Solubilization Behavior And Mechanism Of Tilapia Myosin Induced By Amino Acid In Low Ionic Strength Solution

Myosin is an important component of muscle.Myosin has a long asymmetric structure with a length of about 150 nm,which forms insoluble filaments under physiological conditions.Under the low ionic strength condition in vitro,myosin are highly insoluble,which limits the processing and application of muscle.In order to improve the solubility of myosin in low salt conditions,expand the application of aquatic protein,changes of solubility,turbidity,conformation of tilapia myosin under different ionic strengths was studied.Solubilization effect of type and concentration of amino acids on myosin was determined,and changes of conformation and properties of myosin were analyzed by SDS-PAGE,tryptophan fluorescence spectrum,surface hydrophobicity,circular dichroism,transmission electron microscopy(TEM)under different ionic strength conditions.Solubilization behavior and mechanism of myosin induced by amino acid in low ionic strength solutions was determined.The purpose of this study is to provide the basis for the research and development of low salt aquatic protein food.The main results were as follows:1.Salt concentration(1-600 mmol/L KCl)and pH(2.0,7.0 and 11.0)have significantly influence on tilapia myosin structure.Under the neutral condition,the turbidity of myosin dispersion decreased,solubility increased,and the tertiary structure changes significantly with the increasing of ionic strengths.In low salt solutions,myosin molecules assembled to filament,and myosin was insoluble.In high salt solution(600mmol/L KCl),filament dissociated to monomer or soluble oligomers because of electrostatic interactions between molecules,and solubility increased.The solubility obviously improved by acid treatment(pH 2.0),but the protein molecular structure was unfolded,hydrophobic groups was exposed,surface hydrophobicity was significantly increased(p<0.05).In alkaline condition(pH 11.0),turbidity of myosin dispersion decreased,solubility increased,and changes of secondary and tertiary structure of myosin molecules were significantly.2.Glycine,L-arginine,L-lysine and L-histidine was used for solubilization experiment in low ionic strength condition(1-150 mmol/L KCl),and effect of four aminoacids on turbidity and solubility of myosin was determined.Solubilization effect of four amino acids on myosin was compared.Results showed that,the dialysis against amino acid solution resulted in changes of turbidity and solubility,and this changes was related to the ionic strength and type of amino acids.Comparatively,the dialysis by L-lysine and L-arginine solution caused significantly the decreasing turbidity and increasing solubility(p <0.05),and the myosion solution was clear,and the solubilization effect was obvious.After dialysis treatment by L-histidine solution,a certain solubilization effect was observed in the very low ionic strength(1-150 mmol/L KCl)solutions.But the dialysis by glycine caused no obvious change of turbidity and solubility of myosin(p>0.05).In low ionic strength condition containing 1 mmol/L KCl,myosin was dialyzed against Tris-HCl buffer containing 10 mmol/L L-arginine and L-lysine,and solubility of myosin was respectively78.5% and 82.5%.Therefore,L-arginine and L-lysine was used for the further test.3.The myosin concentration was fixed 2.0 mg/mL,and the solubilization behavior and mechanism of myosin was investigated induced by L-arginine.The results showed that5 mmol/L L-arginine can significantly improve solubility of myosin(p<0.05)under low ionic strength(1-150 mmol/L KCl)condition.The turbidity of myosin dispersion decreased,myosin filament dissociated,the surface potential of molecules and the surface hydrophobicity increased(p<0.05),and α-helix content was decreased.Compared with solubility of myosin adjusted to the corresponding pH values,,solubilization effect induced by L-arginine was more obvious(p<0.05).Under the high ionic strength(600 mmol/L KCl)conditon,amino acids had no significant effect on turbidity and solubility(p>0.05),while surface hydrophobicity increased,α-helix content decreased,and myosin molecule was a single molecular state.Preliminary analysis indicated that solubilazation effect of myosin induced by amino acid in low ionic strength solution might be related to the filament depolymerization and conformation changes by the increasing electrostatic and surface hydrophobicity interaction.4.Under the same conditions,the effect of L-lysine on the solubilization of myosin and its mechanism were discussed.The dialysis treatment by 5 mmol/L L-lysine solution can significantly reduce the turbidity of myosin dispersion in low ionic strength solution(1-150 mmol/L KCl)(p<0.05),the aggregation of myosin molecular was inhibited,and the solubility increased.During the solubilization processing,the myosin molecular conformation was changed,the surface hydrophobicity increased,α-helix content decreased.Compared with the myosin dispersion adjusted to the corresponding pH values,the solubilization effect induced by L-lysine was more obvious(p<0.05),molecular surfacepotential and hydrophobicity increased significantly(p<0.05),α-helix content decreased.Preliminary analysis of effect solubilization dialysis by L-lysine might be related to the filament depolymerization and conformation changes by the increasing electrostatic and surface hydrophobicity interactiont,and might be related to the specific binding between L-lysine and myosin molecules.