Enzymatic breakdown of raffinose oligosaccharides in pea seeds

Abstract

Both alkaline and acidic alpha-galactosidases (alpha-D: -galactoside galactohydrolases, E.C.3.2.1.22) isolated from various plant species have been described, although little is known about their co-occurrence and functions in germinating seeds. Here, we report on the isolation of two cDNAs, encoding for alpha-galactosidases from maturing and germinating seeds of Pisum sativum. One was identified as a member of the acidic alpha-galactosidase of the family 27 glycosyl hydrolase cluster and the other as a member of the family of alkaline alpha-galactosidases, which are highly homologous to seed imbibition proteins (SIPs). PsGAL1 transcripts, encoding for the ACIDIC alpha-GALACTOSIDASE, were predominately expressed during seed maturation and acidic enzyme activities were already present in dry seeds, showing little changes during seed germination. Compartmentation studies revealed that acidic alpha-galactosidases were located in protein storage vacuoles (PSVs). PsAGA1, encoding for the ALKALINE alpha-GALACTOSIDASE, was only expressed after radicle protrusion, when about 50% of RFOs have already been broken down. RFO breakdown was markedly decreased when the translation of the alkaline enzyme was inhibited, providing evidence that PsAGA1 indeed functioned in RFO degradation. Based on these data, we present an integrated model of RFO breakdown by two sequentially active alpha-galactosidases in pea seeds.

Soluble carbohydrates in developing and mature diaspores of polar Caryophyllaceae and Poaceae

“…Changes in the concentrations of hexose and sucrose that reach a developing embryo determine the success of embryogenesis, the transition from the histodifferentiation phase to the reserve accumulation phase, and the rate at which those reserves are accumulated (Borisjuk et al 2004;Weber et al 2005). In mature seeds, low-molecular-weight soluble carbohydrates such as sucrose and raffinose family oligosaccharides (RFOs) account for a small portion (less than 20 % of dry mass, DM) of storage reserves (Obendorf and Górecki 2012), but they are used already in the first hours of germination, which indicates that sucrose and oligosaccharides are important sources of energy during initial stages of seeds germination (Blöchl et al 2008;Lahuta and Goszczyńska 2009). In addition to sucrose and RFOs, fructans and polyhydroxy alcohols, both non-cyclic (mannitol, sorbitol) and cyclic (myoinositol and its isomers or methylated derivatives), increase the resistance of vegetative tissues to abiotic stressors (Valluru and Van den Ende 2011;ElSayed et al 2014).…”

Non-reducing sugar levels in beech (Fagus sylvatica) seeds as related to withstanding desiccation and storage

“…It is located in protein storage vacuoles in seeds (Herman and Shannon, 1985;Bassel et al, 2001;Feurtado et al, 2001). Its activity has been reported in the developed, dry and germinating seeds of monocots and dicots, particularly during the mobilization of RFOs (Obendorf, 1997;Pukacka and Wójkiewicz, 2002;Zhao et al, 2006;Blöchl et al, 2008). There are no data in the literature regarding the activity of this enzyme in stored seeds.…”

Molecular cloning and characterisation of a cDNA encoding a putative alkaline alpha-galactosidase from grapevine (Vitis vinifera L.) that is differentially expressed under osmotic stress

“…We investigated the putative catalytic site(s) of the deduced protein for two well conserved motifs which have been reported (Blöchl et al 2008) using in silico analysis among the different a-gal families (i.e., family 27 Table 3 Intensity ratios (Razegui vs. Syrah) calculated from leaf RNA blot hybridisation results with Vv-a-gal/SIP 5 0 -UTR1-and 5 0 -UTR2-specific cDNA hybridisation probes eukaryotic and plant alkaline a gals/SIPs). These motifs were shown to be required for enzymatic activity for the glycosyl hydrolase family 27.…”

“…Alkaline a-galactosidases have been initially described as a group of SIP of unknown function due to their occurrence in germinating seeds. Evidence was presented for their involvement in RFO degradation in early stages of developing seedlings (Blöchl et al 2008) and recently a role in raffinose phloem unloading in sink tissues in Arabidopsis was suggested (Peters et al 2010).…”