Colin F. Jenner

Identification of multiple isoforms of soluble and granule-bound starch synthase in developing wheat endosperm

We have investigated the nature and locations of isoforms of starch synthase in the developing endosperm of wheat (Triticum aestivum L.). There are three distinct granule-bound isoforms of 60 kDa (the Waxy gene product), 77 kDa and 100–105 kDa. One of these isoforms, the 77-kDa protein, is also present in the soluble fraction of the endosperm but it contributes only a small proportion of the total soluble activity. Most of the soluble activity is contributed by isoforms which are apparently not also granule-bound. The 60-kDa and 77kDa isoforms of wheat are antigenically related to isoforms of very similar size in the developing pea embryo, but the other isoforms in the endosperm appear to have no counterparts in the pea embryo. The significance of these results in terms of the diversity of isoforms of starch synthase and their locations is discussed.

Analysis of effects in wheat of high temperature on grain filling attributes estimated from mathematical models of grain filling

SUMMARY Compared with growth at 20/15 xC (day/night), exposure of wheat (Triticum aestivum L.) plants to moderately high temperature (30/25 xC) significantly decreased grain weight through shortening the duration of grain filling, combined with small (or no) positive increases in the rate of grain filling. Several mathematical models of grain filling were assessed for their suitability as means of analysing these effects of temperature. The ordinary logistic model was found to be the most appropriate model and was used for the analysis of grain filling responses in four cultivars differing in their responses. Genotypic variation in response to temperature was observed for both rate and duration of grain filling, but the variation for the duration of grain filling among cultivars was small at the higher temperature. Significant correlation was found between single grain weight with the rate, but not with the duration, of grain filling at high temperature, which indicated an important role for synthetic processes involved in grain filling in the temperature sensitivity of wheat cultivars. As they are independent traits, both rate and duration are required selection criteria for the improvement of heat tolerance. Responses of one attribute estimated from the logistic model, the inflection point of the course of grain filling, may give insight into a temperature response that is distinguishable from that associated with the duration of grain filling. The inflection point appears to be worth including as a criterion in selecting for high temperature tolerance in wheat.

Thermal Characteristics of Soluble Starch Synthase From Wheat Endosperm

The aim of the work reported in this paper was to characterise the thermal responses of soluble starch synthase (SSS) extracted from the endosperm of the developing wheat grain. Using partially purified preparations of SSS, the reaction obeyed Michaelis-Menten kinetics with both substrates amylopectin and ADPglucose. Both the Vmax and the Km varied with temperature. Values for Vmax were higher at 45oC compared with 25oC. However, the Km values for both substrates were also higher at 45oC than at 25oC indicating that the affinity of the enzyme for its substrates was reduced at the higher temperature. Over the temperature range 15-45oC, the Km for arnylopectin was minimal at 20oC, and rose exponentially between 25 and 45oC. Kinetic analyses indicated that the reaction was sequential and that the substrates could bind to the enzyme in either order. At 25oC the binding of one substrate to the enzyme increased the affinity of the complex for the second substrate but at 45oC these effects were abolished. These thermal characteristics of SSS could explain certain important features of the temperature responses of starch deposition in the wheat grain in vivo.

Nitrogen supply to the grain modifies the effects of temperature on starch and protein accumulation during grain filling in wheat

Effects of nutritional status on the responses to high temperature of 2 wheat cultivars that differed in their sensitivity to high temperature were investigated in plants grown in pots in environmentally controlled growth rooms. The availability of nitrogen to the grains was altered by changing the amount and timing of the nitrogen supplied to the plants, and also by trimming the ears. Single grain weight was significantly decreased in the plants grown at 30/25°C compared with those grown at 20/15°C. The effects of nitrogen level or trimming on the response of grain filling to temperature did not seem to be mediated through alterations in the availability of carbohydrates within the grains. Neither N level nor trimming had substantial effects on the deposition of starch, but both treatments altered the accumulation of protein, and the responses of protein accumulation to the effects of temperature. At the lower temperature, increasing the nitrogen supply resulted in greater single grain weight, more protein per grain, and higher grain protein percentage. At the higher temperature, raising the supply of N increased none of these attributes; the rate of protein accumulation was not accelerated by raising the temperature as much at high as at low N, and high N reduced the duration of protein deposition. There was a linear and positive relationship between the amounts of amino acids in the grain and the rate of accumulation of protein in the endosperm, and the effects of N level and trimming were associated with changes in the concentration of amino acids in the grains. High temperature conditions appear to lower the supply of amino acids to the grain in plants well supplied with nitrogen. There was a positive relationship between the rate of protein deposition and the amount of amino acids in the grain. Although raising the level of nitrogen resulted in an increase in amino acid levels in the grain at low temperature, there was no such increase at high temperature. Trimming, however, did increase grain protein and the rate of protein deposition at high temperature as well as the amino acid levels in the grain. One possible interpretation of this paradox arises from other work showing that under high temperature and/or water deficit there is an inadequate supply of soluble carbohydrates in the shoot to metabolise high levels of nitrogen. Differences in the responses of starch deposition and protein accumulation to N level, trimming, and temperature confirm that these two components accumulate in the grain independently of each other. Consequences of the interactions between nitrogen supply, temperature, and cultivar are discussed in relation to yield and quality.

Effects of high temperature on grain growth and on the metabolites and enzymes in the starch-synthesis pathway in the grains of two wheat cultivars differing in their responses to temperature

The effects of a sustained period of moderately high temperature were evaluated on the availability of substrate and the activity of starch synthase (ADP-glucose: 1,4-α-D-glucan 4-α-D-glucosyltransferase, EC 2.4.1.21) in the developing grains of two wheat Triticum aestivum L. cultivars differing in their tolerance to high temperature. Final grain weight was reduced by 33% in the least sensitive (cv. Kavko) and by 40% in the most sensitive (cv. Lyallpur) cultivar as post-anthesis temperature was raised from 20/15°C (day/night) to 30/25°C. The difference in the response of the two cultivars was mainly due to changes in the rate of grain filling at high temperature. The response of the rate of grain filling at high temperature, and the differential effects on the two cultivars, did not seem to be explained by an effect of temperature on the supply of assimilate (sucrose) or on the availability of the substrate for starch synthesis (ADP-glucose) in the grains. In vitro, but not in vivo, the differential responses of the efficiency (Vmax/Km) of soluble starch synthase in the two cultivars to an increase in temperature were associated with differences in the temperature sensitivity of grain filling. In vivo, the most remarkable difference between the two varieties was in the absolute values of the efficiency of soluble starch synthase, with the most tolerant cultivar having the highest efficiency.

Effect of surfactant treatment on swelling behaviour of normal and waxy cereal starches.

Starch swelling behaviour greatly affects its functionality in a food matrix. For some granular starches pre-treatment with the surfactant, sodium dodecyl sulphate, is known to dramatically affect swelling behaviour. The purpose of this study was to assess the effect of this surfactant treatment on the swelling behaviour of a variety of waxy and normal cereal starches. A concurrent mid-infrared spectroscopy study was carried out to investigate the chemical nature of variations in swelling behaviour. The native normal starches (amylose content 23-28%) showed significant variation in their pasting properties, which was largely eliminated by surfactant treatment. Surfactant treatment had less effect on the behaviour of the waxy starches (amylose contents 1-8%), which still exhibited some residual variation. Waxy durum appeared to be unique within the waxy starches, behaving more similarly to the normal starches. The spectroscopic data highlighted the carbohydrate fingerprint region as the main source of variation between samples.

Optimizing copper-bicinchoninate carbohydrate detection for use with water-elution high-performance liquid chromatography: A technique to measure the major mono- and oligosaccharides in small pieces of wheat endosperm

Abstract Modifications are described to the copper—bicinchoninate detection of reducing sugars to increase sensitivity when used in conjunction with water-elution high- performance liquid chromatography and post-column catalytic hydrolysis of some oligosaccharides to a reducing form. The lowest limit of detection, taken to be the amount of substance that produces a peak height twice the noise level, was about 1 ng for a number of reducing and non-reducing sugars. Colour formation was linear (

Short term transport of iron and copper to various parts of wheat grains

Increasing the micronutrient content of staple foods such as cereal grains is a way of addressing micronutrient deficiencies in humans. This strategy will only be successful if the micronutrients are distributed in parts of the grain that are generally consumed (i.e. the endosperm). To date there are limited reports of micronutrient distribution within wheat grains with most simply reporting content of the whole grain. We investigated the transport of radioactively labelled iron and copper into the grain of cultured wheat ears 18–22 days post anthesis. Ears were loaded for 24 hours and then the grains dissected into various components. The grains were dissected into: epidermis; percarp-testa (combined hypodermis/cross cells/tube cells/testa/nucellar layer); outer endosperm (with aleurone layer attached); central endosperm; embryo; combined tissues of the crease. The majority of the iron was transported into the endosperm (61%), followed by the crease (15%). The majority of the copper was found in the crease (51%) followed by the endosperm (24%). Samples taken from the inner part of the endosperm contained no detectable iron or copper suggesting that the iron and copper present in the endosperm was found in the aleurone layer. If these grains were milled industrially for white flour then most of the iron and copper in the grain would be lost. Although these measurements were short term, they reinforce the point that strategies to increase iron and copper transport into wheat grains must keep in view the destinations of iron and copper within the grain.