Fructan metabolizing enzymes in Vernonia herbacea (VELL.) Rusby. / Enzimas do metabolismo de frutanos em Vernonia herbacea (VELL.) Rusby.

AUTOR(ES)
DATA DE PUBLICAÇÃO

2003

RESUMO

The occurrence of fructans has been reported in native species of a cerrado area of Reserva Biológica e Estação Experimental de Moji Guaçu. Vernonia herbacea, one of these species, presents underground organs named rhizophores which accumulate fructans of the inulin type as reserve carbohydrate. The seasonal growth pattern exhibited by plants of V. herbacea includes sprouting of buds from the rhizophores in spring, followed by a period of flower development and vegetative growth in summer, and dormancy in winter. The fructan content decreases during sprouting and flowering due to mobilization of this carbohydrate during the regeneration of the new shoots. Preliminary studies showed that FEH, the enzyme responsible for the mobilization of fructan, shows high activity only during sprouting. Fructan mobilization was detected by the increase in the amount of reducing sugar released, namely fructose. The aim of this work was to analyze the activities of the enzymes of fructan metabolism, FEH, SST, FFT and invertase and the fructan contents in rhizophores of plants induced to sprouting by defoliation. The enzyme activities were measured every 4 th day for a month after defoliation. Sprouting of new shoots started around the 13 th day, while an increase in FEH activity was detected between 13 and 20 days after defoliation. The results suggest that fructan depolimerization and sprouting are concomitant processes in V. herbacea that occur naturally during the phenological cycle; however, these processes can also be induced by defoliation during other stages of the phenological cycle. FFT seems to act together with FEH by catalyzing the decrease in fructan chain size during shoot regrowth, while SST was inhibited, possibly, due to interruption of sucrose supply to rhizophores from the aerial organs. The characterization and partial purification of FEH were done using rhizophores from plants which were induced to sprouting by defoliation. The optimal pH and temperature for FEH activity were pH4,5 and 30ºC, respectively. The substrate concentration curve exhibited a sigmoid shape, suggesting that FEH of V. herbacea is an alosteric enzyme. Additionally, this enzyme shows more specificity to b-2,1 than to b-2,6 linked fructan and higher affinity for short chain when compared to long chain fructans. Four fractions presenting FEH activity were partially purified by a combination of ammonium sulfate precipitation, affinity chromatography, and anion and cation exchange chromatographies. Two of these were submitted to size exclusion chromatography and the apparent molecular size for one of them was 21 kDa and for the other it was estimated to be between 39 and 155 kDa, due to the wide exclusion limit of the column used. The molecular size of the next two fractions were estimated by SDS- PAGE and the visualized bands corresponded to 81.3 and 57.5 kDa for the first fraction and to 57.5 kDa for the second one.

ASSUNTO(S)

enzima vegetal vegetable carbohydrate compositae metabolismo de carboidrato carboidrato vegetal vegetable enzyme carbohydrate metabolism

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