Carp Myosin Research Articles

Influence of EGCG (Epigallocatechin Gallate) on Physicochemical-Rheological Properties of Surimi Gel and Mechanism Based on Molecular Docking.

The influence of epigallocatechin gallate (EGCG) on the physicochemical-rheological properties of silver carp surimi gel was investigated. The gel strength, texture, water-holding capacity (WHC), dynamic distribution of water, and rheological properties of surimi gels added with different levels (0, 0.02, 0.04, 0.06, 0.08, and 0.1%) of EGCG were measured. The results showed that with the increase of EGCG content, the gel strength, hardness, WHC, and immobilized water contents of surimi gels showed a trend of first increasing and then decreasing, and EGCG 0.02% and EGCG 0.04% showed better gel performance as compared with the control. EGCG 0.02% had the highest gel strength (406.62 g·cm), hardness (356.67 g), WHC (64.37%), and immobilized water contents (98.958%). The gel performance decreased significantly when the amounts of EGCG were higher than 0.06%. The viscosity, G', and G″ of the rheological properties also showed the same trends. The chemical interaction of surimi gels, secondary structure of myofibrillar protein (MP), and molecular docking results of EGCG and silver carp myosin showed that EGCG mainly affected the structure and aggregation behavior of silver carp myosin through non-covalent interactions such as those of hydrogen bonds, hydrophobic interactions, and electrostatic interactions. The microstructures of EGCG 0.02% and EGCG 0.04% were compact and homogeneous, and had better gel formation ability. The lower concentrations of EGCG formed a large number of chemical interactions such as those of disulfide bonds and hydrophobic interactions inside the surimi gels by proper cross-linking with MP, and also increased the ordered β-sheet structure of MP, which facilitated the formation of the compact three-dimensional network gel.

Characteristics and stabilization of Pickering emulsions constructed using myosin from bighead carp (Aristichthys nobilis)

Myosin from bighead carp (Aristichthys nobilis) as a main type of fish protein possesses a good emulsifying ability. However, whether bighead carp myosin (BCM) could construct stable Pickering emulsions is still unclear. Therefore, myosin particles and Pickering emulsions stabilized by bighead carp myosin (BCMPEs) were analyzed. The surface structure of BCM particles at 0.6 mol/L NaCl treatment was uniform and compact with a contact angle of 86.4 ± 2.7°, exhibiting the potential ability to construct O/W Pickering emulsions. The size and flocculation index (FI) of BCMPEs decreased with the increase in BCM concentrations of 1%–4% (w/v). Reversely, the size of BCMPEs increased with the increase in oil-water ratios. BCM particles could uniformly distribute at the oil-water interface to stabilize BCMPEs at a BCM concentration of 4% (w/v) and an oil-water ratio of 6:4 (v/v). This study could help explore fish proteins to construct Pickering emulsions for the deep processing of fish products.

Structural and aggregation changes of silver carp myosin induced with alcohols: Effects of ethanol, 1,2-propanediol, and glycerol

This study investigated the effects of ethanol, 1,2-propanediol, and glycerol on the structure and aggregation behavior of silver carp (Hypophthalmichthys molitrix) myosin. All alcohols induced extensive alteration in the tertiary structure of myosin. Both ethanol and 1,2-propanediol further promoted an increase in the content of β-sheets in myosin and induced myosin aggregation. While glycerol had almost no impact on the secondary structure of myosin. Molecular dynamics simulations revealed that increasing the concentration of ethanol and 1,2-propanediol affected the overall structural changes in the myosin heavy chain (MHC), while glycerol exerted a more pronounced effect on the MHC tail when compared to the MHC head. Disruption of the hydration layers induced by ethanol and 1,2-propanediol contributed to local structural changes in myosin. Glycerol at a concentration of 20% induced the formation of a larger hydration layer around the MHC tail, which facilitated the stabilization of the protein structure.

Myosin light chain of shark fast skeletal muscle exhibits intrinsic urea-resistibility

Marine elasmobranch fish contain urea, a protein denaturant, in their bodies. The urea-trimethylamine N-oxide (TMAO) counteraction mechanism contributes to urea-resistibility, where TMAO compensates for protein denaturation by urea. However, previous studies revealed that shark fast skeletal muscle myosin exhibits native activity at physiological urea concentrations in the absence of TMAO, suggesting that shark myosin has urea-resistibility. In this study, we compared the urea-resistibility of myosin alkali light chains (A1-LC and A2-LC) from banded houndshark and carp by examining the α-helical content at various urea concentrations. The α-helical content of carp myosin A1-LC and A2-LC gradually decreased as urea concentrations increased to 2 M. In contrast, the α-helical content of banded houndshark A1-LC increased between 0 and 0.5 M urea, and the α-helical content of A2-LC remained constant until 0.5 M urea. We determined the full-length sequences of the banded houndshark myosin light chains (A1-LC, A2-LC and DTNB-LC). Hydrophilicity analysis revealed that the N-terminal region (residues 28–34) of A1-LC from banded houndshark is more hydrophilic than the corresponding region of A1-LC from carp. These findings support the notion that shark myosin exhibits urea-resistibility independent of the urea-TMAO counteraction mechanism.

Changes in Gelation Properties of Silver Carp Myosin Treated by Combination of High Intensity Ultrasound and NaCl

The molecular behavior of myosin in a low-salt environment limited the production of surimi-based products. This study aimed to investigate the effect of the combination of high intensity ultrasound (HIU) and NaCl (0.1, 0.3, 0.5 mol/L) on the physicochemical indexes of myosin. The changes were evaluated by solubility, ultraviolet (UV) spectroscopy, dynamic rheological properties, water holding capacity (WHC), microstructures, etc. For control samples, the gelation properties of myosin strengthened upon NaCl increasing. Combination of HIU and NaCl significantly improved the solubility of myosin, which was due to the conformational changes and the exposure of reactive groups. Meanwhile, the particle size of myosin obviously decreased when observed by atomic force microscope, which in turn promoted the stability of myosin. Furthermore, the improvement in solution behaviors of myosin treated by combination of HIU and NaCl contributed to the gelation properties as well as the formation of compact microstructures, which obtained high WHC and low cooking loss of myosin gels. In conclusion, combination of HIU and NaCl induced the unfolding of myosin with the exposure of reactive groups, consequently facilitating the formation of denser microstructures. Moreover, the biggest degree of improvement in gelation properties was observed at 0.1 mol/L NaCl combined with HIU.

Changes in Myosin from Silver Carp (Hypophthalmichthys molitrix) under Microwave-Assisted Water Bath Heating on a Multiscale.

To further prove the advantages of microwave-assisted water bath heating (MWH) in low-value fish processing, the effects of different heating methods (two heating stage method, high temperature section respectively using MWH1, MWH2, MWH3, WH—water heating, MH—microwave heating) on secondary and tertiary myosin structures, SDS-PAGE, surface morphology, scanning electron microscopy (SEM), and particle size distribution were compared and analyzed. The findings revealed that MH and MWH aided in the production of gel formations by promoting myosin aggregation. Myosin from silver carps demonstrated enhanced sulfhydryl group and surface hydrophobicity after MWH treatment, as well as a dense network structure. The distribution of micropores becomes more uniform when the microwave time is increased. Actually, the total effect of microwave time on myosin is not substantially different. The correlation between particle size distribution and protein aggregation was also studied, in terms of time savings, the MWH of short microwave action is preferable since it not only promotes myosin aggregation but also avoids the drawbacks of a rapid warming rate. These discoveries give a theoretical foundation for understanding silver carp myosin under microwave modification, which is critical in the food industry.

Effects of different calcium salts on the physicochemical properties of sliver carp myosin

To investigate the effects of different calcium salts on the physicochemical properties of myosin, the intermolecular interactions, particle size, surface hydrophobicity, reactive sulfhydryl group content and rheological properties of sliver carp myosin with the addition of four different kinds of calcium salts (CaCl2, calcium citrate (CC), nano-hydroxyapatite (HAP), tricalcium phosphate (TCP)) during gel processing were measured. The results showed that the addition of CaCl2 promoted the unwinding of α-helix, the increase in hydrophobic interactions, the formation of disulfide bonds of myosin, and increased the viscoelasticity of myosin. While, the addition of calcium citrate (CC) enhanced the hydrophobic interaction between myosin molecules, and the viscoelasticity of CC-added myosin was second only to the CaCl2-added myosin. On the contrary, the addition of HAP and TCP interfered with the hydrophobic interaction between myosin molecules, and greatly reduced the viscoelasticity of myosin. In summary, calcium with high calcium release rate can improve the physicochemical properties of myosin, HAP and TCP with a very low calcium release rate interfered with the formation of myosin gelation. And it could be concluded that the calcium ions played a leading role in the physicochemical properties of myosin, the more calcium ion that the calcium salt can release, the better physicochemical properties the myosin were. Our work may provide an insight into the phenomenon why different kinds of calcium salts differ from each other on affecting the physicochemical properties of myosin during gel processing.

Effect of deacetylated konjac glucomannan on heat-induced structural changes and flavor binding ability of fish myosin

This work investigated the effects of deacetylated konjac glucomannan (DKGM) on heat-induced structural changes and flavor binding in bighead carp myosin. DKGM could cross-link with fish myosin to form a thermostable complex and improve the gel strength of myosin. The incorporation of DKGM increased the surface hydrophobicity and total sulfhydryl content of heat-induced myosin. Increasing DKGM concentrations resulted in a decrease in the absolute zeta potential and a continuous increase in particle size. DKGM addition significantly reduced the α-helical content of myosin with a concomitant increase in β-sheet, β-turn, and random coil content. The binding abilities of myosin to flavors were significantly enhanced by increasing amounts of DKGM, attributing to the accelerative unfolding of myosin secondary structures and the exposure of additional hydrophobic and thiol binding sites. Increased numbers of available hydroxyl groups after DKGM treatment could also cause an increase of flavor adsorption by hydrogen bonding.

Suppression mechanism of l-arginine in the heat-induced aggregation of bighead carp (Aristichthys nobilis) myosin: The significance of ionic linkage effects and hydrogen bond effects

The suppression mechanism of l-arginine (Arg) in fish myosin heat-induced aggregation was investigated in this work. The hydrodynamic radius determined by dynamic light scattering (DLS) and the turbidity of a myosin solution decreased in a dose-dependently manner with Arg after heating, especially in a solution without pH modification (P < 0.05). The surface hydrophobicity of myosin exhibited the same trend as that for hydrodynamic radius and turbidity (P < 0.05). The myosin secondary structures clearly changed with Arg concentration during heating (P < 0.05). These results demonstrate that positively-charged Arg could form ionic interactions with negatively charged myosin, as well as promote repulsion between myosin molecules induced by higher pH values. Furthermore, the resultant hydrogen bonding observed indicated that, in addition to Arg having ionic linkage effects, the specific structure of Arg played an important role in the suppression of myosin aggregation. Essentially, Arg markedly disturbed the hydrogen bonds of the myosin backbone and formed new hydrogen bonds with myosin molecules via two amino groups (-NH2), which are part of the guanidinium moiety of Arg, preferentially hydrogen bonding with the backbone carbonyl oxygen atoms of myosin molecules. This work provides support for using Arg as an additive to modify the texture of fish products to produce liquid and semiliquid food suitable for the elderly consumers in an aging society.

Effects of pH on self-assembly of silver carp myosin at low temperature

The effects of pH on myosin self-assembly at low temperature to improve the structural attributes by controlling the solution conditions were studied. Myosin from silver carp was dissolved in solutions at various pH (5.0, 5.5, 6.0, 6.5, 7.0, 8.0 and 9.0). Effects of pH on the particle size, intermolecular forces, and micro-morphology of myosin at low temperature were investigated. The structures of myosin assemblies were observed to be influenced by varying pH using the variations in the degree of protonation and surface charge state of myosin molecules. The low electrostatic repulsion in acidic conditions resulted in the assembly before unfolding of myosin, leading to a relatively high turbidity and UV absorption as well as a decreased solubility. Confocal laser scanning microscopy analysis showed that myosin assemblies with coarse filamentous structure were formed in acid conditions. In alkaline conditions, the increased electrostatic repulsion led to a higher rate of unfolding than assembly and exposure of more hydrophobic residues, as showed by increased S0-ANS values and free SH contents. This relatively high rate of unfolding contributed to the formation of assemblies with a fine and ordered structure. At pH 7.0, the most active structure of myosin molecules was maintained as shown by the highest Ca2+-ATPase activity. Myosin assemblies with an ordered and uniform structure were obtained at pH 7.0.

Physico-chemical and functional properties of silver carp myosin glycated with konjac oligo-glucomannan: Effects of deacetylation

The glycation of silver carp myosin (Ms) with konjac oligo-glucomannan (KOG) of different degrees of deacetylation (DD) was investigated. As DD increased, the physico-chemical and functional properties of glycoconjugates changed. The available lysine content decreased, while grafting degree and total sulfhydryl group increased, andmeanwhile, the isoelectric point (pI) reduced. SDS-PAGE analysis indicated that glycation can be promoted as the increase of DD. The solubility increased significantly both in 0.1 M and 0.5 M NaCl solution, and the thermal stability increased when heated for 60 min at 80 °C. The emulsifying activity index (EAI) and emulsifying stability index (ESI) increased as well. These results showed that the highly deacetylated KOG (DKOG) was easier to glycate with myosin, leading to a great improvement in functional properties of myosin. It can be suggested that the reduction in steric hindrance of DKOG as a result of removal of acetyl groups facilitated the glycation.

Structural changes of silver carp myosin glycated with Konjac oligo-glucomannan: Effects of deacetylation

The glycation of silver carp (Hypophthalmichthys molitrix) myosin (Ms) with konjac oligo-glucomannan (KOG) of different deacetylation degrees (DDs) was investigated, in an attempt to detect the structural changes of myosin. As DD increased in KOG, glycation changed the composition of amino acids in myosin, and the surface hydrophobicity (H0) of glycoconjugates decreased. Fourier transform infrared spectroscopy (FT-IR) and circular dichroism (CD) spectroscopy exhibited the significant changes of the primary and secondary structure. The increase of denaturation temperature from DSC spectra and the changes of intrinsic fluorescence emission spectra revealed that the changes in the folded state of myosin occurred, indicating that the thermal stability or the tertiary/quaternary structural stability of myosin can be improved by the glycation with KOG of different DDs. The results from scanning electron microscope (SEM) and atomic force microscope (AFM) demonstrated that when deacetylated KOGs (DKOGs) were used as sugar donor, the glycoconjugates were larger and the surface structure is more dispersed than myosin alone. It can be suggested that myosin holds a more flexible conformation after glycation with DKOGs, which is more suitable for its various excellent functional properties.

Aggregation and conformational changes of silver carp myosin as affected by the ultrasound-calcium combination system.

Ultrasound and Ca2+ have been used separately to increase myosin properties in fish processing. However, little is know about how myosin changes are affected by Ca2+ and ultrasound in combination. The present study aimed to investigate the effects of the ultrasound-calcium combination system on aggregation and conformational changes of silver carp myosin. Ultrasound facilitated a Ca2+ -induced increase in turbidity. As the Ca2+ concentration increased from 0 to 100 mmol L-1 , there was an obvious increase in the turbidity, solubility and mean hydrated particle size of myosin after ultrasound treatment compared to without treatment. Moreover, changes of total and reactive SH contents depended on the ultrasound-calcium combination conditions. Under this combination system, myosin surface hydrophobicity significantly increased for the synergistic effect of ultrasound and Ca2+ . Furthermore, the ultrasound-calcium combination conditions could affect myosin gelation, with better gelation properties being observed for myosin treated with a combination of 60 mmol L-1 Ca2+ and 9min of ultrasound. The combination system reported in the present study was beneficial for myosin unfolding, facilitating intermolecular interactions between Ca2+ and myosin. Ultrasound treatment promoted myosin aggregation via the induction of Ca2+ and reduced the critical concentration of Ca2+ required to aggregate myosin. In the fish processing industry, this combination system can enhance the gelation properties of surimi-based products. © 2018 Society of Chemical Industry.

Effects of different NaCl concentrations on self-assembly of silver carp myosin

The effect of NaCl (0.1 − 3.0 M) on self-assembly of silver carp myosin at 4 °C was investigated in terms of microstructure, conformation, intermolecular interactions, and particle size distribution. During setting at 4 °C, the self-assembly of myosin showed an obvious concentration dependence. At low concentrations (<0.3 M), myosin assembled into filaments mainly through rod-rod ionic linkages, and showed significantly (P < 0.05) higher turbidity and lower solubility than at ≥0.3 M. The addition of NaCl (0.3 − 0.6 M) led to the extension and dissolution of myosin by interfering with electrostatic interactions. The myosin assemblies gradually became smaller and more uniform. The solubility and Ca2+-ATPase activity reached a maximum at 0.6 M NaCl. When the concentrations increased to >1.0 M, myosin further assembled into filaments dominated by hydrophobic interactions. Both rod-rod and head-head interactions contributed to the myosin filaments at high concentrations (1.0 − 3.0 M). The solubility and uniformity of the myosin assemblies decreased as the NaCl concentration increased from 1.0 to 3.0 M.

Mobility and redistribution of waters within bighead carp (Aristichthys nobilis) heat-induced myosin gels

ABSTRACTWater-holding capacity is closely related to gel microstructure, and is a very important quality trait in surimi and surimi product. The changes in the secondary structure, gel microstructure, and the migration of water in bighead carp (Aristichthys nobilis) myosin gel induced by different temperatures (50–90°C) were investigated. The α-helical structure of myosin decreased at temperatures of 40°C or higher. The fractal dimension of the gels increased at 40, 50, and 60°C, but decreased at temperatures over 60°C. The pore size of the gels increased with temperatures up to 50°C, decreased at 60°C, and then increased with temperatures up to 90°C again. The transverse relaxation times also varied; T21 remained constant at temperatures over 40°C; T22 decreased at temperatures lower than 50°C, increased at 60°C, and then decreased with temperatures up to 90°C; and T23 increased at temperatures lower than 50°C and then remained constant until 90°C. Principal component analysis showed that the proportion of T22 water (PT22) was inversely correlated with the unfolding of myosin, whereas directly correlated with the pore size. The proportion of T23 water (PT23) was positively correlated with the fractal dimensions of the gels, whereas negatively correlated with the pore size. The migration of the secondary layer of water was mainly caused by hydrophobic force and the physical space formed by the myosin backbone, and the migration of water within the third layer was mainly caused by capillary pressure. Therefore, the mobility and redistribution of waters depend on the water retention mechanism, which is determined by the physical structure of gels. This study provides further information about the relationship between the NMR data, gel microstructure, water mobility, and distribution.

Effect of l-histidine on the heat-induced aggregation of bighead carp (Aristichthys nobilis) myosin in low/high ionic strength solution

In this study, the effects of l-histidine (L-His) on the aggregation of bighead carp (Aristichthys nobilis) myosin induced by heating in low (0.1) or high (0.5) ionic strength solutions were studied. The solubility of natural myosin was significantly enhanced by L-His both in low and high ionic strength solutions. More small particles are suspended in the heated solution containing L-His, inducing a higher solubility than that solution without L-His. The surface hydrophobicity of myosin in a low ionic strength solution containing L-His is higher than that of myosin without L-His. The results of morphology show that L-His induced myosin to form finer aggregates and a honeycomb-like network in a low/high ionic strength solution during heat treatment. These results demonstrate that L-His might disrupt the electrostatic property of unfolded myosin heat-induced through electrostatic binding with the exposed negatively charged amino acid residues, which might disarrange hydrophobic surfaces, resulting in weakening hydrophobic interactions, which ultimately suppresses the fierce aggregation of carp myosin.

Physicochemical, conformational and functional properties of silver carp myosin glycated with konjac oligo-glucomannan: Implications for structure-function relationships

Silver carp myosin (Ms) was glycated with konjac oligo-glucomannan (KOG) under dry reaction conditions. The physicochemical properties (amino acid composition, total sulfhydryl groups, hydrophobicity, viscosity and microstructure), conformational properties (heat changes, secondary and tertiary conformations) and functional properties (emulsifying properties) were investigated. The amino acid composition (especially relative acidic/basic charged amino acid ratio) was an intrinsic physicochemical parameter that largely determined other physicochemical and conformational properties. Formation of Ms-KOG conjugates reduced the total sulfhydryl groups content and hydrophobicity. Glycation for 12 h increased the viscosity of Ms by 35-fold. Nanoscale structure and secondary conformation, as determined by atomic force microscopy and far-UV circular dichroism spectroscopy, remarkably changed. Combined analysis of differential scanning calorimetry and intrinsic emission fluorescence indicated that Ms may undergo a tertiary conformation unfolding and subsequent rearrangement process. More importantly, we have interestingly found that KOG improved the flexibility in conformation and enhanced the steric hindrance of Ms. This specific structure of Ms-KOG eventually improved the emulsifying properties.

Effect of Mild Ozone Oxidation on Structural Changes of Silver Carp (Hypophthalmichthys molitrix) Myosin

To produce high-quality surimi, we investigated the effect of ozone-induced mild oxidation on structural changes of silver carp myosin by analyzing circular dichroism (CD), intrinsic fluorescence, the sulfhydryl group, surface hydrophobicity, SDS-PAGE, and dynamic light scattering (DLS) of myosin. Mild ozone oxidation of myosin was performed by applying gaseous ozone for a short time. Results showed that the content of sulfhydryl group decreased, and carbonyl content and surface hydrophobicity increased with an increase in ozone treatment time. A red shift in the maximum wavelength of intrinsic fluorescence indicated the unfolding of oxidized myosin. Further conformational change of myosin was illustrated by CD; α-helical content decreased significantly and β-turn, β-sheet, and random coil content increased with increased ozone treatment time, which demonstrated the rearrangement of β-structure in myosin. The results from DLS and SDS-PAGE indicated that soluble oligomers or aggregates were formed by disulfide bonds under mild ozone treatment. Therefore, ozone-induced mild oxidation did not strongly alter myosin structure but promoted myosin partial unfolding, which might result in exposing more cross-linking sites for the formation of surimi gel.

Purification and characterisation of a salt-stable protease from the halophilic archaeon Halogranum rubrum.

Because proteases play an important role in the fermentation of fish sauce, the purification and characterisation of an extracellular protease from the halophilic archaeon Halogranum rubrum was investigated. The molecular mass of the protease was estimated to be approximately 47 kDa based on sodium dodecyl sulfate-polyacrylamide gel electropheresis (SDS-PAGE) and native-PAGE analysis. The optimum conditions for catalytic activity were pH 8.0 and 50°C. The protease showed alkaline stability (pH 7.0-10.0). The protease also exhibited novel catalytic ability over a broad range of salinity (NaCl 0-3 mol L-1 ). Calcium ion enhanced the proteolytic activity of the enzyme. The Km and Vmax values of the purified protease for casein were calculated to be 4.89 mg mL-1 and 1111.11 U mL-1 , respectively. The protease was strongly inhibited by ethylenediamine tetraacetic acid (EDTA) and phenylmethanesulfonyl fluoride (PMSF). Meanwhile, the protease was stable in the presence of Triton X-100, isopropanol, ethanol or dithio-bis-nitrobenzoic (DTNB), but was inhibited by sodium dodecyl sulfate (SDS), dimethyl sulfoxide (DMSO) or methanol. MALDI -TOF/TOF MS analysis revealed that the protease shared some functional traits with protease produced by Halogranum salarium. Furthermore, it exhibited high hydrolytic activity on silver carp myosin protein. The protease is an alkaline and salt-tolerant enzyme that hydrolyses silver carp myosin with high efficiency. These excellent characteristics make this protease an attractive candidate for industrial use in low-salt fish sauce fermentation. © 2016 Society of Chemical Industry.

Characterization of silver carp (Hypophthalmichthys molitrix) myosin protein glycated with konjac oligo-glucomannan

Silver carp (Hypophthalmichthys molitrix) myosin protein (Ms) was glycated with konjac oligo-glucomannan (KOG) by using the controlled Maillard reaction (MR). Colorimeter measurements and SDS-PAGE analysis indicated the occurrence of the MR and glycoconjugates formation, respectively. Thinner sheet and non-homogeneous Ms-KOG conjugates were observed by scanning electron microscope (SEM). Fourier transform infrared spectroscopy (FT-IR) indicated that the amide I, II and III bands of Ms were changed by the glycation. Glycation lowered the isoelectric point from 5.3 to 3.8 when incubated for 96 h. Ms became highly soluble in 0.1 M NaCl with the decrease of 23.6% of the total lysine residues at the reaction time of 12 h. The thermal stability of Ms glycated with KOG for 24 h was effectively improved from 61.4% to 95.8% when heated at 80 °C for 60 min. These results suggested that MR with KOG can be a promising way to improve functional properties of Ms.