Worldwide, plant protein contributes substantially as a food resource because it contains essential amino acids for meeting human physiological requirements. However, many versatile plant proteins are used as medicinal agents as they are produced by using molecular tools of biotechnology. Proteins can be obtained from plants, animals and microorganism cells. The abundant economical proteins can be obtained from plant seeds. These natural proteins are obtained by isolation procedures depending on the physicochemical properties of proteins. Isolation and purification of single protein from cells containing mixtures of unrelated proteins is achievable due to the physical and chemical attributes of proteins. The following characteristics are unique to each protein: Amino acid composition, sequence, subunit structures, size, shape, net charge, isoelectric point, solubility, heat stability and hydrophobicity.

Based on these properties, various methods of isolation exist, like salting out and isoionic precipitation. Purification of proteins is quiet challenging and, therefore, several approaches like sodium dodecyl sulfate gel electrophoresis and chromatography are available. Characterization of proteins can be performed by mass spectrometry/liquid chromatography-mass spectrometry (LC-MS). The amino acid sequence of a protein can be detected by using tandem mass spectrometry. In this article, a review has been made on the sources, isolation, purification and characterization of natural proteins. INTRODUCTION For carrying out different body functions, the body regularly needs nutrients like vitamins, minerals, proteins, fiber and carbohydrates, which are obtained from plant or animal sources or both.

We provide excellent essay writing service 24/7. Enjoy proficient essay writing and custom writing services provided by professional academic writers. Download Proteins: Structures and Molecular Properties, ebook Proteins: Structures and Molecular Properties, pdf Proteins: Structures and Molecular. Book, Book eBook English, Proteins: Structures and Molecular Properties Download, Download Proteins: Structures and Molecular Properties audio book for free, Proteins:.

Among nutrients, the human body requires proteins as the most important compounds because they aid in building cells and tissues and help in repairing the tissues in the body. A high protein diet is recommended for those thinking of building body or muscles. If the body lacks in carbohydrates and fats, the body makes use of proteins for energy production as they are essential for building muscle mass.[] The term “complete protein” refers to foods that contain all nine essential amino acids in the correct proportion to build protein in the body.

In contrast, “incomplete protein” refers to foods that have all essential amino acids but not in the correct proportion, and are termed as “limiting amino acid.”[] Thus, proteins are not only important in the human body but are also widely used in the industry. Hence, an attempt is made to review naturally obtained proteins and its application in pharmaceutical industries. Building blocks of proteins (Amino acids) Naturally occurring organic compounds containing amino and carboxyl groups, which are the chief constituents of protein and are necessary for human and animal growth and nutrition, are termed as essential amino acids. Hence, essential amino acid-rich food consumption is an option to source these as they are not produced by the human body.[] Proteins (or polypeptides) are amino acids joined together by peptide bonds.[] The roles of the various amino acids are highlighted in.[]. Extinction coefficient It indicates at the amount of light absorbed by a protein at a certain wavelength. This coefficient value helps in estimating and identifying a protein when exposed to a spectrophotometer.[] The molar extinction coefficient of a protein can be estimated by knowing its amino acid composition.

By using the following equation, the native protein extinction coefficient in water can be computed using the molar extinction coefficient values at a given wavelength of tyrosine, tryptophan and cystine (at 280 nm, the extinction value of Tyr is 1490, of Trp is 5500 and of Cys is 125 in water).[] E1 = no. Of (Tyr) * Ext (Tyr) + no. Of (Trp) * Ext (Trp) no. Of (Cystine) * Ext (Cystine) E2 = no. Of (Tyr) * Ext (Tyr) + no. Of (Trp) * Ext (Trp) Two values of the proteins produced in water at 280 nm using the above equation indicate that the first value (E1) is due to cysteine residues appearing as half cystines and that the second value (E2) is due to no cysteine appearing as half cystine.

Mass spectrometry The protein sequence can be directly determined by this technique using electro-spray ionization. A protein of any size can be sequenced by this method, but difficulty arises as the protein size increases. Liquid samples for mass spectrometry can be easily prepared due to greater solubility of peptides as compared with whole proteins. In solution, the proteins are digested by an endoprotease and passed through a high-pressure liquid chromatography column.

At the end of this column, the solution is sprayed out of a narrow nozzle charged to a high positive potential into a mass spectrometer. The charge on the droplets causes them to fragment until only single ions remain. Peptides are then fragmented and the mass-to-charge ratio of this is measured.

This process is repeated with a different digestion enzyme, and overlaps in sequences are used to construct a sequence for protein.[]. Soybean proteins Soy proteins are a mixture of globular proteins - conglycinin (140-170 kDa with glycosylated three subunits) and Glycinin (340-375 kDa with six AB subunits comprised of an acidic [A] and a basic [B] polypeptide linked via disulfide bonds) and are obtained from the plant species Glycine max, family Fabaceae.[] Based on the molecular weight and sedimentation coefficient, it separates into fractions 2S, 7S, 11S or 15S.[] The 7S globulin and 11S globulin comprise 37% and 31% of the soy proteins, respectively. In combination with other film-forming proteins, glycinin is known as the gelling agent, emulsifier and foaming agent.[] B-conglycinin is less heat stable than glycinin and gets denaturated at temperatures of 70°C and 80°C.[]. Wheat proteins Based on solubility, the wheat protein fractions are classified as albumins (water soluble), globulins (dilute salt solutions soluble), gliadins (soluble in 70-90% ethanol, comprise 34% of the total protein) and glutenin (insoluble under all of the previously mentioned conditions, comprise 47% of the total protein).[,] Gliadin (40 kDa) is a single-chained peptide of four distinct fractions containing intramolecular disulfide bonds.[] These play a role in film formation, strength and elasticity.

Glutenin, a mixture of proteins, has a molecular weight distribution between 100 and 1000 kDa. The disulfide bonds present in glutenin and gliadin help in determining the strength of the protein matrix.[]. Sunflower Proteins Proteins are majority constituents in sunflower oil cakes. Defatted sunflower flour contains a high quantity of proteins, around 27% in dry weight.[] The dehulled seed consists of about 20-40% crude protein. Four fractions of protein are present in the sunflower protein:[] Globulins, 55-60%; albumins, 17-23% of total proteins; and two minor fractions, glutelins and prolamins, comprising 11-17% and 1-4% of the total protein fractions, respectively. It shows two major fractions: 11S globulins (also named helianthinin) and 2S albumins.

Helianthinin has been reported to be present as a globular oligomeric protein with a molecular weight of 300-350 kDa,[] and this protein mainly exists in the 11S hexametric form. Casein There are four main subunits: s1 α casein (23.6 kDa, 4.94 a pI, net charge - 21.9 at a pH of 6.6), s2 α casein (net charge − 13.8, 5.37 pI, hydrophilic due to high-charge density), β-casein (polar N-terminal amphipathic protein with large hydrophobic domain, Ca 2 + sensitive, at 4°C solubility increases) and κ-casein (not Ca 2 + sensitive), which make up 38%, 10%, 36% and 13% of the casein composition, respectively, and has a unique property to form films.[,,] s1 α casein is amphipathic due to the charge between the hydrophobic N- and C-terminals. It has 8 phosphorylated serine clustered with glutamine residues possessing calcium-binding sites and hence is Ca 2 + sensitive, 17 proline, 25 glutamine residues and no cysteine residues.

Here, the Ca 2 + sensitivity means aggregation and precipitation in low ion concentrations. It does not participate in disulfide bond formation and cross-linking due to the absence of free cysteine.

Caseins are heat stable because they are proline-rich, which interrupt alfa-helix and beta strands, resulting in the absence of disulfide bridges in the structure. Myiptv Hack. It has relatively little secondary or tertiary structure. These undergo proteolytic cleavage due their open structure imparted due by the high proline content. This characteristic, along with acid-soluble calcium–phosphate bridging, makes an excellent target-activated release mechanism for unloading drug in the stomach.[,]. Meat proteins Sarcoplasmic, stromal and myofibrillar are types of meat protein. Sarcoplasmic proteins contain enzymes myoglobulin and cytoplasmic. Collagen and elastin are the content of stromal proteins while myosin, actin, tropomysin and troponins are the content of myofibrillar proteins.

Stromal and myofibrillar proteins, soluble in salt solutions, are used for making edible films and coatings. Collagen, a fibrous stromal protein extracted from connective tissue, tendons, skin, bones and the vascular system, and is a waste products of meat processing. Collagen is a superhelical structure formed by a combination of three parallel alfa-chains, and forms gelatine.[] Collagen exposed to mild heat treatment under acidic or alkaline conditions forms gelatin.[]. Salting out Proteins are salted out as co-precipitate by ammonium sulfate because the saturation concentration provides high molarity that causes precipitation of most proteins. It does not have a large heat of solution and hence the generated heat get easily dissipated; a saturated solution (4.04 M at 20°C) of proteins has a density of 1.235 g/cm 3, which does not interfere with the precipitated protein sedimentation by centrifugation. Its concentrated solutions are generally bacteriostatic and protect most proteins from denaturation in solution state. Illustrates the procedures for a pilot experiment.

Isoionic precipitation Column method Proteins are frequently least soluble and most precipitable when they are isoionic. In an isoionic, salt-free state, the protein molecules are in their most compact, least-hydrated conformation – a phenomenon that is closely related to the condition of proteins at their isoelectric point. The distinction between isoionic and isoelectric properties is determined by the procedure described by Tanford.[] Deionization using a column or dialysis aims at rendering proteins isoionic to precipitate them. Two important parameters determine the solubility of many proteins: Solution pH with respect to each proteins’ isoionic point (pI) and low salt concentration (0 to 0.1 to 0.2 M salt). The column method used is appropriate only for proteins that remain soluble at their isoionic point.

In addition to adjusting the proteins to their isoionic pH, the general method is using mixed-bed resin deionization to strip away all salts from proteins. Инструкция Для Ответственного За Лифты. Salts, even in small concentration, often have large effects on protein solubility and, therefore, on perceptibility.

Inorganic salts tend to salt in many proteins thus enhancing their solubility. Dialysis method One of the older methods of rendering proteins salt free or nearly salt free (i.e., isoionic) is dialysis. However, two problems frequently arise with conventional dialysis: (1) When appreciable amounts of protein are present, osmotic effects result in swelling of the dialysis bags as the salt diffuses outward and (2) often, it is quite uncertain where the isoionic point is even if it is feasible to deionize by dialysis against a buffer. The resin deionization method, sometimes called the Dintzis method,[] automatically adjusts a protein precisely to its isoionic pH without prior knowledge of the same.

Salt ions and protein counterions exchange through the membrane and are trapped outside in the exchanger resins. After exchange is complected, the precipitated protein in the dialysis tubing is recovered by centrifugation of the bag's contents. This general technique requires several hours. Salting out and diffusion become slow as the protein concentraction inside the dialysis bag decreases; hence, this method is slower than the flow-through column method. When proteins are insoluble and precipitate at their isoionic point, deionization is accomplished by placing the dialysis tubing containing the protein sample in a slurry of mixed-bed resin exchangers and incubating with rocking.

Proteins insoluble at their isoionic point precipitate inside the bag. The mixed-bed exchange resins remove free salts, forcing the protein to its isoionic pH. Coomassie blue staining This staining requires an acidic medium for generation of an electrostatic attraction between the dye molecules and the amino groups of proteins. Ionic attraction, together with van der Waals forces, binds the dye–protein complex together.

Coomassie Blue stains exhibit three-times staining intensity of Fast Green and six-times intensity of Amido Black. Usually, 0.2-0.5 μg of any protein in a sharp band can be detected using Coomassie Blue staining. Because Coomassie Blue is predominantly non-polar, it is usually used in methanolic solution and excess dye is removed from the gel later by distaining. Stacking gels are usually discarded before staining the resolving gels. Gel slabs are placed in staining solution and are stained fully within 4-6 h at room temperature if only 1.5-mm thick.

Silver staining Silver staining is 50-100-times more sensitive than Coomassie Blue staining. The principal reactive groups are free amines and sulfur groups (basic and sulfur-containing amino acids) contained on proteins. Most proteins stain with monochromatic brown or black colors. Lipoproteins tend to stain blue while some glycoproteins appear yellow, brown or red.

Artefact bands with molecular weights ranging from 50 kDa to 68 kDa have been commonly observed in silver-stained gels. Evidence has been presented indicating that these contaminating bands are due to keratin skin proteins.

Liquid chromatography–mass spectrometry and tandem mass spectrometry of peptides and proteins Protein and peptide tandem mass spectrometry involves application of two separate stages: LC–MS and LC–MS/MS, to solve problems that require protein identification and software tools to identify proteins by matching mass spectrometric data to protein sequence databases.[,] Additionally, LC–MS provides high-quality data for peptide mass as compared with MALDI mass spectrometry in fingerprinting searches. Tessier et al. Reported the prediction of plant protein hydrolysate containing small peptide amino acids’ composition performed by LC-MS and capillary electrophoresis-mass spectrometry.[].