Increased climatic variability is resulting in an increase of both the frequency and the magnitude of extreme climate events. Therefore, cereals may be exposed to more than one stress event in the growing season, which may ultimately affect crop yield and quality. Here, effects are reported of interaction of water deficits and/or a high-temperature event (32°C) during vegetative growth (terminal spikelet) with either of these stress events applied during generative growth (anthesis) in wheat. Influence of combinations of stress on protein fractions (albumins, globulins, gliadins and glutenins) in grains and stress-induced changes on the albumin and gliadin proteomes were investigated by 2-DE and MS. The synthesis of individual protein fractions was shown to be affected by both the type and time of the applied stresses. Identified drought or high-temperature-responsive proteins included proteins involved in primary metabolism, storage and stress response such as late embryogenesis abundant proteins, peroxiredoxins and α-amylase/trypsin inhibitors. Several proteins, e.g. heat shock protein and 14-3-3 protein changed in abundance only under multiple high temperatures.

The interaction and physical/structural effects of aroma compounds, at high concentrations on dry native starch granules were studied using eight selected model compounds: acetaldehyde, dimethyl sulphide, diacetyl, allyl isothiocyanate, ethyl butyrate, citral, octanol and butyric acid. The maize, potato and pea starches used represent different typical structural and chemical starch characteristics. Retention of the different aroma compounds varied from a few to one hundred percent and starch was found to induce as well as reduce aroma evaporation depending on the aroma compound and the starch type. As deduced from DSC, powder XRD and SEM analyses, citral, butyric acid and octanol exerted specific effects on the starch granules manifested in local melting of crystalline layers and partial disruption of the granular meso structure. The most prominent effect was obtained with citral that generated surface wrinkles on B-and C-type polymorphic granules and aggregation of A-type polymorphic granules, decreased the melting temperature and suppressed the crystallinity of the starch.

The industrially important glucoamylase 1 is an exo-acting glycosidase with substrate preference for ␣-1,4 and ␣-1,6 linkages at non-reducing ends of starch. It consists of a starch binding and a catalytic domain interspersed by a highly glycosylated polypeptide linker. The linker function is poorly understood and structurally undescribed, and data regarding domain organization and intramolecular functional cooperativity are conflicting or non-comprehensive. Here, we report a combined small angle x-ray scattering and calorimetry study of Aspergillus niger glucoamylase 1, glucoamylase 2, which lacks a starch binding domain, and an engineered low-glycosylated variant of glucoamylase 1 with a short linker. Low resolution solution structures show that the linker adopts a compact structure rendering a well defined extended overall conformation to glucoamylase. We demonstrate that binding of a short heterobidentate inhibitor simultaneously directed toward the catalytic and starch binding domains causes dimerization of glucoamylase and not, as suggested previously, an intramolecular conformational rearrangement mediated by linker flexibility. Our results suggest that glucoamylase functions via transient dimer formation during hydrolysis of insoluble substrates and address the question of the cooperative effect of starch binding and hydrolysis.Glucoamylase (GA, 3 1,4-␣-D-glucan glucohydrolase; EC 3.2.1.3; glycoside hydrolase family 15) is an exo-acting glycosidase that cleaves ␣-1,4 and, less efficiently, ␣-1,6 linkages at non-reducing ends of starch and related oligo-and polysaccharides (1, 2). GA is industrially important in production of bioethanol, glucose, and fructose syrups. The glucoamylase 1 form (GA1) from Aspergillus niger has an N-terminal catalytic domain (CD) and a C-terminal starch binding domain (SBD) connected by a 69-amino acid-long linker that is decorated by short, predominantly mannose-containing O-glycosylations corresponding to a minimum content of 63 mol of hexose attached to about 32 serines and threonines (2-4) (see the schematic in Fig. 1). The functional role of the linker region is not fully understood. The isolated CD and the GA2 form (Fig. 1), which includes the linker and lacks the SBD, are both able to hydrolyze soluble substrates, but not starch granules, in contrast to GA1 (4 -6). Moreover, it has been shown that the addition of free SBD to a mixture of GA2 and starch granules increases the rate of hydrolysis (7). The SBD is, therefore, suggested to enhance the substrate accessibility by disentangling ␣-glucan helices on the surface of the starch granule rather than to act as an intramolecular guide directing the substrate chain to the active site pocket of the CD (7, 8). Conformational constraints on the linker are not required in this function and engineered linker variants, low-glycosylated or shortened ones inclusive, behave very similarly to wild-type GA1. Thus, the specific sequence of the linker seems to have a modest if any effect on the action of GA1 (9). This is much in contr...