further, they are not active on the peroxidase substrate pair H₂O₂/3-amino-9-ethyl carbazole. They are recognized on Western blots by polyclonal antibodies to the seed lipoxygenase-1 isozyme and the major induced species has a molecular weight of approximately 100,000, similar to that of the cotyledon lipoxygenases. These lipoxygenases appear to be synthesized de novo upon germination since they comigrate with radioactive protein species from seeds germinated in [³⁵S]methionine.     

Lipoxygenase-generated hydroperoxides account for the nonphysiological features of ethylene formation from 1-aminocyclopropane-1-carboxylic acid by microsomal membranes of carnations

Several lines of evidence indicate that the conversion of 1-aminocyclopropane-1-carboxylic acid (ACC) to ethylene by microsomal membranes from carnation flowers is attributable to hydroperoxides generated by membrane-associated lipoxygenase (EC 1.13.11.12). As the flowers senesce, the capability of isolated microsomal membranes to convert ACC to ethylene changes. This pattern of change, which is distinguishable from that for senescing intact flowers, shows a close temporal correlation with levels of lipid hydroperoxides formed by lipoxygenase in the same membranes. Specific inhibitors of lipoxygenase curtail the formation of lipid hydroperoxides and the production of ethylene from ACC to much the same extent, whereas treatment of microsomes with phospholipase A₂, which generates fatty-acid substrates for lipoxygenase, enhances the production of hydroperoxides as well as the conversion of ACC to ethylene. Lipoxygenase-generated lipid hydroperoxides mediate the conversion of ACC to ethylene in a strictly chemical system and also enhance ethylene production by microsomal membranes. The data collectively indicate that the in-vitro conversion ACC to ethylene by microsomal membranes of carnation flowers is not reflective of the reaction mediated by the native in-situ ethylene-forming enzyme.Abbreviations ACC 1-aminocyclopropane-1-carboxylic acid - EDTA ethylenediaminetetraacetic acid     

Induction of accumulation and degradation of the 18.4-kDa oleosin in a triacylglycerol-storing cell culture of anise (Pimpinella anisum L.)

Two closely related anise cell-culture lines, Pa15 and Pa19, differ considerably in growth rate, potential to form somatic embryoids, triacylglycerol (TAG) storage and pattern of lipid-body proteins. Line Pa15 grows very fast (doubling rate: 3 d), mainly as single cells, exhibits a low potential for somatic embryogenesis and its TAG content is relatively low (5–20 mg TAG per g dry weight). In contrast, the line Pa19 shows lower growth rates (doubling rate: 8 d), tends to form clusters of somatic cells, has a higher TAG content (100–150 mg TAG per g dry weight), and somatic embryoids are easily induced. Under defined culture conditions, the TAG content of the line Pa19 can be increased to approximately 70% of that of ripe anise seeds (150 and 220 mg TAG per g dry weight, respectively). Polyclonal antibodies prepared against the most abundant protein (relative molecular mass 18.4 kDa) from the lipid-body fraction of anise seeds (Radetzky et al. 1993, Planta 191, 166–172) react also with a 18.4-kDa protein from the lipid-body fraction of cells of the Pa19 culture. In contrast, only fairly low levels of the 18.4-kDa oleosin were detected in Pal5. Limited sucrose supply in the medium resulted in TAG degradation and the concomitant decrease in the amount of immunodetectible 18.4-kDa protein in the Pa19 cell culture. Treatment with sorbitol, or abscisic acid and sorbitol in combination, enhanced TAG contents and also the amount of immunostained 18.4-kDa protein in the cell culture Pa19, whereas no effect was found on either TAG content or 18.4-kDa protein in the cell-culture line Pa15. The 18.4-kDa protein can be classified as an oleosin, a proposal which is supported by the similarity in molecular mass compared with other known oleosins, its occurrence in the lipid-body fraction and the fact that its amount correlates with the TAG content. The results of this study indicate that the Pa19 cell culture provides a valid model system for investigations of lipid storage and mobilization in higher-plant cells.Abbreviations ABA cis-abscisic acid - TAG triacylglycerol(s) - 2,4-D 2,4-dichlorophenoxyacetic acid The authors thank Christiane Bernshausen for kind technical assistance.     

Lipoxygenase heterogeneity in Pisum sativum

Antibodies raised against two pea (Pisum sativum L. cv. Birte) seed lipoxygenases have been used to analyze lipoxygenase heterogeneity in seeds and in other organs. At least seven different polypeptides were identified in vivo; five of these were identified as precursors synthesized in vitro. The developmental appearance of the seed polypeptides has been analyzed and early and late forms were identified. Limited N-terminal sequence data indicated further heterogeneity when compared with sequences predicted from cDNAs.Abbreviations cDNA complementary DNA - DAF days after flowering - HPLC high-performance liquid chromatography - Ig immunoglobulin - kb kilobase - M_r relative molecular mass - PAGE polyacrylamide gel electrophoresis - PVDF polyvinylidene difluoride - SDS sodium dodecyl sulphate - SSC 0.15 M sodium chloride, 0.015 M sodium citrate, pH 7.0 This work was supported by the Agricultural and Food Research Council via a grant-in-aid to the John Innes Institute. We acknowledge financial support from the Commission of the European Communities Biotechnology Action Programme; grant No. 0063-UK.     

Properties of a Lipoxygenase in Green Algae (Oscillatoria sp.)

A lipoxygenase preparation was obtained from green algae Oscillatoria sp. and was shown to differ from previous described lipoxygenases in the positional specificity and pH characteristics of the dioxygenation reaction. The enzyme had a pH optimum at 8.8 and was inactive at pH 6. Oscillatoria lipoxygenase converted linoleic acid into two products: 13-hydroperoxylinoleic acid (52%) and 9-hydroperoxylinoleic acid (48%). The molecular weight of the enzyme was estimated at 124,000. Esculetin was found to be the best inhibitor of the enzyme activity.     

Lipoxygenase pathway in olive callus cultures (Olea europaea)

Stimulation of the lipoxygenase pathway in olive fruit initiates a cascade of reactions that begins with the regio- and stereospecific di-oxygenation of polyunsaturated fatty acids containing a cis, cis-1,4 pentadiene moiety. Later products of the pathway include volatiles that influence the organoleptic properties of harvested olive oil. In this study, we have investigated lipoxygenase activity in olive callus cultures, and found that there is evidence of several isoforms of the enzyme with different pH optima and substrate specificities. Endogenous lipoxygenase activity was detected throughout the growth cycle of olive callus, particularly during the log phase of growth, suggesting that olive lipoxygenases are intimately involved in growth. The most prominent lipoxygenase activity in tissue cultures was found to be soluble but significant activities were detected in the plastid fraction. In addition, hydroperoxide lyase (HPL) activity was measured in the calli; both 13- and 9-HPL activities were found which were particulate.     

Immunological comparison of lipoxygenase isozymes-1 and -2 with soybean seedling lipoxygenases

An affinity-purified polyclonal antibody against soybean seed lipoxygenase-2 was prepared and used to characterize the immunological relatedness of lipoxygenase isozymes 1 and 2 and lipoxygenases from soybean seedling roots, hypocotyls, leaves, and cotyledons. All soybean lipoxygenases tested cross-reacted with the anti-lipoxygenase-2. Cross-reactivity of seed-derived lipoxygenases was evidenced by formation of a line of identity in double-diffusion tests, by positive results on an immunoblot, and by antibody precipitation of enzyme activity. Levels of anti-lipoxygenase-2, which inhibited lipoxygenase-2 activity, had no effect on lipoxygenase-1 activity. Root, hypocotyl, and leaf lipoxygenases did not form precipitation lines in double-diffusion tests but the anti-lipoxygenase-2 did inhibit and precipitate lipoxygenase activity from these sources as well as cross-react on immunoblots. All the cross-reactive lipoxygenases examined were found to have the same apparent molecular weight. Lipoxygenase activity found in soybean seedling roots, hypocotyls, leaves, and cotyledons is associated with proteins which are all immunologically related to the seed-derived enzymes.     

Trehalase transformation in silkworm midgut during metamorphosis

Summary The particulate trehalase from silkworm larval midgut was effectively solubilized by repeated freezing and thawing, and by incubation with snake venom and non-ionic detergents (Lubrol PX and WX and Triton X-100). With solubilization the activity was enhanced and the activation behaviour was dependent upon the developmental stage of silkworms, being highest (up to about 3-fold) at the spinning stage.When chromatographed on DEAE-cellulose columns separately, the enzyme solubilized by freezing and thawing and the soluble trehalases from feeding larval midgut were respectively eluted as single peaks, P I and P II. However, both P I and P II trehalases were demonstrated after solubilization of the particulate fraction from feeding larvae with Triton X-100, or after treatment of the midgut of spinning larvae by freezing and thawing.The apparent molecular weights of P I and P II trehalases as estimated by Sephadex G-200 chromatography were about 70,000 and 140,000, respectively. The optimum pH was 6.0 for P I and about 5.0 for P II trehalase. TheK _m values were about 1.0 mM for P I trehalase and 0.30 mM for P II trehalase.These results suggest that in feeding larval midgut there are two types of trehalase which are distinguishable from each other by intracellular localization, protein nature and kinetic properties. Furthermore, when the midgut undergoes metamorphosis, the P I enzyme found predominantly in feeding stages seems to be transformed to the P II enzyme via an intermediate form (Ppt-P II) with transitional properties.     

Activity of soybean lipoxygenase isoforms against esterified fatty acids indicates functional specificity

In soybean (Glycine max L.) vegetative tissue at least five lipoxygenase isozymes are present. Four of these proteins have been localized to the paraveinal mesophyll, a layer of cells that is thought to function in assimilate partitioning. In order to determine the role of the lipoxygenase isozymes within the soybean plant, the leaf lipoxygenases were cloned into bacterial expression vectors and expressed in Escherichia coil. The recombinant lipoxygenases were then characterized as to substrate preference, pH profiles for the most common plant lipoxygenase substrates, linoleic acid, and alpha-linolenic acid, and the reaction products with the substrates linoleic acid, alpha-linolenic acid, arachidonic acid, gamma-linolenic acid, and the triacylglycerol trilinolein. All five enzymes were shown to be (13S)-lipoxygenases against linoleic acid. The results of these assays also indicate that two of these isozymes are highly active against esterified fatty acid groups, such as those found in triacylglycerols. Lipid analysis of leaves from plants subjected to sink limitation conditions indicates that the soybean leaf lipoxygenases are active in vivo against both free fatty acids and esterified lipids, and that the quantities of lipoxygenase products found in leaf tissue show a positive correlation with the level of lipoxygenase in the leaf. Implications for the putative role of these enzymes in the paraveinal mesophyll are discussed.     

Molecular cloning and biochemical characterization of a lipoxygenase in almond (Prunus dulcis) seed.

We have characterized an almond (Prunus dulcis) lipoxygenase (LOX) that is expressed early in seed development. The presence of an active lipoxygenase was confirmed by western blot analysis and by measuring the enzymatic activity in microsomal and soluble protein samples purified from almond seeds at this stage of development. The almond lipoxygenase, which had a pH optimum around 6, was identified as a 9-LOX on the basis of the isomers of linoleic acid hydroperoxides produced in the enzymatic reaction. A genomic clone containing a complete lipoxygenase gene was isolated from an almond DNA library. The 6776-bp sequence reported includes an open reading frame of 4667 bp encoding a putative polypeptide of 862 amino acids with a calculated molecular mass of 98.0 kDa and a predicted pI of 5.61. Almond seed lipoxygenase shows 71% identity with an Arabidopsis LOX1 gene and is closely related to tomato fruit and potato tuber lipoxygenases. The sequence of the active site was consistent with the isolated gene encoding a 9-LOX.     

Resolution of the isoenzymes of soybean lipoxygenase using isoelectric focusing and chromatofocusing

Isoelectric focusing in thin-layer polyacrylamide gels has been applied to the analysis of the enzymes involved in the formation and destruction of peroxides in soybeans [Glycine max (L.)], lipoxygenases and peroxidases, respectively. As a result of differences in pH optima for catalytic activity, lipoxygenases were selectively detected by adjusting the pH employed for activity-specific staining. Type-1 lipoxygenase was revealed not only by staining based on the conversion of linoleic acid to hydroperoxide but also by two stains based on the reduction of the hydroperoxide. These methods were found to be suitable for the analysis and characterization of isoenzyme patterns in different soybean cultivars. A substantial difference in the distribution of lipoxygenases maximally active near pH 7 was observed for cultivars Provar and Vickery. A similar degree of separation of the isoenzymes was achieved on a larger scale using chromato-focusing in the pH range 7.4-5.0.     

Metabolic profiling of oxylipins in germinating cucumber seedlings--lipoxygenase-dependent degradation of triacylglycerols and biosynthesis of volatile aldehydes

A particular isoform of lipoxygenase (LOX) localized on lipid bodies was shown by earlier investigations to play a role in initiating the mobilization of triacylglycerols during seed germination. Here, further physiological functions of LOXs within whole cotyledons of cucumber (Cucumis sativus L.) were analyzed by measuring the endogenous amounts of LOX-derived products. The lipid-body LOX-derived esterified (13 S)-hydroperoxy linoleic acid was the dominant metabolite of the LOX pathway in this tissue. It accumulated to about 14 micromol/g fresh weight, which represented about 6% of the total amount of linoleic acid in cotyledons. This LOX product was not only reduced to its hydroxy derivative, leading to degradation by beta-oxidation, but alternatively it was metabolized by fatty acid hydroperoxide lyase leading to formation of hexanal as well. Furthermore, the activities of LOX forms metabolizing linolenic acid were detected by measuring the accumulation of volatile aldehydes and the allene oxide synthase-derived metabolite jasmonic acid. The first evidence is presented for an involvement of a lipid-body LOX form in the production of volatile aldehydes.     

Molecular characterization of L2 lipoxygenase from maize embryos

We investigated the expression and accumulation pattern of lipoxygenaseisoforms throughout the maize plant life. Two forms of lipoxygenase L1and L2 have been identified as acidic proteins of 100 kDa (pI 6.4) and90 kDa (pI 5.5-5.7) which accumulate in dry embryos and in variousorgans of maize seedlings. In young embryos, only the L2 form wasdetected and accumulation of L2 mRNA decreased during embryodevelopment. Identification of lipoxygenases from in vivo and in vitro synthesized proteins indicates that similar levels of both L1and L2 forms accumulated during treatment with abscisic acid, (ABA)gibberellic acid (GA₃) and jasmonic acid (JA). However,differences in the activity of both enzymes were detected. By using anantiserum directed against purified L2 we isolated and characterized apartial cDNA clone of maize embryos encoding a lipoxygenase. The deducedamino acid sequence of L2 cDNA shares 78% identity with the rice L2protein, and 51-56% identity with lipoxygenases from thedicotyledonous plants soybean and Arabidopsis/. DNA blotanalysis indicated that maize contains a family of lipoxygenase geneswhich are presently being characterized.