The involvement of lipoxygenase isozymes in several physiological processes of plants has been described but their role is not well understood and more biochemical studies are needed to elucidate the role of the "Lipoxygenase Pathway" in plant physiology. Thus, the biochemical and kinetic characterization of a lipoxygenases "pool" from soybean leaves was carried out. Two genotypes were used: IAC-100 (a normal variety having lipoxygenases in the seeds) and IAC-100 TN (genetically modified genotype, which is devoid of lipoxygenases in the seeds). The plants were submitted to the application of fatty acids (lipoxygenase substrates) on leaves. The results of the biochemical and kinetic studies of lipoxygenase isozymes from leaves of the two genotypes analysed showed that genetic removal of lipoxygenase from seeds did not affect the response of the plant to the treatment, since both genotypes showed similar results.

O objetivo deste trabalho foi avaliar a capacidade da planta de soja em responder à aplicação de ácidos graxos, substratos das lipoxigenases, pela "via das lipoxigenases" por meio da caracterização bioquímica e cinética do "pool" das lipoxigenases foliares. Dois genótipos de soja foram usados: um normal (variedade IAC-100) e outro desprovido das três lipoxigenases nas sementes (genótipo IAC-100 TN). As plantas foram tratadas com os ácidos araquidônico, linoléico e linolênico, no início do estádio V2 de desenvolvimento, até atingirem o estádio V3. Em seguida, os trifolíolos foram coletados 0, 24 e 48 horas após a última aplicação dos ácidos graxos. Os resultados mostraram um pico de atividade das lipoxigenases a pH 6,0 e 25°C de temperatura. Os valores de atividade das lipoxigenases nos dois genótipos foram maiores nos tratamentos que nos respectivos controles, e os valores de K M app diminuíram 48 horas após a última aplicação dos ácidos linoléico e linolênico no tratamento em relação ao controle. Estes resultados sugerem que a planta de soja responde à aplicação de ácidos graxos nas folhas, com o aumento na atividade das lipoxigenases, e que a remoção das lipoxigenases da semente não afetou a resposta da planta a esse tipo de tratamento, uma vez que IAC-100 e IAC-100 TN apresentaram comportamentos bioquímico e cinético semelhantes.

The objective of this work was to evaluate the capacity of soybean plant to respond to spraying of fatty acids, lipoxygenase substract, by lipoxygenase pathway through the biochemical and kinetic characterization of the "pool" of lipoxygenases from leaves. Two cultivars of soybean were used: one normal (cultivar IAC-100) and the other devoid of three lipoxygenases in the seeds (cultivar IAC-100 TN). The plants were treated with arachidonic, linoleic and linolenic acids at the beginning of the development stage V2, up to the stage V3. The trifoliates were collected at 0, 24 and 48 hours after the last spraying of fatty acids. The results of the biochemical and kinetic studies showed a peak of lipoxygenase activity at pH 6.0 and temperature at 25°C. The activity values for both genotypes were higher in the treatment than in the respective control, and the K M app values decreased 48 hours after the last spray with linoleic and linolenic acids, in the treatments towards respective controls. These results suggest that the soybean plant responds to the application of fatty acids in the leaves by increasing lipoxygenase activity, and that removing lipoxygenases from the seed has not changed the response of the plant to the fatty acids treatment, since IAC-100 and IAC-100 TN presented similar biochemical and kinetic behaviors.

Lipoxigenases (linoleato: oxigênio oxido-redutase - EC 1.13.11.12) são dioxigenases que catalisam a adição do oxigênio molecular ao sistema cis, cis, 1,4 - pentadieno dos ácidos graxos polinsaturados, formando hidroperóxidos dos ácidos graxos correspondentes. As lipoxigenases vegetais utilizam o ácido linolênico (C18:3) ou ácido linoléico (C18:2) como substrato e estão associadas a importantes processos fisiológicos, tais como: biossíntese de compostos regulatórios, crescimento e desenvolvimento, senescência, germinação de sementes, resposta a ferimento, proteína de reserva vegetativa e resistência a insetos e patógenos. Quando os tecidos da planta são danificados por patógenos ou mecanicamente, ocorre uma degradação seqüencial de lipídeos, cujo produto inicial são os hidroperóxidos resultante da ação das lipoxigenases. Entre os vários produtos formados, têm-se a traumatina, o ácido jasmônico, os aldeídos voláteis e os oxiácidos. Para avaliar a capacidade de as plantas de soja responderem, por meio da Via das Lipoxigenases, ao ataque do fungo causador da doença cancro-da-haste - Diaphorte phaseolorum (Cke. e Ell) f. sp. meridionalis - foram realizadas a caracterização bioquímico-cinética do pool de lipoxigenases foliares de plantas de soja (Glycine max (L.) Merril ) e a quantificação dos produtos da Via das Lipoxigenases, utilizando-se um cultivar resistente (FT-Cristalina RCH) e um susceptível (FT-Cristalina) ao patógeno. As plantas dos dois cultivares, infectadas com o patógeno ou apenas injuriadas mecanicamente, apresentaram valores de atividade específica de lipoxigenases maiores que os respectivos controles, em diferentes valores de pH e temperatura, e o pool de lipoxigenases dos dois genótipos apresentou cinética de Michaelis-Menten na faixa de concentração de substrato analisada. Ocorreu aumento dos níveis de inibidores de proteases e aldeídos totais, os quais são produtos dessa via bioquímica. Pelos parâmetros bioquímico e cinético, há a sugestão de uma resposta das plantas de soja através das Via das Lipoxigenases .

Lipoxygenases (linoleate: oxygen oxido-reductases, EC 1.13.11.12) are a class of enzymes that catalyze the addition of molecular oxygen to 1,4 - cis,cis - pentadiene motifs of unsaturated fatty acids to form hydroperoxide products. Plant lipoxygenases use linoleic (18:2) or linolenic (18:3) acids as substrates and are involved in important physiological process such as biosynthesis of regulatory molecules, growth and development, senescence, responses to wounding, vegetative storage protein and may directly mediate host resistance to insect, fungal and bacterial pathogens. When plant tissues are damaged, by pathogens or mecanically injured, a series of lipid degradation occurs, via lipoxygenase pathway, giving rise to hydroperoxydes. Among several products formed it can be found traumatin, jasmonic acid, volatile aldehydes and oxyacids. In this work we evaluated the capacity of the soybean plant to respond to the attack of stem canker fungi (Diaporthe phaseolorum (Cke. e Ell.) f. sp. meridionalis), through the lipoxygenase pathway. Leaves from resistant (FT-Cristalina RCH) and susceptible (FT-Cristalina) cultivars were used for kinetic evaluation of lipoxygenase and biochemical quantitation of products from the lipoxygenase pathway. Plants from both cultivars, infected with the pathogen or mecanically injured, showed values of lipoxygenase specific activity higher than the controls in differents pH and temperature values. The lipoxygenase pool of both cultivars showed kinetic of Michaelis-Menten within the range of substrate concentration assayed. There were an increase in protease inhibitors and of total aldehydes contents, which are products of the lipoxygenase pathway. The biochemical and kinetic parameters suggest a response of the soybean plant through the lipoxygenase pathway.