Structural similarities among the high molecular weight protein fractions of oilseeds.

Data on the physico-chemical properties of proteins from soybean, groundnut, sesame seed, sunflower seed, safflower seed, mustard seed, rapeseed and cotton seed are fairly extensive. An examination of the available data on high molecular weight proteins suggests that there are similarities in many of their properties. In this report the similarity in amino acid composition, size and shape, molecular weight, secondary structure, subunit composition, association-dissociation at high and low pH, stability towards denaturants, hydrolysis by enzymes and quaternary structure of the high molecular weight proteins is discussed. Based on these similarities a model has been proposed for the associationdissociation, denaturation and reassociation behaviour of the high molecular weight proteins of oilseeds.

Effect of chemical modification on the binding of gossypol by gossypin (11S protein) and congossypin (7S protein) of cottonseed.

Binding of gossypol by gossypin and congossypin and their succinylated and sulfhydryl group-blocked derivatives has been measured. The binding by gossypin and congossypin is characterized by weak interaction. Succinylation of gossypin decreases the binding affinity whereas that of congossypin increases it. Blocking of sulfhydryl groups of both the proteins does not significantly affect gossypol binding, Succinylation dissociates gossypin and causes conformational changes whereas it does not dissociate congossypin but causes conformational changes. Sulfhydryl group blocking does not dissociate gossypin or congossypin, nor does it cause any conformational changes.

Effect of alkaline pH on sunflower 11S protein.

The effect of alkaline pH on sunflower 11S protein has been monitored by the techniques of ultracentrifugation, polyacrylamide gel electrophoresis, turbidity, viscosity, ultraviolet absorption spectra and fluorescence spectra· Both ultracentrifugation and polyacrylamide gel electrophoresis show the dissociation of the protein with increase in pH. Turbidity values decrease with pH while viscosity increases· With increase in pH absorbance of the protein solution increases and there is a red shift in the absorption maximum. Fluorescence quenching and a red shift in the emission maximum are also observed. Both dissociation and denaturation of the protein occur. Analysis of turbidity, viscosity and fluorescence data suggests that apparently denaturation follows dissociation.

Association-dissociation of glycinin in urea, guanidine hydrochloride and sodium dodecyl sulphate solutions.

The effect of urea, guanidine hydrochloride and sodium dodecyl sulphate on glycinin, the high molecular weight protein fraction from soybean has been investigated by analytical ultracentrifugation. Urea and guanidine hydrochloride dissociate the protein to a '2S' protein through the intermediary 7S and 4S proteins. However, in sodium dodecyl sulphate the protein directly dissociates to a 2S protein. Analysis of the data by calculation of per cent fraction and S20,w value indicates that dissociation and denaturation of glycinin occur simultaneously in the presence of the above reagents but to different extents.

Denaturation of the high molecular weight protein fraction of mustard (Brassica juncea) and rapeseed (Brassica campestris) by urea or guanidinium hydrochloride.

Urea and guanidinium hydrochloride dissociate the 12S protein of mustard and rapeseed to 1.8 S protein and the extent of dissociation depends on the concentration of the denaturant. Mustard (Brassica juncea) protein is more readily dissociated than the rapeseed (Brassica campestris) protein. The reagents denature the protein as evidenced by increase in viscosity, appearance of difference spectra and quenching of fluorescence. Rapeseed protein is denatured more readily than the mustard protein. Analysis of visctosity, spectral and fluoresence data suggests that the first event in the denaturation reaction is the perturbation of the aromatic amino acid residues followed by their exposure to the solvent medium and unfolding of the protein molecule.

Effect of sodium dodecyl sulphate on the high molecular weight protein fraction of mustard (Brassica juncea) and rapeseed (Brassica campestris).

The effect of sodium dodecyl sulphate on mustard and rapeseed 12S protein has been monitored by the techniques of ultracentrifugation, viscosity, difference spectra and fluorescence spectrophotometry. At low concentration of sodium dodecyl sulphate (<3.47 mM) mustard protein undergoes aggregation and at higher concentrations it dissociates to 1.8 S protein, the dissociation being complete at 17.3 mM sodium dodecyl sulphate. The rapeseed protein, on the other hand, undergoes dissociation at all the concentrations of sodium dodecyl sulphate. The reduced viscosity values of mustard protein in the presence of the denaturant are higher than those of rapeseed protein. Similarly in difference spectra change in absorbance values of mustard protein are higher.' The relative fluorescence intensity of the mustard protein increases with sodium dodecyl sulphate concentration, upto 0.87 mM and this is followed by fluorescence quneching at higher denaturant concentrations. However, with the rapeseed protein fluorescence quenching was observed at all concentrations of sodium dodecyl sulphate.

Effect of denaturants on the 11S protein of guar seed.

The effect of sodium dodecyl sulphate, urea or guanidinium hydrochloride on the sedimentation velocity, viscosity, ultra violet spectra and fluorescence spectra of the 11S protein of guar seed has been determined. Sodium dodecyl sulphate dissociates the protein directly to the 2S protein, whereas urea or guanidinium hydrochloride produces an intermediate 7S protein. These reagents denature the protein also. Both the dissociative and the denaturation effect follow the order, sodium dodecyl sulphate > guanidinium hydrochloride > urea when the concentration are expressed as mols per litre. The denatured states in the three cases probably differ.

Isolation and characterization of the 7S protein from glanded cottonseed (Gossypium herbacium).

The major protein from glanded cottonseed has been isolated in a homogeneous form. Its S20, w value at 1% protein concentration is 6S in 1 Μ NaCl solution. It contains 1% carbohydrate and is free from phosphorus, gossypol (bound or free) and nucleic acid impurities. It consists of atleast seven non-identical subunits. The protein has an ultraviolet absorption maximum at 278 nm and fluorescence excitation and emission maxima at 280 nm and 325 nm respectively. Optical rotatory dispersion and circular dichroism measurements indicate that the protein consists predominantly of β-structure and random coil. The observed near-ultraviolet circular dichroic bands can be attributed to tyrosine, phenylalanine and tryptophan residues of the protein.

Extractability of polyphenols of sunflower seed in various solvents.

The extractability of chlorogenic acid from defatted sunflower seed flour in water and salt solutions at different pH values and also in aqueous organic solvents was determined. It increased with increase in pH and at pH 8 in water nearly 70% chlorogenic acid was removed in a single extraction, while NaCl did not increase the extraction, and, MgCl2 and CaCl2 increased it, especially at higher concentrations. Methanol, ethanol, isopropanol and acetone, at 20% concentration in water, caused the maximum extraction of polyphenol. These organic solvents without added water were poor solvents for the extraction of polyphenol from the flour.

Effect of denaturants on the oligomeric structure of the 11 S protein of sunflower (Helianthus annum).

The effect of sodium dodecyl sulphate, urea, guanidinium hydrochloride and heat on the oligomeric structure of the 11 S protein of sunflower has been determined. Sodium dodecyl sulphate directly dissociates the protein to 2 S subunits, whereas urea and guanidinium hydrochloride dissociate it through an intermediate 7 S protein. Heating the protein at 90°C for 20 min caused dissociation of the 11 S protein, without any precipitation.

Interaction of allylisothiocyanate with bovine serum albumin.

The interaction of allylisothiocyanate with bovine serum albumin was monitored by fluorescence titration. The interaction was weak with an apparent association constant of 2 × 102. The interaction was unaffected in the pH range of 5·0 to 8·3 and by NaCl. However, the addition of dioxane upto 4% increased the value of the association constant. N-Methyl bovine serum albumin and bovine serum albumin with sulphydryl groups blocked had the same affinity for allylisothiocyanate suggesting that amino and sulphydryl groups may not be involved in the interaction. Polyacrylamide gel electrophoresis and estimation of available lysine suggested that there were perhaps two types of groups involved in the interaction of allylisothiocyanate with bovine serum albumin.