Alpha-Helical Structure of Fish Actomyosin: Changes during Setting

Participation of the α-helix in setting was investigated using circular dichroism. The α-helicity of the actomyosin from eight species of fish decreased during incubation at 30°C or at 40°C. The extent and pattern of decrease differed among species. When rate of decrease was plotted vs rate of increase in strength of gel preincubated at 30°C or at 40°C, the two factors correlated closely. We propose that the unfolding of α-helix initiated setting.     

Protein Concentration, pH and Ionic Strength Affect Apparent Viscosity of Actomyosin

Response surface methodology was used to compare effects of NaCl protein concentration, and pH on apparent viscosity (ζapp) of natural actomyosin from hake, pork and chicken muscle. Models for the three species had R² of 0.921, 0.915 and 0.814 (P<0.01) for chicken, pork and hake, respectively. Protein concentration and pH increased qapp, whereas higher ionic strength decreased it and had less influence. Interactions occurred between protein concentration and pH. Models did not reveal inter-species differences (P<0.05). However, analysis of variance of regression coefficients for each variable showed some differences due to pH.     

Polymerization of Beef Actomyosin Induced by Transglutaminase

Polymerization of beef actomyosin was induced by addition of transglutaminase. The relative intensity analyzed by densitometry after sodium dodecyl sulfate polyacrylamide gel electrophoresis indicated that bands containing the polymerized myosin increased from 10.1 ± 2.2% to 20.7 ± 3.5% while the myosin monomer band decreased from 20.9 ± 3.4% to 13.0 ± 2.7% as the reaction time extended from 10 to 120 min at 35°C. Polymerization of actomyosin induced by transglutaminase resulted in gelation of the actomyosin that was visualized by confocal laser scanning microscopy.     

Effect of microbial transglutaminase on autolysis and gelation of lizardfish surimi

In the absence of microbial transglutaminase (MTGase), the textural properties of lizardfish surimi (Saurida spp) improved when pre‐incubated at 4 and 25 °C for 24 and 4 h, respectively. MTGase optimally catalyzed incorporation of monodansylcadaverine (MDC) into surimi at 40 °C. Addition of MTGase appeared to reduce autolytic activity at 25 and 40 °C, but had no effect on autolytic activity at 65 °C. Breaking force and deformation of lizardfish surimi significantly improved when 0.1 unit MTGase g^?1 surimi (1.8 g kg^?1) was added and pre‐incubated at either 25 or 40 °C. Textural properties improved concomitant with cross‐linked polymers of myosin heavy chain and tropomyosin, but not actin. Addition of MTGase also improved the storage modulus (G′). The gel network of surimi mixed with MTGase and pre‐incubated at 40 °C readily formed during the pre‐incubation period, while formation of the gel network began at 48.1 °C in the absence of MTGase. Copyright © 2005 Society of Chemical Industry     

Transglutaminase Effects on Low Temperature Gelation of Fish Protein Sols

Myosin polymerization and formation of ?-(γ-glutamyl)lysine linkages were quantified in Alaska pollock surimi gels which contained no additive (control), or a commercial microbial transglutaminase (MTGase). As preincubation (“setting”) time at 25°C was increased, the gel strength of control and 0.2% MTGase-added samples increased, with greater increases at higher MTGase levels. SDS-PAGE and HPLC analyses showed increasing nondisulfide polymerization and ?-(γ-glutamyl)lysine dipeptide content, with increasing setting time and/or added MTGase. Content of ?-(γ-glutamyl)lysine dipeptide correlated with gel strength (shear stress) and shear modulus at failure (G_f) for these gels. Higher stresses were measured in samples containing 0.2% MTGase than in controls at corresponding levels of ?-(γ-glutamyl)lysine dipeptide, indicating that rate of myosin polymerization may affect ultimate gel strength.     

Comparative study on protein cross-linking and gel enhancing effect of microbial transglutaminase on surimi from different fish

BACKGROUND: Microbial transglutaminase (MTGase) has been used to increase the gel strength of surimi. Nevertheless, its effectiveness varies with fish species. The aim of this study was to elucidate the effect of MTGase at different levels on protein cross‐linking and gel property of surimi from threadfin bream, Indian mackerel and sardine in the presence and absence of endogenous transglutaminase. RESULT: Breaking force of all surimi gels increased as MTGase levels (0–0.6 U g^?1) increased except for threadfin bream surimi gel, where the breaking force decreased at 0.6 U g^?1 (P < 0.05). In the presence of EDTA, the gel strengthening effect was lower, suggesting the combined effect of endogenous transglutaminase with MTGase. With the addition of MTGase, the gel with the highest increase in breaking force showed highest decrease in myosin heavy chain. When cross‐linking activity of MTGase on natural actomyosin (NAM) was determined, the highest decreasing rate in ε‐amino group content with the concomitant increased formation of cross‐linked proteins was found in NAM from threadfin bream. The reactivity of muscle proteins toward MTGase‐induced cross‐linking was in agreement with surimi gel strengthening. CONCLUSION: The composition and properties of muscle proteins of varying fish species more likely determined protein cross‐linking induced by MTGase, thereby affecting their gel properties. Copyright © 2011 Society of Chemical Industry     

Gel‐enhancing effect and protein cross‐ linking ability of tilapia sarcoplasmic proteins

BACKGROUND: Tilapia (Oreochromis niloticus) sarcoplasmic proteins contain substantial transglutaminase (TGase) activity. The enzyme catalyzes the protein cross‐linking reaction, resulting in a more elastic gel. The objective was to investigate the gel‐enhancing effect of sarcoplasmic proteins from tilapia as related to TGase activity. RESULTS: Total TGase activity of sarcoplasmic proteins concentrate (SpC) increased about 3.6‐fold after ultrafiltration using 30 kDa membrane, but specific activity remained unchanged, indicating minimal TGase purification by ultrafiltration. Addition of 1 mg mL^?1 SpC containing 40 units TGase activity induced cross‐linking of tilapia actomyosin, and the extent of cross‐linking increased with added level of SpC. Myosin heavy chain (MHC) and troponin were preferably cross‐linked by tilapia SpC, while actin and tropomyosin were not affected. Higher retention of MHC was observed concomitantly with greater content of cross‐linked protein when SpC was added to lizardfish surimi. Lizardfish surimi with 10 g kg^?1 SpC added and pre‐incubated at 37 °C for 1 h exhibited 91.6% and 26.7% increase in breaking force and deformation, respectively, when compared to the control. CONCLUSIONS: Residual TGase activity in SpC played an important role in catalyzing the protein cross‐linking and enhancing actomyosin gelation. SpC could be a potential ingredient for improving textural properties of fish protein gel. Copyright © 2007 Society of Chemical Industry     

Differences in Gelation Characteristics of Natural Actomyosin from Two Species of Bigeye Snapper, Priacanthus tayenus and Priacanthus macracanthus

ABSTRACT: Natural actomyosin (NAM) of P. tayenus exhibited higher turbidity and storage modulus (G') upon heating, compared to that of P. macracanthus , suggesting the higher protein aggregation and rigidity. At temperature above 35 °C, P. tayenus NAM had higher surface hydrophobicity and disulfide bond formation than P. macracanthus NAM. The α-helix content of NAM from both fish species decreased as the temperature increased, indicating changes in structural conformation during heating. NAM gel from P. tayenus rendered more three-dimensional network than that from P. macracanthus. These results indicated that P. tayenus NAM possessed superior gelling characteristic to P. macracanthus NAM due to the higher aggregation of protein caused by both hydrophobic interaction and disulfide bond.     

Influence of Thermal Treatment on Gelation of Actomyosin from Different Myosystems

We analyzed the influence of temperature and heating time on rheological properties, and types of proteins involved in the gelation process of natural actomyosin from pork, chicken, and hake. At low temperatures (40–50°C) hake gels were stiffer than chicken or pork gels (due to setting); at higher temperatures (60°C), actomyosin from all three formed gels with similar rheological characteristics. Types of protein in the different fractions (analyzed by electrophoresis) were consonant with the rheological behavior of the AM gels for each heat treatment and species.     

Covalent Cross-linking Effects on Thermo-Rheological Profiles of Fish Protein Gels

The extent of covalent cross-linking in surimi sols by transglutaminase was modified by addition of either EDTA (inhibition by calcium sequestering), variation of preincubation time at 25°C to allow action of endogenous transglutaminase, or by addition of a microbial transglutaminase. Gels were cooked following preincubation to fix the effects of the cross-linking reaction. Torsion, dynamic (small strain) test and stress relaxation measurements were made on the gels over a range of temperatures. Gels preincubated with EDTA had similar rheological response to temperature as gels cooked without preincubation. The stress relaxation and thermo-rheological profiles of gels with increased covalent cross-linking (induced by added or endogenous transglutaminase) were increasingly rubber elastic in their behavior.     

肌原纤维蛋白转谷氨酰胺酶交联程度对鱼糜凝胶及其风味释放影响的研究进展

转谷氨酰胺酶可以催化肌原纤维蛋白发生交联反应。随着交联程度的增加,鱼糜凝胶从弹黏体变为弹脆体,风味也随之改变。风味物质的扩散、释放与凝胶网络的交联程度密切相关,因此深入了解交联程度与食品品质的关系及风味物质在凝胶网络中的扩散行为显得尤为重要。本文总结转谷氨酰胺酶催化肌原纤维蛋白的交联机理,归纳了国内外对肌原纤维蛋白交联程郭彩霞度的测定方法与影响因素,探讨交联程度对鱼糜蛋白的凝胶特性与风味释放的影响,并对今后的研究方向提出思考与展望。     

Application of Antifreeze Protein for Food Preservation: Effect of Type III Antifreeze Protein for Preservation of Gel-forming of Frozen and Chilled Actomyosin

Type III antifreeze protein (AFP) remarkably preserved Ca²⁺ ATPase activity of actomyosin during frozen and chilled storage. Under frozen conditions, AFP helped to retain the Ca²⁺ ATPase activity of actomyosin much higher than that of conventional cryoprotectants (sucrose‐sorbitol mixture). The Ca²⁺ ATPase activity increased with increasing AFP concentration and reached a maximum at 50g/L AFP. After 3‐d chilled storage, the Ca²⁺ ATPase activity of control and sucrose‐sorbitol samples had lost 80% and 50%, respectively, while the AFP samples remained unchanged. A Type III AFP mechanism based on freezing temperature depression (more unfrozen water) and inhibition of ice recrystallization that protects against the freezing of muscle proteins in chilled or frozen conditions is proposed.     

Chemically Induced Covalent Crosslinks Affect Thermo-Rheological Profiles of Fish Protein Gels

Effects of inducing covalent crosslinks in surimi gels by adding potassium bromate or 1-ethyl-3(3-dimethylamino propyl) carbodiimide (EDC) were investigated. Potassium bromate induces disulfide crosslinking of proteins and EDC induces crosslinking between amino and carboxyl groups. Thermorheological properties compared to a control (no additive) were determined by torsion, dynamic (small strain) and stress relaxation testing. Profiles of gels with increased covalent cross-linking induced by potassium bromate or EDC showed increased rubber-elastic behavior. At the levels used, potassium bromate effected greater change in thermo-rheological response than EDC. These results confirmed that as covalent crosslinks increased, surimi gels become more rubber elastic in thermo-rheological responses.     

Suppression of Surimi Gel Setting by Transglutaminase Inhibitors

Three types of salted meat paste (3% NaCl, 3% NaCl plus 0.66% NH₄Cl or 3% NaCl plus 0.2% EDTA) were prepared from high and second grade surimi, set at 30°C up to 4 br, and subsequently heated at 85°C for 30 min. The gel strength, crosslinking of myosin heavy chain (MHC) and ?-(γ-glutamyl)lysine (?-(γ-Glu)Lys) content were determined. With extended setting time, gel strength, crosslinking of MHC and the content of a crosslinked product, ?-(γ-Glu)Lys, increased markedly in the gel from the high grade surimi. Such changes were suppressed considerably in the presence of NH₄Cl and EDTA and were not observed in the gel prepared from second grade surimi. These results indicated an active participation of intrinsic transglutaminase in the setting process.     

谷氨酰胺转氨酶对鳜鱼鱼糜凝胶的品质影响

以鳜鱼鱼糜热诱导凝胶为研究对象,检测盐度、持水性、色泽、质构、感官评分、蛋白质二级结构、微观结构以及十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(sodium dodecyl sulfate-polyacrylamide gel electrophoresis,SDS-PAGE)指标.研究谷氨酰胺转氨酶(transglutami...     

A study on the preparation and some functional properties of a cross‐linked casein–gelatin composite by a microbial transglutaminase

A microbial transglutaminase (TGase) was used in this work as biocatalyst to prepare a cross‐linked casein–gelatin composite, a modified protein product with 4‐hydroxyproline about 41 mg g^?1 peptides. Some cross‐linking conditions such as total protein content, the ratio of caseinate to gelatin and original pH were fixed at 5% (w/v), 4:1 (w/w) and 7.5, respectively. Other suitable conditions selected by single factor trials were TGase addition of 20 U g^?1 peptides, reaction temperature of 45 °C and reaction time of 4 h. Peptide profiles from SDS‐PAGE analysis showed the composite was peptide polymers. Compared to that of the original caseinate or the cross‐linked caseinate, the dispersion of the composite exhibited a markedly enhanced apparent viscosity, storage modulus and viscous modulus. Meanwhile, the composite also showed a better water holding capacity, unchanged oil binding capacity and lower enzymatic digestibility in vitro, conferring its applicability as a new protein ingredient.     

Draw pH and Storage Affect Rheological Properties of Mozzarella Cheese

Cheeses with whey pH at draw of 6.15 or 6.40 were stored at 4°for 50 days. Compression test at 10°(Instron) showed a decrease in compressive stress with storage. Failure points were near a true strain value of 0.5 for the cheeses at day 3. The failure region became less obvious by day 50. The hypothetical equilibrium stress (HES) in the stress-relaxation test was higher with a draw pH of 6.40 than 6.15, possibly indicating that higher calcium led to a stronger network structure. The HES decreased with storage, suggesting cleavage of structural crosslinks caused by the coagulant.     

Biochemical and Conformation Changes of Actomyosin from Threadfin Bream Stored in Ice

ABSTRACT: Biochemical and conformational changes of actomyosin stored in ice were investigated. The K-value of threadfin bream increased from 9% to 40% after storage for 12 d. Ca²⁺-, EDTA-, Mg²⁺-, and Mg²⁺-Ca²⁺-ATPase activities of actomyosin decreased, whereas Mg²⁺-EGTA ATPase activities increased. Total SH content of actomyosin increased after 3 d and decreased thereafter. Surface hydrophobicity gradually increased within 6 d. Protein loss during washing increased with storage time. A significant reduction (50%) of breaking force of thrice-washed mince was observed in fish stored in ice for 6 d. There was no evidence of proteolysis of muscle proteins stored up to 9 d as shown with SDS-PAGE.     

Protein Oxidation at Different Salt Concentrations Affects the Cross‐Linking and Gelation of Pork Myofibrillar Protein Catalyzed by Microbial Transglutaminase

In a fabricated then restructured meat product, protein gelation plays an essential role in producing desirable binding and fat‐immobilization properties. In the present study, myofibrillar protein (MFP) suspended in 0.15, 0.45, and 0.6 M NaCl was subjected to hydroxyl radical stress for 2 or 24 h and then treated with microbial transglutaminase (MTGase) in 0.6 M NaCl (E : S = 1 : 20) at 4 and 15 °C for 2 h. Protein cross‐linking and dynamic rheological tests were performed to assess the efficacy of MTGase for mediating the gelation of oxidized MFP. MTGase treatments affected more remarkable polymerization of myosin in oxidized MFP than in nonoxidized, especially for samples oxidized at 0.6 M NaCl. Notably, the extent of MTGase‐induced myosin cross‐linking at 15 °C in oxidized MFP improved up to 46.8%, compared to 31.6% in nonoxidized MFP. MTGase treatment at 4 °C for MFP oxidized in 0.6 M NaCl, but not MFP oxidized in 0.15 M NaCl, produced stronger gels than nonoxidized MFP (P < 0.05). The final (75 °C) storage modulus (G′) of oxidized MFP gels was significantly greater than that of nonoxidized, although the G′ of the transient peak (～44.5 °C) showed the opposite trend. Overall, oxidation at high salt concentrations significantly improved MTGase‐mediated myosin cross‐linking and MFP gelation. This might be because under this condition, MTGase had an increased accessibility to glutamine and lysine residues to effectively initiate protein–protein interactions and gel network formation.     

白鲢鱼糜流变特性的研究

本实验利用ARES-RDA流变仪,通过动态振荡测量,研究了不同温度和浓度下的白鲢鱼糜的流变特性、凝胶过程及动态粘弹性的变化规律。结果表明白鲢鱼糜的粘度随剪切速率增大而减小,存在剪切变稀行为;鱼糜的动态剪切模量(包括弹性模量和损耗模量)在5℃和10℃时几乎与动态频率(f)(2～16Hz)无关,其中G’远高于G’’,20℃时,当动态频率(f)>8Hz,糊状鱼糜的动态剪切模量与动态频率有关,随着频率的增加而增加;温度扫描(温度范围:5～70℃)结果表明鱼糜凝胶过程经历了三个阶段:第一段在30℃之前,第二段出现在38～43℃,第三段出现在46℃之后,蛋白开始形成了最终的凝胶。该结果为进一步扩宽白鲢鱼糜的应用范围有一定的借鉴意义。