Study On The Interactions Of Plant Polyphenols With White Croaker Myosin

Polyphenols are a group of secondary metabolites with a polyphenolic structure that are widely found in plants.During the processing of surimi products,plant polyphenols as exogenous additives can interact with myosin,the main component of surimi,to improve the gelation properties of surimi products,however,the intrinsic molecular mechanism is not clear.In this paper,thirteen plant polyphenols(five hydroxybenzoic acids,three hydroxycinnamic acids,two flavonoids and three flavonols)with different skeletal structures were selected to study the binding and thermodynamic behaviors of myosin and plant polyphenols in white grouper by fluorescence spectroscopy,circular dichroism and dynamic light scattering,in combination with computerized molecular simulation techniques.The study was carried out to investigate the conformational changes and aggregation patterns of myosin in the presence of polyphenols,to analyze the recognition sites and binding modes of myosin to plant polyphenols,and to resolve the interaction mechanism between plant polyphenols and myosin.The main findings are as follows:(1)The type of quenching,rate constants,binding constants,number of binding sites,type of force and thermodynamic parameters of the interaction between different plant polyphenols and myosin of white croaker were investigated by fluorescence spectroscopy.The results showed that thirteen polyphenolic compounds quenched the endogenous fluorescence of myosin in a static manner,and the two could form complexes spontaneously driven mainly by hydrogen bonding and van der Waals forces.The quenching and binding ability of different polyphenols on myosin were different.For the five hydroxybenzoic acids,the above-mentioned ability was ranked as follows: Gallic acid > Protocatechuic acid > pHydroxybenzoic acid > Syringic acid > Vanillic acid;for the three hydroxycinnamic acids,the ability was ranked as follows: p-coumaric acid >ferulic acid > Sinapic acid;for the two flavonoids,the ability was ranked as follows: Luteolin > Apigenin;for the three flavonols,the ability was ranked as follows: Myricetin > Quercetin > Kaempferol.In addition,the number of binding sites of thirteen polyphenols and myosin was close to one.(2)The effects of different plant polyphenols on the aggregation state of white croaker myosin were investigated by turbidity and dynamic light scattering.It was found that all thirteen polyphenols caused an increase in turbidity and particle size,leading to some degree of aggregation and weakening of the dispersion of white croaker myosin,but the effects of polyphenols with different structures on the aggregation degree of white croaker myosin were different.Using surface hydrophobicity,synchronous fluorescence spectroscopy and circular dichroism spectroscopy to investigate the effects of different plant polyphenols on the structure of white croaker myosin.The results of surface hydrophobicity and synchronous fluorescence spectroscopy showed that thirteen polyphenols could change the microenvironment of hydrophobic amino acid residues(Tryptophan,Tyrosine and Alanine)of myosin.The results of circular dichroism spectroscopy showed that thirteen polyphenols could also change the secondary structure of white croaker myosin,ferulic acid,p-coumaric acid,Sinapic acid,Protocatechuic acid,p-Hydroxybenzoic acid,Syringic acid and Vanillic acid decreased the α-helix structure of myosin and relaxed the peptide chain;Gallic acid,Luteolin,Apigenin,Kaempferol,Myricetin and Quercetin increased the α-helix structure of myosin and contracted the peptide chain.(3)Using homology modeling,molecular docking and molecular dynamics simulation technology to investigate the interactions between different plant polyphenols and white croaker myosin at the molecular level.Based on the PDB 5H53 and 5TBY templates,a heavy chain model of white croaker myosin was established,in which the dihedral angle of amino acid residues in a reasonable region was 99.3% and the ERRAT value was 98.83,which was of good quality.With the help of site prediction and molecular docking,the binding sites of thirteen polyphenols on the myosin of white croaker myosin were identified,among which,the binding sites of flavonoids were close to the BIT site,and the binding sites of hydroxycinnamic acids and hydroxybenzoic acids were located in the top region of myosin S1,which were newly discovered binding sites.Molecular dynamics simulation results showed that all four polyphenols had hydrogen-bonding interactions with white croaker myosin.In addition,π-π stacking was one of the important interactions in the complex composed of p-coumaric acid,myricetin,luteolin and white croaker myosin.