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Barley lines producing grains with either amylopectin-free or hyper-phosphorylated starches were engineered by transgenic methods. Amylopectin-free barley was generated by simultaneously silencing the three genes encoding the starch branching enzymes (SBEIIa, SBEIIb and SBEI) by a chimeric hairpin. This construct was inherited as a single locus with a 3:1 segregation, which makes the method useful for breeding as compared to combing alleles of the three different SBE genes segregating independently. Transgenic grains were wrinkled. This is a phenocopy of Mendel’s wrinkled peas, which were also based on a non-functional allele of an SBE gene. Amylopectin content was below detection level (< 1%) measured by both size exclusion chromatography (SEC) and differential scanning calorimetry (DSC). Scanning electron microscopy (SEM) showed that this amylopectin-free starch is organized into amorphic granules. Plants were propagated for three generations. Yield and grain phenotypes were determined in grains grown under semi-field conditions. Loss of SBE activity was compensated by the cereal grain by up-regulated gene expression of starch synthases. As an effect, yield loss is limited to 20%. This demonstrates for the first time a way for production of pure amylose in plants with limited yield loss. In a different barley line endosperm specific overexpression of glucan water dikinase from potato (StGWD) was conducted. The content of phosphate esters in starch from consecutive generations (T0 and T1) of transgenic grains was tenfold higher than from vector control and wild type grains. Amylose content was not affected in hyperphosphorylated grains. Hyper-phosphorylated starch granules had several pores on the surfaces, similar to pores seen on enzymatically semi-degraded granules. This provides support for the presence of a general mechanism in starch degradation in the plant kingdom where phosphorylation carried out by ectopic expression of StGWD tags barley starch granules for degradation by endogenous enzymes. Together this work shows two new strategies for in planta starch bioengineering of cereals. It demonstrates that bioengineering may be used to obtain novel and technologically interesting cereal starches, and to elucidate the complex pathways of starch biosynthesis and the roles of individual starch biosynthetic enzymes.

The b-D-glucans from the cell wall of Saccharomyces cerevisiae have shown in vitro affinity for zearalenone. For this reason, their utilization as dietary adsorbent, to reduce the bioavailability of zearalenone, is of practical interest. Our study used powerful devices to elucidate the spatial conformation and molecular sites of interaction between ZEN and b-D-glucans. In this respect, 1H NMR spectroscopy implicated the hydroxyl groups of the phenol moiety of zearalenone in the complexation by laminarin, a pure b-(1,3)-D-glucan. X-ray diffraction determined that laminarin displays the conformation of a single-helix with six b-D-glucopyranose residues per turn. At this stage, molecular modeling was useful to locate the interaction sites and to propose highly probable complexes of zearalenone with laminarin fragment. Interestingly, the b-(1,3)-D-glucan chain favors a very stable intra-helical association with zearalenone, nicely stabilized by b-(1,6)-D-glucans side chains. Both hydrogen bonds and van der Waals interactions were precisely identified in the complex and could thus be proposed as driving interactions to monitor the association between the two molecules

Native starches were extracted from 21 cultivars of four yam species representative of the yam population of Ivory Coast. They were first characterized for their proximate composition, starch physico-chemical properties (amylose content, particle size distribution, crystallinity, thermal properties and intrinsic viscosity). Some functional properties (swelling, solubility and pasting behaviour and paste clarity) were then determined. Analysis of variance and principal component analysis showed that three homogenous groups could be distinguished, mainly based on starch physico-chemical properties. The first group contained all yam starches of the D alata and the D cayenensis-rotundata complex species. It was characterized by a large diameter grain (approximately 25 [mu]m), a high amylose content (around 25% db), a high intrinsic viscosity (mean of 190 cm3 g-1), and a high apparent viscosity and clarity of the paste. The second group contained the D esculenta varieties, characterized by a small granule size (diameter 6 [mu]m), a low intrinsic viscosity (121 cm3 g-1), a high gelatinization enthalpy change (19 J g-1) and a low paste viscosity. The D dumetorum sample differed from the D esculenta group by having a pure A-type crystalline form and an opaque paste. A multiple regression showed that the volume fraction of the dispersed phase and native granule size (or amylose content) could account for close to 80% of the variability of paste apparent viscosity. Gel clarity appeared mainly linked to granule size, small granules from D dumetorum and D esculenta giving the most opaque gels.

Les amidons natifs de 21 cultivars ont été extraits de 4 espèces d'igname de Côte d'Ivoire. Afin de caractériser ces échantillons, des analyses chimiques, physiques, morphologiques et rhéologiques ont été réalisées. Les gels d'amidon de l'espèce D. alata sont les plus visqueux (valeur moyenne: 292 mPa.s; 4% bs) alors que le gel d'amidon de D. dumetorum ne dépasse pas 61 mPa.s . La clarté des gels varie de 9% chez D. dumetorum à 40,7% de transmitance chez D. cayenensis-rotundata. L'étude de la diffraction aux rayons X met en évidence une cristallinité de 36% pour toutes les espèces. Une analyse en composantes principales (ACP) montre que les amidons d'igname de Côte d'Ivoire peuvent être regroupés en 3 classes homogènes: 1) La classe regroupant la totalité des amidons d'igname des espèces D. alata et D. cayenensis-rotundata, est caractérisée par un grain de gros diamètre (25 [mu]m environ), une teneur élevées en amylose (voisin de 26% bs), une forte viscosité intrinsèque (190 cm3.g-1 en moyenne), une viscosité apparente élevée, et une enthalpie de gélatinisation faible (?H = 15 j.g-1), 2) la classe regroupant les D. esculenta, est caractérisée par une petite taille de grains (Ø 6 [mu]m), une viscosité intrinsèque faible (121 cm3.g-1), un ?H élevé (19 J.g-1); 3) la classe de D. dumetorum se différencie de la classe des D. esculenta par un type cristallin de type A pur. Une régression multiple montre que la viscosité apparente des gels est a la fois liée à la fraction dispersée et à la taille des grains d'amidon. Cette étude met en évidence la grande diversité des propriétés physico-chimiques et fonctionnelles des amidons d'igname et ouvre des perspectives de valorisation spécifique en industrie alimentaire.

International audience; Recently, debranching enzymes were shown to be required during starch biosynthesis either for shaping amylopectin structure or for preventing synthesis of abnormal soluble polysaccharides in the plasmid stroma. We now describe a novel mutation in Clamydomonas which results in significantly reduced granular starch deposition displaying major modifications in structure. This defect simultaneously leads to the accumulation of linear malto-oligosaccharides. The mutation co-segregates with the absence of an -1,4 glucanotransferase , known as disproportionating enzyme (D-enzyme). We have cloned the gDNA and cDNA sequences corresponding to the Clamydomonas Reinhardtii -1,4 glucanotransferase.We demonstrate co-segregation between allele specific variations evidenced through Southern blots using the -1,4 glucanotransferase gDNA as a probe and the corresponding genotype at the STA11 locus. These results establish STA11 as the locus encoding D-enzyme.While the D-enzyme was known to transfer -1,4 linked chain segments between small-size acceptor and donor malto-oligosaccharides, we show that the missing enzyme readily catalyses the incorporation of oligosaccharides into amylopectin external chains in vitro.We therefore propose that D-enzyme plays a key role in starch biosynthesis by efficiently clearing those malto-oligosaccharides released by debranching enzyme. Oligosaccharide clearance can proceed by either reinserting spliced glucans into external chains of the putative maturing preamylopectin molecule or by facilitating their hydrolytic or phospholytic degradation.

La structure des réseaux amylacés (33 % matière sèche) obtenus à partir d'amidons de maYs normal, modifiés par différents traitments (cuissonextrusion, cylindres chauffants et empesage) a été étudiée à l'aide de diverses méthodes physico-chimiques : analyse enthalpique différentielle, diffractométrie des rayons X et susceptibilité à l'hydrolyse acide. Après empesage ou traitement par cylindres chauffants, l'amylose cristallise indépendamment de l'amylopectine. Ce phénomène s'explique par la lixiviation aqueuse de l'amylose pendant le traitement thermique, suivie d'une séparation de phase au sein du volume occupé par les molécules d'amylose. Selon les conditions de mise en oeuvre, différents modes de cristallisation peuvent se produire pour l'amylose : - après empesage de l'amidon natif, la complexation de l'amylose conduit à la formation de cristallites de type V, formés par des chaînes de degré de polymérisation DP de 40-45, faiblement attaquables par l'acide chlorhydrique et fondant à 100-120°c ; - après un traitement par cylindres chauffants suivi d'une réhydratation pour avoir un gel, l'amylose cristallise dans un type B, avec des chaînes de DP 35-40 , et fondant à de hautes températures : 125-135°C avant hydrolyse acide et 165 °C après hydrolyse acide. Par contre, après cuisson-extrusion, !'amylose et l'amylopectine cocristallisent dans un système mixte. Dans tous ces gels, les chaînes externes (DP 15-25) de l'amylopectine cristallisent lentement dans une structure de type B. Ces cristallites qui sont très "minces" sont facilement érodés par l'acide et fondent à 50 °C.