soybean proteins

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SOY AND WHEAT PROTEINS By TAHSEEN FATIMA MIANO

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Page 1: Soybean proteins

SOY AND WHEAT PROTEINS

ByTAHSEEN FATIMA MIANO

Page 2: Soybean proteins

Outline:

1. Soybean protein

IntroductionClassificationProperties ApplicationProcessingAlleging effects

2. Wheat ProteinIntroductionClassificationProperties Germ proteinBran ProteinEndosperm protein

3. Conclusion

Page 5: Soybean proteins

Soybean storage proteins

Globulins 85%Major globulins are 2S, 7S, 11S, and 15S molecular masses of approximately 25, 160, 350, and 600 kDa,

7S and 11S globulins, are β-conglycinin and glycinin,

multi-subunit storage proteins Salt-soluble glycinin (11S) 40% β-conglycinin (7S), 25% total seed endosperm protein.

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Factor affecting on composition of the storage proteins Maturity,

Nutrient supply from the soil,

Fertilizer treatment,

And with environmental factors.

Poor accumulation of sulfur- β-subunit of β-conglycinin is promoted by excess application of nitrogen or by sulfur deficiency,

Application of sulfur fertilization increases the synthesis of glycinin.

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Kunitz trypsin, Bowman–Birk,

protease inhibitors,

β-amylases, lipoxygenases, urease, and seed lectins,

each of which accounts for about 2–5% of the total seed proteins in soybean.

functional properties of soy proteins:

Gelation,

Gels made from glycinin being harder

Gels from β-conglycinin.

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Chemical structure of soybean proteins

Glycinin: Glycinin is a hexamer with a molecular mass of 300–380 kDa and is formed by associating six acidic and six basic polypeptides.

The acidic subunits have an isoelectric point (pI) ranging from 4.5 to 5.5.

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Glycinin’s five subunits further classified into group I (A1aB1b, A2B1a, A1bB2) and group II (A5A4B3, A3B4)on the basis of the extent of their homology. Group I subunits contain two cysteine and three cystine residues, group II subunits contain two cysteine and two cystine residues.

3D structure:Soybean storage proteins by using optical rotatory dispersion,

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CRYSTALLIZED PROTEIN

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𝛃-CONGLYCININ: STRUCTURE AND PROPERTIES Β-CONGLYCININ IS A VICILIN STORAGE PROTEIN

Molecular mass of 150-200 kDa

a glycoprotein and a trimer consisting of three subunits namely α, α , and β with molecular masses of 68, 72, and 52 kDa, ′ Subunits α and α contain one cysteine residue near the N-′terminus.

β subunit is devoid of any cysteine,

None of these three subunits contain cystine residues by strong hydrophobic interaction and hydrogen bonding.

β-Conglycinin undergoes the association–disassociation.

Ionic strength and Ph.

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Separation of soybean glycinin and -conglycinin subunits𝛃

The separation of glycinin and β-conglycinin is important for chemistry and physiological activity of each subunit.

Food industrial applications.

high-purity preparation and appropriate storage conditions of the protein

Industrial Application Of Soybean

Storage proteins requires less purity and less-expensive techniques,

High yield, simplicity, and feasibility for large-scale industrial production.

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Centrifugation at 2–5◦C; To collect insoulble material.

Supernatant pH was adjusted to pH 6.4 to precipitate glycinin.

pH adjusted to 4.8 to precipitate β-conglycinin.

Both precipitates were washed and pH adjusted to 7.8 (for glycinin extraction using NaOH)

and to 6.2 (for β-conglycinin extraction using HCl) to solubilize the purified fractions.

The remaining precipitate was removed by centrifugation and the supernatants were called glycinin-rich and β-conglycinin-rich fractions.

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SOYBEAN ENZYMES AND ENZYME INHIBITORS

1. Urease Soybean urease, EC 3.5.1.5, also known as urea amidohydrolase is a homooligomeric protein.

approximately 90-kDa subunits

It is rich in cysteine residues with 34 sulfhydryl groups per hexamer

This enzyme catalyzes the degradation of urea into carbon dioxide and ammonia,

enhancing the rate of reaction by 8 × 1017 units

Urease activity is present in all tissues of the soybean plants and has two isoforms with 87% amino acid homology.

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One of the isoforms called the ubiquitous urease

responsible for recycling all urea delivered to the embryo assimilatory function

urease called the seed urease has defense roles associated with its ureolytic activity, as a result high release of ammonia

Soybean meal and feeds, residual urease activity is used as an index for adequate heat treatment that the product has received.

soybean meals used for poultry feed, an acceptable range of 0.20–0.05 pH unit.

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LIPOXYGENASE

Lipoxygenase (LOX), E.C. 1.13.11.12, also known as linoleate: oxygen oxidoreductase, is a non-heme iron-containing dioxygenase.

catalyzes the insertion of molecular oxygen into polyunsaturated fatty acid

2% of total soybean seed protein.

functions in the plant including

vegetative growth, wounding, herbivore, and pathogen attack response and mobilization of storage lipids during germination

During germination, no substantial oxygenation of polyunsaturated fatty acids happens; suggesting that LOX is not used for lipid mobilization.

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These three LOXs are controlled by separate genes: Lox1, Lox2 and Lox3

Soybean LOX-1 is mostly α-helical that was described to becomposed of 2 domains and 5 domains

In soybean-containing foods, LOX plays a role in the development of unpleasant flavorsbecause of oxidation of polyunsaturated fatty acids;

volatile degradation products of linoleic and linolenic acids in soybean oil rancid off-flavors leading to reduce consumer acceptability of soybean products.

These off flavors have been associated with the production of volatile compounds such as N-hexanal and cis-3-hexenal.

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Bowman–Birk inhibitor

Soybean Bowman–Birk inhibitor (BBI) is composed of 71 amino acids

with a molecular mass of 8 kDa

The isoelectric point of BBI is between pH 4.0 and 4.2

BBI is rich in cysteine residues

contains seven disulfide bonds between amino acids residues 9 and 24, 14 and 22, 8 and 62, 12 and 58, 36 and 51, 32 and 39, and 41 and 49.

BBI secondary structure is composed of 61% β-sheet, 38% unordered structure, 1% β-turn and no helices,

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X-ray crystallographic study showed that BBI contains two domains; one is responsible for

its trypsin-inhibitory activity,

whereas the other domain is responsible for inhibiting.

Kunitz trypsin inhibitor

Kunitz trypsin inhibitor (KTI) is a 21.5-kDa molecule consisting of 12 crisscrossing β-sheets oriented in antiparallel fashion stabilized by

hydrophobic side chains.

It has two disulfide bonds between cysteine 39 and cysteine 86, and cysteine 136 and cysteine 145

KTI has been found to have eight distinguishable isoforms namely, Tia, Tib, Tic, ti-null, Tid, Tie, Tif, and Tibi5,

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Isolation of KTI from soybean was based on its capability to precipitate out of the solution when heated in 2.5% trichloroacetic acid with minimum solubility in water at pH 4.2– 4.8.

Combination of centrifugation, ammonium sulfate precipitation, gel filtration chromatography, and anion exchange chromatography used to isolate and purify KTI from soybean.

A simple affinity chromatography method was also used to purify KTI from soybean.

Trypsin was immobilized in an affinity chromatographic resin and an aqueous solution of defatted soybean meal was loaded into the column.

Page 24: Soybean proteins

Proteins as allergens in soybean

Soybean is considered one of the “Big 8” most allergenic foods.

The other seven are milk, eggs, fish, crustacean shellfish, tree nuts, peanuts, and wheat.

Soybean allergens consist of proteins with molecular masses ranging from 7 to 71 kDa.

Thirty-four reactive proteins have been identified and characterized as related to soybean allergy.

any allergen in a food system, soy allergen reactivity is dominated by epitopes or(Immunoglobulin E )IgE-binding sites, categorized as linear or conformational.

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•The epitopes are not only fully characterized by their primary protein structure,• but also by their tertiary structural conformations.

•These structures offer opportunities for different thermal and nonthermal food processing technologies to alter the nature of epitopes and allergen reactivity with IgE antibody.

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SOYBEAN PROCESSING

such as extrusion, may inactivate some antinutritional factors through heat treatment;

This process may also reduce availability of amino acids, especially when the soybean product is overcooked.

Extrusion is a high-temperature/short-time process in which moistened, expansive, andstarchy and/or protein rich food materials are plasticized.

and cooked by a combination of moisture, pressure, temperature and mechanical shear, resulting in molecular transformation and chemical reactions.

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SODIUM DODECYL SULFATE POLYACRYLAMIDE GEL ELECTROPHORESIS (SDS–PAGE)

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acid chain is composed of Gly1, Gly2, Gly3, Gly4, and Gly5 that corresponds to 35, 32, 38, 30, and 10 kDa,.

Proteins identified as allergenic proteins in SPI were 22-kDa KTI and 17-kDa 2S globulin. The allergenic protein LOX (102 kDa) was not found in SPI.

For the corn meal, Zea profilin, Zea m 1, γ-zein, and Zea m 14 were identified as allergenic proteins which corresponded to 60, 35, 27, 9 kDa proteins.

. Extrusion process at lower temperatures and higher moisture content produced samples that were very hard in texture .

The quick loss of moisture after expansion at higher temperature may have resulted in crispier samples

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THE WHEAT PROTEIN

1 NOMENCLATURE:

Wheat seeds comprise three main parts a protective seed shell,

a small embryo, and a

starchy endosperm.

easy solubility wheat proteins

using different molecular and proteomic.

Page 33: Soybean proteins

The classic nomenclature of wheat-seed proteins is based on their solubility

The term “Osborne fractions” covers four groups of proteins,

water-soluble albumins,

the salt-soluble globulins,

the alcohol-soluble gliadins and

the alcohol-insoluble glutenins

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Page 35: Soybean proteins

The protein composition of wheat endosperm consists about 80% of storage proteins.

Deposits of nitrogen and sulfur for germination .

Gluten proteins function as building blocks wheat gluten .

Contains two osborne fractions, the gliadins and the alcohol-insoluble glutenins.

complex classification

Which is mainly based on their Genetic composition,

Structural features and molecular function.

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prolamin groups are further divided into three distinct groups of proteins.

namely the S-rich, S-poor and HMW prolamins.

HMW glutenin subunits evolved in a different way compared to the S-rich and sulfurpoor prolamins.

While HMW glutenin subunits share similarities to rice glutelins.

Which belong to globulin type of proteins, S-rich and sulfur-poor prolamins have developed due to some duplication events.

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HMW glutenin subunits share similarities to rice glutelins.

High molecular weight glutenin subunits

The HMW-GS of wheat endosperm glutenin fraction represents only about 17% of the total gluten protein.

Significant effect on dough strength and dough stability.

The HMW glutenins are encoded by Glu-1 genes.

Each HMW glutenin locus harbors two adjacent genes, an x-type and a y-type gene.

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The polypeptides coded by the x-type genes have larger molecular weights.

Due to the presence of inner stop codons, as well as gene inactivation events, the numbers of HMW glutenin subunits that are actually expressed vary from 3 to 5.

In hexaploid wheat, the y-type subunit coded at chromosome 1A is generally absent.

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Polymorphisms have been detected at all three Glu-1 loci.

present number of identified alleles based on the Grain Genes 2.0 database is 22 for Glu-A1, 52 for Glu-B1, and 36 for Glu-D1.

The ratio of x-type to y-type subunits varies between 1.7 and 3.2,depending on the genotype characterized.

most of the over-expressing gene variants, a duplicated matrix attachment region has been identified in the distal promoter region.

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Prolamins, HMW glutenin polypeptides show unusually high levels of proline, glutamine and glycine in their sequences.

Their secondary structure reveals short N- and C-terminal domains with high cysteine content, α-helical structure and a large repetitive region with frequent repeats in the sequences.

Cysteine residues enable the formation of intra- and inter-molecular disulfide bonds which are responsible for the formation of the polymeric glutenin structure.

Repeats of the repetitive domain of x-typeand y-type sequences show special conserved patterns.

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Low molecular weight glutenin subunits

The LMW-GS are controlled by genes at the Glu-A3, Glu-B3, and Glu-D3 loci on the short arms of chromosomes 1A, 1B, and 1D.

N-terminal sequences originally distinguished by the first amino acid of their mature peptide sequences

Isoleucine (LMW i-type), serine (LMW s-type) or methionine (LMW m-type) at their N-terminal, as identified by N-terminal sequencing

Sequences starting with SHIPGL were supposed to belongto the s-type sequences.

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Deduced amino acid sequences of their encoding genes started either with IENSHIPGLEK, MENSHIPGLEK, or MENSHIPGLER,

Post-translational modification due the effect of the asparaginyl peptidase at position N.

M-type sequences were described as

mature peptides starting with MET, including types with METRCIPGLER, METSCIPGLER, METSCISGLER, METSRVPGLEK, METSHIPGLEK, METSHIPSLEK, and MDTSCIPGLER.

LMW i-type genes are considered as gene types completely lacking their N-terminal region.

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LMW-GS consist of between 250 and 300 amino acid residues.

Gliadins

Gliadins form the most diverse group of wheat prolamins.

Gliadins are usually characterized as monomeric and alcohol-soluble proteins with a molecular range between 30 and 75 kDa

Initially, based on their electrophoretic mobility on acid-PAGE analysis, they have been differentiated into four groups, α-, β-, γ-, andω-gliadins,

with α-gliadins as the fastest group and ω-gliadins as the slowest group.

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𝛄-Gliadins

γ-Gliadins share similarities in their sequence and molecular size to LMW glutenin polypeptides.

They contain eight cysteine residues, all of which are involved in the formation of intra-molecular disulfide bonds.

Their structure consists of a short (12 amino acids long) N-terminal domain, a relatively large repetitive domain with 78–161 residues, and a large C-terminal domain of over 130 residues.

The γ-gliadins differ from α-gliadins in their amino acid compositions, possessing higher amounts of aspartic acid, methionine, tryptophan, and lower amounts of tyrosine and phenylalanine.

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𝛂-Gliadins

The estimated number of genes encoding α-type gliadins ranges from 25 to 150 copies

gene amplification

retrotransposon insertion

major molecular mechanisms

large number of α-gliadin genes.

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Other gliadin loci

Genes encoding most of the ω- and γ-gliadins

tightly clustered at the Gli-A1, Gli-B1, and Gli-D1 loci on the distal ends of the short

arms of chromosomes 1A, 1B, and 1D.

A few gliadin components and LMW subunits have been shown to be encoded by additional.

short arms of chromosomes 1A, 1B, and 1D, tightly linked to one or another of the major Gli loci. Gli-5 is a minor locus tightly linked to Gli-1 and it encodes several ω-gliadin genes

Their encoding loci Glu-2, Gli-3, Gli-5, and Gli-6.

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Minor prolamins

1 Wheat -amylase and protease inhibitors 𝛂

Wheat α-amylase and serine/ trypsin protease inhibitors are salt-soluble proteins that inhibit various insect proteases and amylases.

The wheat α-amylase inhibitors (WAI) are present in three distinct forms,

in monomeric (WMAI), dimeric (WDAI), and tetrameric (WTAI) forms.

Their tertiary structure consists of four major α-helices and a one-turn helix arranged in an up-and-down manner.

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Wheat farinins and purinins

In addition to gliadin and glutenin, wheat flour contains a number of minor storage protein types.

Name avenin-like proteins or more recently farinins based on the sequence similarities to both oat avenins and γ-gliadins.

Another group of minor prolamins called purinins or LMW gliadins consists of proteins with molecular weights less than 20 kDa.

The purinin proteins all contain 14 cysteine residues, 8 of which correspond to the characteristic conserved cysteine skeleton of the prolamin super family.

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Both purinins and farinins are prolamin-like in composition but have lower contents of glutamine and proline than the γ-gliadins.

Puroindolines and grain-softness proteins

Friabilin, a protein fraction of roughly 15 kDa, has been identified from the surface of starch granules isolated from soft wheats.

Friabilin comprises more than 10 closely related components,

of which puroindoline a, puroindoline b, and the grain-softness proteins (GSPs) are the main components.

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Lipid-transfer proteins: LTPs are small, basic proteins that are characteristic of different

tissues of higher plants.

They are generally 7–15 kDa in size and are basic proteins.

prolamin super family, they contain eight cysteine residues which are able to form four intra-molecular disulfide bonds.

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Thionins

Thionins comprise a group of small ( 5 kDa) Cys-rich peptides ∼that also occur in cereal species.

They are generally 45–47 amino acids long and are classified into five subclasses (Type I–Type V).

Purothionins, or Type I thionines, first isolated from wheat endosperm, have been identified in three variants: that is, α1, α2, and β.

Their genes are controlled by loci on the long arms of the group 1 chromosomes (Pur-A1, Pur-B1, Pur-D1.

They are basic polypeptides, only 45 amino acids long, containing 8 cysteine residues

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Water-soluble proteins

The 3S and 7S globulins Several types of globulins are also present among the flour proteins.

Proteins termed globulin-1, 3S, or α-globulin are encoded at the highly conserved Glo-2 locus between the loci for the x- and y-type HMW-GS on chromosome

Globulin-2 or 7S globulins are members of the cupin superfamily, similar to known food allergens.

Analysis of wheat endosperm cDNA libraries shows that α-globulin transcripts are present and encode a mature protein.

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Triticins

The triticins of wheat endosperm are legumin-like 11S globulin storage proteins encoded at Tri-A1, Tri-B1, and Tri-D1 on the short arms of chromosomes 1A, 1B, and 1D.

They account for about 5% of the total seed protein and form small polymeric proteins, consisting of hetero-tetramers linked by disulfide bonds.

The triticin locus Tri-B1, identified on chromosome 1B, seems to be an inactive locus.

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HMW albumins

non-prolamin proteins of 60 kDa, revealed by SDSPAGE ∼fractionation of reduced extracts, are mostly enzyme related and water soluble

Certain HMW-albumin bands (65, 63, and 60 kDa) occur as both disulfide-linked oligomers and monomeric forms in their native state.

They are β-amylases controlled by chromosome arms 4DL, 4AL, and 5AL (β-Amy-1 loci). HMW albumins are absent from protein bodies.

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A limited amount of allelic variation in these bands has been served.

A fourth HMW albumin with faster mobility and MW of 45,000 ∼Da was distinct from the β-amylase group according to immunoblotting analysis.

HMW albumins tend to be present either in monomer form or they form small aggregates, stabilized through disulfide bonds.

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LMW albumins

LMW albumins share similar characteristics to HMW albumins with a lower molecular mass of 45 kDa.

They show low polymorphism on SDSPAGE.

The amounts of them depend strongly on sulfur availability during the grain

development process.

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Glutenin polymers Because of their strong relationships with the bread-making

properties

The polymeric proteins have received intense research attention.

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Proteins of non-endosperm tissues

healthy cereal products, wheat germ and wheat bran have become a more valuable food source,

especially in the development of dietary products or products for patients with special nutritional needs.

Due to the very efficient commercial milling process, the separation of endosperm from other parts, such as germ or bran.

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Germ proteins The proteomic composition of wheat germ is of nutritional

significance and helps to elucidate processes that occur during germination, especially during pre-harvest sprouting.

Protein composition of wheat germ has been characterized by using gel-based two-dimensional proteomics tools.

In total, 612 protein spots have been characterized by peptide mass fingerprint analysis.

The majority of the proteins identified were in functional categories associated with processes involved in embryonic development and activating growth,

including different transcriptional, translational processes, energy and general metabolism, transport, cell division, and signaling processes.

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Bran proteins.

Bran has an important function as a protective barrier for the grain.

The grain is a potential food for insects, fungi, and bacteria, and it is also exposed to many environmental stresses,

Proteomic analysis of wheat bran and bran tissue fractions were analyzed by spatial and temporal expression of peripheral

layer proteins .

Due to the strong bonds present between various tissue layers and the endosperm tissue in mature grain.

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Peripheral layers (inner pericarp, hyaline, testa, and aleurone layer) were identified and classified into 16 different functional categories.

metabolic activity, photosynthesis and all the metabolic pathways link to the reactive oxygen species (ROS) production and

detoxification.

Protein synthesis, protein turnover, signal transduction, membrane transport, and biosynthesis of secondary metabolites were the mediating functions of this shift.

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CONCLUSION:

It is concluded from the above study the soy and wheat protein has numerous effect and mostly they are rich in protein. Whole seed of soy and wheat of various amount of proteins and its use in daily life has also great importance because of there inhibiting properties of reducing the effect of allergy creating components. Commercial application of soy protein has been of importance due to consumption of pure protein. By analyzing their 3D structure of protein we can easily identify the components and their properties.