Asega Amanda Francine, do Nascimento João Roberto O, Schroeven Lindsey, Van den Ende Wim, Carvalho Maria Angela M
Seção de Fisiologia e Bioquímica de Plantas, Instituto de Botânica, CP 3005, 01061-970, São Paulo, SP, Brazil.
Plant Cell Physiol. 2008 Aug;49(8):1185-95. doi: 10.1093/pcp/pcn094. Epub 2008 Jun 20.
Variations in the inulin contents have been detected in rhizophores of Vernonia herbacea during the phenological cycle. These variations indicate the occurrence of active inulin synthesis and depolymerization throughout the cycle and a role for this carbohydrate as a reserve compound. 1-Fructan exohydrolase (1-FEH) is the enzyme responsible for inulin depolymerization, and its activity has been detected in rhizophores of sprouting plants. Defoliation and low temperature are enhancer conditions of this 1-FEH activity. The aim of the present work was the cloning of this enzyme. Rhizophores were collected from plants induced to sprout, followed by storage at 5 degrees C. A full length 1-FEH cDNA sequence was obtained by PCR and inverse PCR techniques, and expressed in Pichia pastoris. Cold storage enhances FEH gene expression. Vh1-FEH was shown to be a functional 1-FEH, hydrolyzing predominantly beta-2,1 linkages, sharing high identity with chicory FEH sequences, and its activity was inhibited by 81% in the presence of 10 mM sucrose. In V. herbacea, low temperature and sucrose play a role in the control of fructan degradation. This is the first study concerning the cloning and functional analysis of a 1-FEH cDNA of a native species from the Brazilian Cerrado. Results will contribute to understanding the role of fructans in the establishment of a very successful fructan flora of the Brazilian Cerrado, subjected to water limitation and low temperature during winter.
在草本斑鸠菊的根状茎物候周期中已检测到菊粉含量的变化。这些变化表明在整个周期中存在活跃的菊粉合成和解聚现象，并且这种碳水化合物作为储备化合物发挥着作用。1-果聚糖外切水解酶（1-FEH）是负责菊粉解聚的酶，其活性已在发芽植物的根状茎中检测到。去叶和低温是这种1-FEH活性的增强条件。本研究的目的是克隆这种酶。从诱导发芽的植物中收集根状茎，然后在5℃下储存。通过PCR和反向PCR技术获得了全长1-FEH cDNA序列，并在毕赤酵母中表达。冷藏增强了FEH基因的表达。Vh1-FEH被证明是一种功能性的1-FEH，主要水解β-2,1键，与菊苣FEH序列具有高度同源性，并且在10 mM蔗糖存在下其活性被抑制81%。在草本斑鸠菊中，低温和蔗糖在果聚糖降解的控制中发挥作用。这是关于巴西塞拉多本土物种1-FEH cDNA克隆和功能分析的首次研究。研究结果将有助于理解果聚糖在巴西塞拉多非常成功的果聚糖植物群建立中的作用，该植物群在冬季受到水分限制和低温影响。
