Endosperm Proteins Research Articles

Japanese Sake‐Brewing Rice Cultivars Show High Levels of Globulin‐like Protein and a Chloroplast Stromal HSP70

ABSTRACTWhite‐core is an important trait in the selection of rice (Oryza sativa L. ssp. japonica) for sake (Japanese rice wine) production. The present study was conducted to find differences between the endosperm protein pattern of white‐core–bearing sake‐brewing cultivars and those of normal rice grains. Protein patterns of white‐core sake‐brewing grains were compared with those of normal cultivars by two‐dimensional polyacrylamide gel electrophoresis (2D‐PAGE). Several proteins showing different amounts were detected. These were identified by microsequencing and found to be known carbohydrate‐metabolizing enzymes as well as proteins with no known involvement in carbohydrate metabolism. Two units of globulin‐like protein (GLP) and a chloroplast stromal heat shock protein 70 (ctHSP70) showed the greatest difference in the white‐core group. Analysis of amyloplast proteins showed slightly higher amounts of GLP and ctHSP70 in the amyloplast than in the cytosol. Also, time‐course analysis of developing rice grains indicated that both proteins were elevated during the later stages of grain development. From this we conclude that GLP and ctHSP70‐II are associated with white‐core appearance and thus behave as biochemical markers for this character in sake‐brewing rice cultivars.

Homoeologous recombination within bread wheat to develop novel combinations of HMW-GS genes: transfer of the Glu-A1 locus to chromosome 1D

In an attempt to improve the bread-making quality within hexaploid wheat by elaborating novel high-molecular weight glutenin subunits (HMW-GS) combinations useful in wheat-breeding programmes, a 1A chromosome fragment carrying the Glu-A1 locus encoding the subunit Ax2*, was translocated to the long arm of chromosome 1D. The partially isohomoeoallelic line, designated RR239, had a meiotic behaviour as regular as cv. Courtot. It was characterised using genomic in situ hybridization and microsatellite markers as well as biochemical and proteomic approaches. The translocated 1D chromosome had an interstitial 1AL segment representing in average 30% of the recombinant arm length that was confirmed by molecular analysis. The genetic length of the removed segment in chromosome 1DL was estimated to be at least 51 cM, and that of the interstitial 1AL translocation to be at least 33 cM. Proteome analysis performed on total endosperm proteins revealed variation in amounts, 8 spots and 1 spot being up- and downregulated, respectively. Quantitative variations in HMW-GS were observed for the Glu-A1 (Ax2*) and Glu-B1 (Bx7 + By8) loci in response to duplication of the Glu-A1 locus.

Enzymatic properties of the 20S proteasome in wheat endosperm and its biochemical characteristics after seed imbibition

The 20S proteasome from wheat (Triticum aestivum L., Yangmai 158) endosperm was purified to apparent homogeneity by three sequential centrifugations and gradient PAGE (GPAGE). The purified 20S proteasome clearly cleaved peptidyl-arylamide bonds in the model synthetic substrates Z-GGL-AMC and Z-GGR-AMC, which are used to reflect chymotrypsin-like and trypsin-like activity, respectively. For both substrates, the optimum pH was 8.0, but the optimum temperatures for chymotrypsin-like and trypsin-like activity were 55 degrees C and 37 degrees C, respectively. Both enzyme activities were clearly inhibited by MG115 and PMSF. Polyubiquitinated proteins remained constant from 0 to 7 days after seed imbibition, but caseinolytic activity and the amount of the 20S proteasome associated with the aleurone layer decreased from 1 to 2 days after imbibition (DAI), then increased from 2 to 4 DAI, and reached a maximum at 4 DAI that was retained until 7 DAI. An increase was seen in the mRNA level of the beta5 subunit of the 20S proteasome from 2 DAI, and caseinolytic activity and the amount of the 20S proteasome increased from 3 DAI onwards. In addition, the main storage proteins of the wheat endosperm could not be hydrolyzed by the 20S proteasome. The evidence suggests that the main role of the 20S proteasome may not be to degrade massive proteins of the wheat endosperm after seed imbibition.

Identification of Proteinaceous Inhibitors of a Cysteine Proteinase (an Arg-Specific Gingipain) from Porphyromonas gingivalis in Rice Grain, Using Targeted-Proteomics Approaches

Porphyromonas gingivalis is known to be a major etiologic agent in the onset and progression of chronic periodontitis. Among various virulence factors that this bacterium produces, Arg- and Lys-specific cysteine proteinases (gingipains) are believed to be major determinants of the pathogenicity of P. gingivalis. Here, we report on our finding that there are inhibitors of these cysteine proteinases in a rice protein fraction. Comprehensive affinity chromatography and MS analyses resulted in the identification of 17 Arg-gingipain (Rgp)-interacting proteins in the rice endosperm. Of these, four proteins (i.e., a 26 kDa globulin, a plant lipid transfer/trypsin-alpha amylase inhibitor, the RA17 seed allergen, and an alpha amylase/trypsin inhibitor) were estimated to account for 90% of the Rgp inhibitory activity in the rice protein fraction, using a two-dimensional gel system of double-layer reverse zymography. In addition, a synthetic peptide derived from an Rgp-interacting protein, cyanate hydratase, could inhibit the growth of P. gingivalis and showed inhibitory activity against both the Arg- and Lys-gingipains. These results suggest that these rice proteins may be useful as nutraceutical ingredients for the prevention and management of periodontal diseases.

A Comparative Analysis of Embryo and Endosperm Proteome from Seeds of Jatropha curcas

Jatropha curcas is an important economic plant for biodiesel, which is extracted mainly from the endosperm of its mature seeds. Despite the morphological and functional differences between the embryo and endosperm, proteomic characteristics of the two tissues are not yet known. Similar proteomic profiles were observed in the two-dimensional gel electrophoresis maps from the two tissues. There were 380 and 533 major protein spots in the embryo and endosperm, respectively. Fourteen identical spots, showing a notable change, were selected and identified by tandem mass spectrometry. Among these proteins, dihydrolipoamide acetyltransferase (spot 27) participates in tricarboxylic acid cycle, which is an amphibolic pathway. The two parts both included proteins related to stress (spots 8, 115, 118, 125, 130) and signal transduction (spots 7, 100, 108). According to the volume percentage of proteins in embryo and endosperm, the proteins in endosperm (spots 54, 61, 73) were catabolism-related enzymes and reserves to provide the nutrition for seed germination; the proteins in embryo (spots 27, 62, 122) were inclined to anabolism and utilized the nutrition from the endosperm to generate a new life.

Antioxidant activities of the rice endosperm protein hydrolysate: identification of the active peptide

The defatted rice endosperm protein (REP) was digested using five different proteases (Alcalase, Chymotrypsin, Neutrase, Papain, and Flavorase) to produce the antioxidative peptide. The degree of hydrolysis of REP by Neutrase (20.00%) was slightly lower than that of Chymotrypsin, but higher than those of other enzymes. The Neutrase hydrolysate from rice endosperm protein (NHREP) took on 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity similar to α-tocopherol. Also, the reaction condition of Neutrase hydrolysis was moderate (pH of 7.0 and temperature of 37 °C). Therefore, Neutrase was chosen to be the optimum enzyme for producing the antioxidative peptide from REP. In succession, the antioxidant activities of NHREP were further evaluated. The median effective concentration (EC50) value of NHREP for hydroxyl radical scavenging activity was 2.0 mg/mL, while its superoxide radical scavenging activity did not surpassed 50% even at 6 mg/mL. The percentage inhibition of autooxidation in linoleic acid system by NHREP was 82.09%, similar to that of α-tocopherol (86.59%) on day 5 at the same concentration. NHREP displayed 89.15% chelating effect on ferrous ion at a concentration of 1000 μg/mL, and a high correlation was also observed between the reducing power and antioxidant activity of NHREP (r2 = 0.99678). Consequently, NHREP was purified and identified, and the sequence determination by MALDI-TOF/TOF MS/MS revealed that the active constituent contained eight amino acids in its sequence (Lys-His-Asn-Arg-Gly-Asp-Glu-Phe), with the molecular mass of 1002.5217 Da. After sequence interpretation and database searching, the MS/MS spectrum was matched to glutelin protein f (460–465).

Isolation and identification of antioxidative peptides from rice endosperm protein enzymatic hydrolysate by consecutive chromatography and MALDI-TOF/TOF MS/MS

The defatted rice endosperm protein (REP) was, respectively, digested by five different protease treatments (Alcalase, Chymotrypsin, Neutrase, Papain and Flavorase), and Neutrase appears to be the most desirable for producing high quality antioxidant peptides from REP. Specially, the DPPH and hydroxyl radical scavenging activities of NHREP were higher than its superoxide radical scavenging activity, and the percentage inhibition of autooxidation of NHREP (80.09%) was similar to that of α-tocopherol (86.59%) on day 5. Furthermore, NHREP was purified consecutively, and the antioxidant peptides were identified to be Phe-Arg-Asp-Glu-His-Lys-Lys (FRDEHKK, 959.5 Da) and Lys-His-Asp-Arg-Gly-Asp-Glu-Phe (1002.5 Da) by MALDI-TOF/TOF MS/MS. Lastly, FRDEHKK was chemically synthesised. It significantly inhibited lipid peroxidation in an linoleic acid emulsion system more effectively than α-tocopherol, and enhanced the viability of t-BHP induced cytotoxicity up to 74.38% (for MRC-5) and 78.39% (for RAW264.7) at 80 μg/ml. Conclusively, it was feasible to produce natural antioxidants from REP.

Development of isohomoeoallelic lines within the wheat cv. Courtot for high molecular weight glutenin subunits: transfer of the Glu-D1 locus to chromosome 1A

Wheat quality depends on protein composition and grain protein content. High molecular weight glutenin subunits (HMW-GS) play an important role in determining the viscoelastic properties of gluten. In an attempt to improve the bread-making quality of hexaploid wheat by elaborating novel HMW-GS combinations, a fragment of wheat chromosome 1D containing the Glu-D1 locus encoding the Dx2+Dy12 subunits was translocated to the long arm of chromosome 1A using the ph1b mutation. The partially isohomoeoallelic line selected was characterized using cytogenetical and molecular approaches to assess the amount of chromatin introgressed in the translocated 1A chromosome. Triple-target genomic in situ hybridization indicated that the translocated 1A chromosome had a terminal 1D segment representing 25% of the length of the recombinant long arm. The translocation was also identified on the long arm using molecular markers, and its length was estimated with a minimum of 91 cM. Proteome analysis was performed on total endosperm proteins. Out of the 152 major spots detected, 9 spots were up-regulated and 4 spots were down-regulated. Most of these proteins were identified as alpha-, beta-, gamma-gliadins assigned to the chromosomes of homoeologous groups 1 and 6. Quantitative variations in the HMW-GS were only observed in subunit Dy12 in response to duplication of the Glu-D1 locus.

The amino-acid composition in endosperm and embryo of a barley variety and its high lysine mutant

The differences in amino-acid composition between Bomi barley and the high lysine 1508-mutant are exclusively confined to the endosperm proteins. Despite the increased embryo size of the 1508 seed the total lysine content is not appreciably affected by the changed embryo/endosperm weight ratio, and the amino-acid pattern of the embryo is unaltered when compared with Bomi barley.

Molecular characterization of mutations induced by gamma irradiation in rice

In order to analyze mutations induced by gamma irradiation in higher plants, we irradiated rice with gamma rays and screened for mutations expressing phenotypes of glutinous endosperm (wx), chlorophyll b deficiency, endosperm protein deficiency, gibberellin-related dwarfism, and shortened plastochron-in order to clarify types of mutations. Nucleotide sequence analysis showed that the most frequent mutation induced by gamma rays was deletion, particularly small deletion. Of the 24 mutations, 15 were small deletions (1-16 bp), four were large deletions (9.4-129.7 kbp), three were single-base substitutions, and two were inversions. Deletions 100 bp-8 kbp in length were not found, suggesting that gamma irradiation is unlikely to induce deletions of 100 bp to 8 kbp but is more likely to induce deletions between 1 and several ten bp or those of around 10 kbp or more. Based on the results, reverse genetics applications may be possible for gamma irradiation-induced deletions in rice by mismatch cleavage analysis used in Targeting Induced Local Lesions IN Genomes (TILLING) to detect small deletions and base substitutions or by using array comparative genomic hybridization (aCGH) to detect large deletions.

Characters of Cysteine Endopeptidases in Wheat Endosperm during Seed Germination and Subsequent Seedling Growth

The endopeptidases (EPs) in wheat endosperm during seed germination and subsequent seedling growth were characterized by gradient-polyacrylamide gel electrophoresis with gelatin copolymerized into the gel. Four cysteine EPs (EP1, EP2, EP3 and EP4) were detected in wheat endosperm during the 7 d growth after seed imbibition. The results also showed that the activities of all of these EPs increased continuously, and EP2 first appeared and had the highest proteolytic activity among the four EPs in this experimental process. The optimum pH and temperature of all four EPs were 4.0 and 40.0 degrees C. All EPs were completely inhibited by 25 micromol/L E-64 and had no good thermal stabilities, especially EP1. In addition, these EPs had different substrate specificities to albumins, globulins, gliadins and glutenins; the main storage proteins of mature wheat endosperm. Among them, EP2 had the highest proteolytic activities on globulins, gliadins and glutenins, and might be the most important and specific EP with potential to be tightly correlated with seedling development.

Sorghum protein digestibility is affected by dosage of mutant alleles in endosperm cells

AbstractProtein digestibility in grain sorghum (Sorghum bicolor (L.) Moench) is affected by the structure of endosperm protein bodies that limits access of digestive enzymes to storage protein. In this experiment, we studied the effect of endosperm dosage of high digestible alleles (hd+) on protein digestibility in sorghum hybrids carefully constituted for such a study. We also examined the effect that these alleles may have on endosperm protein body structure and agronomic performance of the hybrids. The results showed that protein digestibility, endosperm texture and protein body morphology were highly influenced by dosage of mutant alleles. Genotypes carrying two or more hd+ alleles had significantly higher protein digestibility than those with one or no mutant allele. Similarly, higher doses of these alleles resulted in opaque endosperm phenotype while the wild type alleles produced normal vitreous texture. Moreover, hybrids carrying variable proportions of mutant alleles had uniquely irregular but variable protein body structure that was distinct from spherical protein bodies found in normal genotypes. Agronomic performance was not affected by dosage of digestibility alleles.

Physical and Biochemical Properties of Airborne Flour Particles Involved in Occupational Asthma

Aerosol particles which deeply penetrate the human airways and which trigger baker's asthma manifestations are known to represent only a part of flour and of airborne particles found in bakeries. They were a major focus of this study. To this end, aerosols were produced from different wheat and rye flours, using an automatic generator designed for bronchial challenge. Particles were characterized for their size distribution, their ability to be deposited in the airways, their protein content, their histological composition and their reactivity with immunoglobulin E (IgE) present in sera from asthmatic bakers. Like dust particles collected in the bakery, the aerosols produced showed increased protein content but decreased IgE reactive protein content when compared to the corresponding bulk flours. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of these particles showed a predominance of endosperm gluten proteins. Under scanning electron microscopy, flour particles displayed various tissue fragments with entrapped large A-starch and small B- or C-starch granules, whereas aerosol particles appeared primarily as a mixture of the endosperm intracellular interstitial protein matrix and small B- or C-starch granules free or still associated. These observations showed that aerosols supposed to penetrate deeply the airways, mainly correspond to intracellular fragments of endosperm cells enriched in gluten proteins but with lower amount of allergens belonging to albumins or globulins.

Specific aspects of detection of peroxidase isozymes and their genetic control in barley

Identical specimens were separated by electrophoresis in two gels to detect and fix peroxidase isozymes. Both gels were stained by Coomassie brilliant blue for detecting proteins. One gel was previously incubated for detecting peroxidase activity. The differences in electrophoretic patterns between the gels indicate the zones of peroxidase activity. It has been shown that locus Prx 6H, controlling a low-mobility grain peroxidase (PRX 6H), is localized to barley chromosome 6. Two loci, Alb 4H and Alb 7H, controlling the biosyntheses of water-soluble proteins of barley endosperm, were localized to chromosomes 4 and 7. It has been demonstrated that barley culture is polymorphic at multiple molecular forms of peroxidase.

Structure and Orientation of Puroindolines into Wheat Galactolipid Monolayers

Puroindolines (PINs), basic and cysteine-rich proteins of wheat endosperm, are composed of two proteins, puroindoline-a (PIN-a) and puroindoline-b (PIN-b). Using a monolayer assay at the air/liquid interface, both PIN-a and PIN-b were studied in pure components and mixed with wheat galactolipids, 1,2-di-O-acyl-3-O-(beta-d-galactopyranosyl)- sn-glycerol (MGDG) and 2-di-O-acyl-3-O-(beta-d-galactopyranosyl-1,6-beta-d-galactopyranosyl)-sn-glycerol (DGDG). Following the adsorption of PINs at the air/liquid interface thanks to surface pressure increases, we concluded that PIN-a displays a more amphipathic character than PIN-b. Compression isotherms combined with ellipsometric measurements showed that the area per molecule is smaller and the protein film is more condensed for PIN-a than for PIN-b. According to the polarization modulation-infrared reflection-absorption spectroscopy data, both proteins display a highly alpha-helical structure and the alpha-helices are oriented rather parallel to the interface. By measuring the overpressure due to PIN adsorption into MGDG and DGDG monolayers, we observed that PIN-a interacts more strongly into lipid films than PIN-b. The observation by atomic force microscopy of mixed protein/lipid films showed that the nature of the lipid plays a significant role in the organization of PINs, particularly for PIN-a. The presence of galactolipids at the interface stabilizes the alpha-helical structure of PINs, but significant changes were observed concerning the orientation of the alpha-helices. They adopt a perfect parallel orientation to the interface in the MGDG monolayer, whereas the bundle of alpha-helices orients normal to the interface in the DGDG film.

Comparison of promoters in transgenic rice

Reports of the use of rice storage protein gene promoters to express transgenes in rice grain have demonstrated that rice grain can be used as a production platform for end-use quality or seed-based edible vaccines. The generation of transgenic rice with multitraits (gene stacking), which requires the use of multiple transgenes under the control of different promoters, necessitates the use of promoters from rice and other cereals, as this is highly advantageous in reducing homology-based transcriptional gene silencing. Using the green fluorescent protein gene (gfp) as a reporter gene and a transgenic rice platform, promoters of storage protein and non-storage protein genes from barley, wheat and rice were compared with regard to their spatial and temporal control of expression. Storage protein promoters from barley (549-bp B-hordein and 433-bp D-hordein) and wheat (425-bp high-molecular-weight glutenin) directed the expression of green fluorescent protein (GFP) in endosperm but not embryo; however, expression was leaky, as it was also observed in seed maternal tissues, leaf and root tissues. As expected, the rice promoters (1350-bp alpha-glutelin B-1 and 1007-bp alpha-globulin) directed the endosperm-specific expression of GFP in transgenic rice. Our results indicate that seed-specific promoters from barley and wheat, although containing endosperm and GCN4 motifs, which are important for endosperm-specific expression in rice, may not be spatially regulated in the same manner as they are in their native species. The analysis of GFP expression under the control of various promoters in rice grain indicates that promoters from other cereals can drive high levels of endosperm-specific expression in rice, but their utility for seed-specific expression may depend on their tissue specificity.

Proteome and phosphoproteome analysis of starch granule-associated proteins from normal maize and mutants affected in starch biosynthesis.

In addition to the exclusively granule-bound starch synthase GBSSI, starch granules also bind significant proportions of other starch biosynthetic enzymes, particularly starch synthases (SS) SSI and SSIIa, and starch branching enzyme (BE) BEIIb. Whether this association is a functional aspect of starch biosynthesis, or results from non-specific entrapment during amylopectin crystallization, is not known. This study utilized genetic, immunological, and proteomic approaches to investigate comprehensively the proteome and phosphoproteome of Zea mays endosperm starch granules. SSIII, BEI, BEIIa, and starch phosphorylase were identified as internal granule-associated proteins in maize endosperm, along with the previously identified proteins GBSS, SSI, SSIIa, and BEIIb. Genetic analyses revealed three instances in which granule association of one protein is affected by the absence of another biosynthetic enzyme. First, eliminating SSIIa caused reduced granule association of SSI and BEIIb, without affecting GBSS abundance. Second, eliminating SSIII caused the appearance of two distinct electrophoretic mobility forms of BEIIb, whereas only a single migration form of BEIIb was observed in wild type or any other mutant granules examined. Third, eliminating BEIIb caused significant increases in the abundance of BEI, BEIIa, SSIII, and starch phosphorylase in the granule, without affecting SSI or SSIIa. Analysis of the granule phosphoproteome with a phosphorylation-specific dye indicated that GBSS, BEIIb, and starch phosphorylase are all phosphorylated as they occur in the granule. These results suggest the possibility that starch metabolic enzymes located in granules are regulated by post-translational modification and/or protein–protein interactions.

Comparative proteomic and transcriptional profiling of a bread wheat cultivar and its derived transgenic line overexpressing a low molecular weight glutenin subunit gene in the endosperm

We carried out a parallel transcriptional and proteomic comparison of seeds from a transformed bread wheat line that overexpresses a transgenic low molecular weight glutenin subunit gene relative to the corresponding nontransformed genotype. Proteomic analyses showed that, during seed development, several classes of endosperm proteins were differentially accumulated in the transformed endosperm. As a result of the strong increase in the amount of the transgenic protein, the endogenous glutenin subunit, all subclasses of gliadins, and metabolic as well as chloroform/methanol soluble proteins were diminished in the transgenic genotype. The differential accumulation detected by proteomic analyses, both in mature and developing seeds, was paralleled by the corresponding changes in transcript levels detected by microarray experiments. Our results suggest that the most evident effect of the strong overexpression of the transgenic glutenin gene consists in a global compensatory response involving a significant decrease in the amounts of polypeptides belonging to the prolamin superfamily. It is likely that such compensation is a consequence of the diversion of amino acid reserves and translation machinery to the synthesis of the transgenic glutenin subunit.

Development and evaluation of transgenic rice seeds accumulating a type II-collagen tolerogenic peptide

Type II collagen (CII) in joint cartilage is known to be a major auto-antigen in human rheumatoid arthritis. Several animal model- and clinical-studies on tolerance-based immunotherapy for the arthritis have been conducted by administrating synthetic immunodominant peptides through an oral route. In the present study, to produce a tolerogenic peptide with therapeutic potential in transgenic rice plants, a gene construct producing glutelin fusion protein with tandem four repeats of a CII(250-270) peptide (residues 250-270) (GluA-4XCII(250-270)) containing a human T-cell epitope was introduced with a selection marker, hygromycin phosphotransferase gene (hygromycin-resistance gene) (hph), by co-transformation. Several transgenic plants with high and stable expression of gluA-4XCII ( 250-270 ), but no hph, were selected based on both DNA and protein analyses. The GluA-4XCII(250-270) fusion proteins were detected as both precursor and processed forms mainly in a glutelin fraction of rice endosperm protein extracts and in protein-body rich fractions prepared by density gradient ultracentrifugation. The amount of accumulated CII(250-270) peptide was immunochemically estimated to be about 1 microg per seed. Feeding DBA/1 mice the transgenic rice seeds (25 microg of the peptide per mouse a day) for 2 weeks showed tendencies lowering and delaying serum specific-IgG2a response against subsequent and repeated intraperitoneal-injection of type II collagen. Taken these together, the CII-immunodominant peptide could effectively be produced and accumulated as a glutelin-fusion protein in the transgenic rice seeds, which might be useful as pharmaceutical materials and functional food for prevention and therapy for anti-CII autoimmune diseases like human rheumatoid arthritis.

Influence of Cooking Conditions on the Protein Matrix of Sorghum and Maize Endosperm Flours

ABSTRACTTo understand the influence of the sorghum and maize endosperm protein matrix honeycomb structure on starch hydrolysis in flours, three‐dimensional fluorescence microscopy was applied to floury and vitreous endosperm flours cooked under various conditions. Cooking caused the collapse and matting of the sorghum and maize vitreous endosperm matrices, with the effect being greater in sorghum. The effect of cooking was rather different in the floury endosperm in that the protein matrices expanded and broke up to some extent. These effects were a consequence of expansion of the starch granules through water uptake during gelatinization. Cooking in the presence of 2‐mercaptoethanol caused an expansion of the vitreous endosperm matrix mesh due to breakage of disulfide bonds in the protein matrix. Mercaptoethanol also caused an increase in the proportion of β‐sheet structure relative to α‐helical structure of the endosperm proteins. Increased energy of cooking caused collapse of the sorghum matrix. Disulfide bonding and an increase in β‐sheet structure occurred with cooking, with the increase in disulfide bonding being greatest in sorghum vitreous endosperm. The tendency for the sorghum protein matrix to collapse and mat more with cooking than the maize matrix appears to be due to greater disulfide bonding. This is responsible for the observed low starch digestibility of cooked sorghum flour as a result of the more disulfide‐bonded protein matrix limiting the expansion of the starch granules and hence amylase access.