A starch granule bound endoamylase and its possible role during cold acclimation of parenchyma cells in poplar wood (Populus X canadensis Moench robusta)

摘要

Three-to-five-year-old twigs of poplar (Populus X canadensis Moench robusta) and willow (Salix caprea) trees were used to study the starch grain bound alpha-amylase (EC 3.2.1.1) in the parenchyma cells of the wood. These exhibit a remarkable temperature dependent binding to starch grains. Its endoamylase activity was measured using the starch azure method. In addition, amylase activities were measured in the desalted supernatant of the crude wood extract and also in the starch grain fraction with amylopectin as substrate. The changes in the enzyme activities were followed during autumn, winter, and spring in relation to the changes in starch content and frost hardiness, which was followed with the LT50 method. In contrast to the soluble amylase activity that was present at a considerable level before autumn and showing relatively little changes during winter, starch grain bound endoamylase activity started in autumn, was high during winter and disappeared during spring. This endoamylase was removed from the starch grain surface by elevated levels of maltose at 25deg C without destroying its amylolytic activity. Its binding on starch grains at low temperature, which has been proven earlier, is thus considered to be one key for the understanding of cold-induced starch hydrolysis. Additional support of this view was obtained when twigs in the winter stage were stored at +10deg C and at -5deg C, respectively. A dramatic decrease in starch grain bound endoamylase occurred at 10deg C while almost no change was detected at -5deg C. In contrast, total amylase activity of the supernatant did not change. At the same time, starch resynthesis was found at 10deg C while the low level of starch was maintained at -5deg C. Both poplar and willow showed identical behaviour. The present results thus give additional evidence for the peculiar role of this endoamylase in the cold-induced starch break-down during autumn and winter, which in turn is a most essential prerequisite for the abundant accumulation of sucrose and its galactosides in these cells at the beginning of cold acclimation.