Degradation Of Myosin Heavy Chain Research Articles

Effect of preheating temperature on the microstructure of walleye pollack surimi gels under the inhibition of the polymerisation and degradation of myosin heavy chain

The physical attribute of heat-induced gel texture is highly dependent on the microstructure of the gel. In this study the microstructures of walleye pollack surimi gels preheated at various temperatures with and without inhibitors (ethylenediamine-N,N,N',N'-tetraacetic acid, iodoacetamide and leupeptin) were observed with a natural scanning electron microscope. Without inhibitors, gels preheated at 30 °C showed a fine and uniform network structure together with the highest polymerisation of myosin heavy chain (MHC) and the highest gel strength. At 60 °C, gels exhibited a broken, disrupted and loose cluster-like structure together with the highest degradation of MHC and the lowest gel strength. Under the inhibition of polymerisation and degradation of MHC a fine network was observed up to 40 °C during preheating. However, after a second step of heating at 80 °C the microstructures were disrupted and resembled each other regardless of the preheating temperature. Heat-induced gel formation is related to the polymerisation and degradation of MHC and the microstructure of the gel during preheating. Gelation occurred during setting even under the inhibitory condition, and the formation of covalent bonding by transglutaminase is not essential to the formation of a three-dimensional network during setting but is essential to the gel strength enhancement effect of setting by subsequent heating at 80 °C.

Original article: Gel properties of red tilapia surimi: effects of setting condition, fish freshness and frozen storage

SummaryThis study aimed to determine effects of setting condition, fish freshness and storage time of frozen surimi on properties of red tilapia surimi gel. To investigate the effect of setting condition, a combination of eight setting temperatures (35–70 °C) and four setting times (30–120 min) was used. Maximum breaking force, deformation and gel strength were obtained after the gel had been set at 40 °C for 90 or 120 min. Setting at 65 °C resulted in the lowest obtained gel strength, because of proteolytic degradation of myosin heavy chain. Increasing storage time of raw fish material in ice caused a significant decrease in gel strength of the resultant surimi gel (P < 0.05). Gels produced from surimi kept in frozen storage for up to 9 months also exhibited reduced gel strength, with a concomitant increase in the expressible drip, with increasing storage time (P < 0.05).

Post-mortem degradation of myosin heavy chain in intact fish muscle: Effects of pH and enzyme inhibitors

Fish muscle is rapidly degraded during post-mortem storage, due to proteolytic enzymes acting probably both on muscle cells and connective tissue. In this work we have developed a model system which may be used to study the enzymatic degradation occurring in intact post-mortem fish muscle. Degradation of myosin heavy chain (MHC) was monitored in muscle with pH adjusted to 6.05, 6.3 and 6.9 and in the presence of the enzyme inhibitors PMSF, EDTA, phenanthroline, pepstatin A, antipain, E-64 and the cysteine proteinase activator dithiothreithol (DTT). After storage, myofibrillar proteins were isolated and MHC-specific antibodies used to study the degradation in the different samples. MHC from muscle with pH 6.05 and 6.3 was degraded, while no severe degradation was observed at pH 6.9. Introduction of enzyme inhibitors into the muscle tissue clearly showed that mainly cysteine and aspartic proteinases are responsible for the in situ MHC degradation. This is supported by the severe breakdown of MHC in the muscle samples containing DTT.

Effect of sodium lactate as cryostabilizer on physico-chemical attributes of croaker (Johnius gangeticus) muscle protein

Effect of sodium lactate as cryostabilizer on physico-chemical attributes of croaker (Johnius gangeticus) fish muscle protein was studied during freezing and frozen (-20 ± 2°C) storage for 3 months. Minced meat was mixed with 4% sucrose, 4% sorbitol, and 0.3% sodium tri poly phosphate (STPP) (T1), minced meat was mixed with 6% (w/v) sodium lactate and 0.3% STPP (T2) and control (C) was without any additive. The decreasing rate of Ca(2+) ATPase activity, thaw drip, water holding capacity and relative viscosity in T1 and T2 samples from that of C was significantly lower, indicating higher protective effect of additives. In case of cryoprotectant treated samples, the degradation of myosin heavy chain was much lower than that of C which prevents the aggregation and subsequent insolubilization of myosin during frozen storage. The sodium lactate prevented Ca(2+)ATPase activity more than that of sucrose/sorbitol during isothermal storage at -20 ± 2°C for 3 months. This inferred that sodium lactate can effectively be used as an alternative cryostabilizer to sucrose/sorbitol for stabilization of croaker muscle protein native structure.

Edible film from squid ( Todarodes pacificus) mantle muscle

Mantle muscle meat of Japanese common squid ( Todarodes pacificus) was used to produce edible films in this study. The solubility of squid mantle muscle proteins increased upon addition of NaCl and organic salts (Na-citrate, Na-benzoate, Na-acetate and Na-tartrate). At concentrations below 2%, among organic salts, Na-citrate appeared to have the highest ability to dissolve proteins. Film-forming solutions were prepared by dissolving squid mantle meat in 0.5% or 1% salt solutions at neutral pH. Physical properties and SDS–PAGE of the films were determined. The film-forming solution prepared with 0.5% Na-citrate gave the transparent film with the highest tensile strength and least degradation of myosin heavy chain. Organic salt type at the same concentration had no significant effects ( p ⩾ 0.05) on water vapour permeability (WVP) of the films. The films showed excellent UV barrier properties. In addition, organic salts gave no significant effects ( p ⩾ 0.05) on light barrier properties and transparency of the films except for NaCl and Na-tartrate which were crystallised during the drying process. These results suggested that Na-citrate is the most useful for producing the edible films from squid mantle muscle among the organic salts examined in this study.

Preventive effect of tannic acid in combination with modified atmospheric packaging on the quality losses of the refrigerated ground beef

Chemical, microbiological and sensorial changes of ground beef treated without and with tannic acid (200 mg/kg) and stored in air and under modified atmosphere packaging (MAP) (80%O 2/20%CO 2 or 10%O 2/20%CO 2/70%N 2) were monitored during 15 days of storage at 4 °C. During the storage, samples treated with tannic acid and kept under all packaging conditions contained lower peroxide value (PV) and thiobarbituric acid-reactive substances (TBARS) with coincidental lower non-haem iron content, compared with non-treated counterparts ( P < 0.05). The sample packed in high oxygen MAP treated without and with tannic acid had the higher oxymyoglobin and a * values and received the higher likeness scores for colour, whereas the samples stored in air and under low oxygen MAP showed the lower values, regardless of tannic acid treatment. After 15 days of storage, myosin heavy chain (MHC) and actin of all tannic acid treated samples underwent less degradation than those without tannic acid treatment for all packaging conditions. Degradation of MHC was more pronounced in samples kept under MAP with high oxygen. Psychrophilic bacterial count (PBC) of all tannic acid treated samples was lower, compared with that of non-treated samples ( P < 0.05), irrespective of packaging condition. Therefore, tannic acid treated samples stored under high oxygen MAP could maintain the red colour and retard lipid oxidation and microbial growth of ground beef during refrigerated storage.

Protein nitration is associated with increased proteolysis in skeletal muscle of bile duct ligation–induced cirrhotic rats

Cirrhosis is characterized by skeletal muscle wasting. In this study, the effects of nitric oxide production on skeletal muscle protein nitration and degradation in cirrhosis were investigated. Cirrhosis was induced by bile duct ligation (BDL) in Sprague-Dawley rats for 4 weeks. The BDL-induced cirrhotic rats and sham-operated rats were then injected daily with either saline or N G - l-nitro-arginine methyl ester ( l-NAME) for 7 days from week 4 to week 5, after which nitrite/nitrate, glutathione reduction, as well as protein nitration, ubiquitination, and degradation were assessed in skeletal muscle. Elevated muscular nitrite/nitrate concentrations, protein nitration, total ubiquitin conjugates, and degradation fragments of myosin heavy chain as well as diminished glutathione reduction levels were observed in BDL-induced cirrhotic rats as compared with controls. Administration of l-NAME for 1 week led to reduction of nitrite/nitrate levels; protein nitration was also decreased in the skeletal muscle. In addition, ubiquitination of muscular proteins and degradation of myosin heavy chain were significantly diminished after treatment of l-NAME. In conclusion, nitrosative stress occurred in the skeletal muscle of BDL-induced cirrhotic rats and may lead to increased proteolysis of muscle-specific structural proteins.

Application of a Kiwifruit (Actinidia chinensis) to Improve the Textural Quality on Beef Bulgogi Treated with Hydrostatic Pressure

In order to reduce the increased hardness of beef bulgogi due to hydrostatic pressure (HP), kiwifruit (Actinidia chinensis) was applied. To understand the changes of shear force in beef bulgogi with kiwifruit induced by HP, changes in chemical properties of myofibril (Mf) with 10% kiwifruit induced by HP were investigated. From the SDS-PAGE patterns of Mf with 10% kiwifruit, there was an observed increase in the degradation of myosin heavy chain (MHC) by HP (300-500 MPa) to that by 0.1 MPa. This result indicates that HP may enhance enzyme action from a kiwifruit for the degradation of MHC, and the similar phenomenon occurred in the beef bulgogi with kiwifruit induced by HP. The shear force of beef bulgogi without a kiwifruit induced by 400 and 500 MPa significantly increased compared to that by 0.1 MPa (p

Myosin heavy chain degradation during post mortem storage of Atlantic cod (Gadus morhua L.)

Abstract Post mortem proteolytic degradation of fish fillets leads to textural changes like muscle softening and gaping. In this study proteolytic degradation of myosin heavy chain (MHC) was monitored during storage of muscle and of isolated myofibrils at different temperatures and pH-values by the use of MHC-specific antibodies. The ability of cathepsin D to associate to myofibrillar proteins was also studied. Muscle stored at 6 °C and isolated myofibrils stored at 0 °C, 6 °C and 20 °C were degraded at pH 6.3 or lower. Cathepsin D could be found associated with extensively washed myofibrils. Inhibition of cathepsin D during storage affected the observed MHC-degradation at pH 5.5, but not at pH 6.3. This indicates that cathepsin D to a less extend than formerly believed, is responsible post mortem degradation of MHC.

Rheological and Textural Properties of Pacific Whiting Surimi Gels As Influenced by Chicken Plasma

Rheological properties of Pacific whiting surimi, in the absence and presence of chicken plasma (CP) at different levels (0.3–3.0%, w/w), were studied by dynamic rheological (small strain) and torsion fracture measurements, respectively. The surimi paste exhibited two major distinctive rheological transitions during heating (1°C/min) from 20 to 90°C with peaks observed at 33 and 56°C. The abrupt loss of G′ upon heating from 47 to 57°C, and the occurrence of small peak of phase angle at the same temperature range were prevented by the addition of CP. Nevertheless, the final G′ of the surimi paste added with CP was lower than that of the control. But shear fracture stress of both kamaboko and modori gels increased as the CP levels increased and shear strain increased with the addition of CP up to 2% (P < 0.05). CP inhibited the degradation of myosin heavy chains (MHC) caused by endogenous proteinases as indicated by more retained MHC and lowered TCA-soluble peptide content. Whiteness of gels decreased somewhat with increasing CP levels. CP, thus, could be a helpful additive for improving gelling properties of Pacific whiting surimi

EFFECTS OF ENDOGENOUS CATHEPSINS B AND L ON DEGRADATION OF SILVER CARP (HYPOPHTHALMICHTHYS MOLITRIX) MYOFIBRILLAR PROTEINS

ABSTRACT The activities of cathepsins B and L in minced silver carp meat were 4.89 and 3.81 units/g, respectively. About 11.30% activity for cathepsin B and 25.61% activity for cathepsin L were left after washing with distilled water, suggesting a short wash removed a large portion of both cathepsins B and L, but the activities of cathepsins still remained to some extent, and the remaining activity of cathepsin L was higher than that of cathepsin B. Higher cathepsin residual activity was correlated with the autolytic rate of myosin heavy chain (MHC) during the autolysis of myofibrillar proteins at 50C. The sodium dodecyl sulfate‐polyacrylamide gel electrophoresis analysis indicated that both of the purified cathepsins B and L could result in significant degradation of MHC at 50 and 35C, which was close to the temperature of gel softening and setting, respectively, but cathepsin L seemed to be a key proteinase for hydrolyzing the myofibrillar protein. PRACTICAL APPLICATIONThere is a possibility of consuming silver carp surimi in the future because the catches are large. Silver carp is very suitable for surimi because of its attractive white color and taste. Consumption in China and India as fresh fish is large. However, silver carp surimi exhibited considerable gel softening (modori) in the course of heating, resulting in a decrease in textural quality, eventually having a negative effect on acceptance and price of the surimi products. For this purpose, this paper investigates the effects of cathepsins B and L on modori, and compares the roles of pure cathepsins in degradation of myosin at similar conditions as for processing of silver carp surimi products.

Properties of protein-based film from round scad (Decapterus maruadsi) muscle as influenced by fish quality

The properties of film prepared from round scad (Decapterus maruadsi) stored in ice for different times were investigated. Degradation of myosin heavy chain (MHC) was more pronounced with the coincidental increase in total volatile base and trimethylamine contents as the storage time increased (P 0.05). For the films prepared from washed mince, TS decreased, whereas EAB increased when the storage time of fish increased (P<0.05). However, films prepared from washed mince showed the greater mechanical properties with the lower film solubility and protein solubility than did those from mince (P<0.05). Generally, films prepared from fish stored in ice for a longer time became less transparent, darker and more yellowish. The electrophoretic study revealed that similar protein patterns were observed between films, irrespective of storage time of fish and washing. Therefore, the quality of fish did not show the marked impact on the mechanical property of the resulting films, while washing likely affected the film forming ability.

Effect of Fish Meat Quality on the Properties of Biodegradable Protein Films

Fish muscle protein films were prepared from blue marlin (Makaira mazara Jordan & Evermann) meat which had been stored at 30°C to intentionally lower the meat quality. In this study, the effects of meat quality and pH on the formation of these protein films were investigated. Moreover, ε-polylysine was added to the film-forming solutions to reduce the microbial population of films. The mechanical properties of the films were slightly affected by acidic and alkaline pHs. However, the water vapor permeability of muscle protein films was not affected by either the quality of the fish meat or the pH of the film-forming solutions. SDS-PAGE (sodium dodecyl sulfate- polyacrylamide gel electrophoresis) showed the degradation of myosin heavy chain in acidic films and polymerization in alkaline films. It was revealed that biodegradable films can be produced even from very low quality fish meat, and that the bacterial population of films could be drastically reduced by the addition of ε-polylysine.

Effect of endogenous acid proteinases on the properties of edible films prepared from Alaska pollack surimi

The existence of endogenous acid proteinases in Alaska pollack surimi and their effect on mechanical properties of surimi films were investigated. The optimum pH of acid proteinases involved in the degradation of myosin heavy chain (MHC) was 3.0, and the optimum temperature was 45 °C. The degradation of MHC was completely inhibited by pepstatin A together with any one of cysteine proteinase inhibitors, suggesting that acid proteinases present in surimi are mainly cathepsin D and cysteine proteinases. The concomitant decrease of surimi film strength with the extent of MHC degradation was observed, but surimi films were formed even when most of MHC was degraded. The main associative forces responsible for the surimi films prepared at pH 3.0 were ionic bonds and hydrophobic interactions.

Properties of Edible Surimi Film as Affected by Heat Treatment of Film-Forming Solution

The effect of heat treatment of film-forming solutions on the properties of edible surimi films was investigated. The film-forming solutions prepared at pH 3 from frozen Alaska pollack surimi were heated to 45, 70 or 100°C to promote unfolding of surimi protein molecules. As a result, solubility, surface hydrophobicity, and reactive SH group of surimi proteins increased. After 45°C -treatment, the mechanical properties, film solubility, and protein solubility of surimi films were not affected and myosin heavy chain (MHC) of surimi proteins was degraded by endogenous acid proteinases. Conversely, at higher heating temperatures (70°C, 100°C), degradation of MHC was effectively inhibited and mechanical properties were improved, while the film solubility and protein solubility of surimi films decreased. It is revealed that the prevention of MHC degradation by heat treatment could improve mechanical properties of surimi films. The optimum condition was found to be heating the film-forming solutions (pH 3) at 70°C for 20min.

Effect of cysteine proteinase inhibitor containing fraction from chicken plasma on autolysis and gelation of Pacific whiting surimi

The effects of cysteine proteinase inhibitor containing fraction (CPI fraction) from chicken plasma on autolysis inhibition and gelling properties of Pacific whiting surimi were investigated. The CPI fraction exhibited the inhibitory activity against autolysis of Pacific whiting surimi in a concentration-dependent manner. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis pattern revealed that the CPI fraction effectively prevented the degradation of myosin heavy chain, tropomyosin and troponin-T in Pacific whiting surimi incubated at 55 °C for 60 min as well as modori gel (55 °C for 30 min/90 °C for 20 min). The breaking force and deformation of modori gels increased as the concentration of CPI fraction increased up to 2.0% with a concomitant decrease in TCA-soluble peptides ( p<0.05). However, a decrease in breaking force and deformation was noticeable with the modori gel added with 3% CPI fraction. The addition of CPI fraction had no detrimental effect on whiteness of surimi gels ( p>0.05). The result suggested that CPI fraction would be an alternative additive for surimi containing the active cysteine proteinase causing the gel weakening.

Myofibrillar proteolysis by myofibril-bound serine protease from white croaker Argyrosomus argentatus

The modori phenomenon is defined as heat-induced myofibrillar degradation caused by endogenous serine protease(s) of fish muscle during Kamaboko fish meat gel production. This study was undertaken to analyze myofibrillar proteolysis of white croaker Argyrosomus argentatus muscle, which is an ingredient of high quality Kamaboko, by myofibril-bound serine protease (MBSP) under conditions corresponding to the modori phenomenon. White croaker MBSP was stable between pH 2–11 and below 65°C, and about 60% of its initial activity remained after incubation for 2 h under the conditions at 65°C and pH 7.5. About 60% of the enzyme activity was suppressed by 0.5 M NaCl. White croaker MBSP degraded various myofibrillar proteins between 40 and 70°C and pH 6.0–9.0, and preferentially degraded myosin heavy chain rather than other myofibrillar proteins. The enzyme degraded the myosin heavy chain most strongly at 55°C and pH 7.0, and a major part of the bands of myosin heavy chain and its degradation products disappeared for a period of 2 h. These degradation characteristics are very similar to those observed during the modori phenomenon, indicating that MBSP could be a modori-inducing protease involved in the modori phenomenon of white croaker Kamaboko production.

Degradation of myofibrillar proteins by a myofibril-bound serine proteinase in the skeletal muscle of crucian carp ( Carasius auratus)

A myofibril-bound serine proteinase (MBSP) in the skeletal muscle of crucian carp ( Carasius auratus) was identified. Hydrolysis of myofibrillar proteins by the endogenous MBSP was studied. Myosin heavy chain (MHC) was degraded markedly when crucian carp myofibril was incubated at 55 °C, as shown by SDS-PAGE. Prolonged incubation of myofibril at 55 °C also caused the obvious degradation of tropomyosin, while the decomposition of other myofibrillar proteins, such as α-actinin and actin, was slight, as detected by Western blotting. The results suggest the existence of an endogenous myofibril-associated proteinase in crucian carp myofibril, which efficiently cleaves MHC and tropomyosin. Serine proteinase inhibitors (Lima bean trypsin inhibitor, PMSF and benzamidine) greatly suppressed the degradation of MHC, caused by the enzyme, while inhibitors for cysteine, metallo-, and aspartic proteinases showed only partial or incomplete inhibitory effects, indicating that the endogenous proteinase is a serine proteinase. Substrate specificity analysis, using partially purified crucian carp MBSP, suggested that the enzyme is a trypsin-like serine proteinase.

IDENTIFICATION OF A MYOFIBRIL-BOUND SERINE PROTEINASE IN THE SKELETAL MUSCLE OF SILVER CARP

Myofibril-bound serine proteinase (MBSP) in the skeletal muscle of silver carp was characterized. Myosin heavy chain (MHC) degraded markedly when silver carp myofibril was incubated at 55-60C as shown by SDS-PAGE. Prolonged incubation of myofibrils also caused the degradation of other myofibrillar proteins such as α-actinin, actin and tropomyosin to some degree. The results suggest the existence of an endogenous myofibril associated proteinase. Serine proteinase inhibitors (Pefabloc SC and Lima bean trypsin inhibitor) greatly suppressed the degradation of myosin heavy chain, while inhibitors for cysteine, metallo, and aspartic proteinases did not show any effect, indicating that the endogeneous proteinase is a myofibril-bound serine proteinase.

Effects of Alkali and Acid Solubilization on Gelation Characteristics of Rockfish Muscle Proteins

ABSTRACT: Solubility of rockfish whole muscle and actomyosin was minimum at pH 5 and gradually increased as the pH was shifted to acidic or alkaline pH. Acidic and alkaline solubilization was followed by isoelectric precipitation induced degradation of myosin heavy chain, resulting in a protein band of about 120 kDa. Both myofibrillar and sarcoplasmic proteins underwent denaturation after acidic and alkaline treatment, exhibiting minimal solubility and absence of endothermic peaks. Acid‐ and alkali‐treated muscle proteins readily aggregated upon heating, showing different dynamic rheological patterns compared with whole muscle and washed mince. Disulfide linkages occurred at a greater extent in gel prepared by alkaline solubilization, resulting in higher breaking force and deformation.