Myosin myofibrillar proteins

In addition to the major proteins of striated muscle (myosin, actin, tropomyosin, and the troponins), numerous other proteins play important roles in the maintenance of muscle structure and the regulation of muscle contraction. Myosin and actin together account for 65% of the total muscle protein, and tropomyosin and the troponins each contribute an additional 5% (Table 17.1). The other regulatory and structural proteins thus comprise approximately 25% of the myofibrillar protein. The regulatory proteins can be classified as either myosin-associated proteins or actin-associated proteins. 

The smdy of tissue protein breakdown in vivo is difficult, because amino acids released during intracellular breakdown of proteins can be extensively reutilized for protein synthesis within the cell, or the amino acids may be transported to other organs where they enter anabohc pathways. However, actin and myosin are methylated by a posttranslational reaction, forming d-methylliistidine. During intracellular breakdown of actin and myosin, 3-methylhistidine is released and excreted into the urine. The urinary output of the methylated amino acid provides a rehable index of the rate of myofibrillar protein breakdown in the musculature of human subjects. 

In contrast to milk, where samples are primarily derived from cows, meat analysis has to be performed in samples of a widely different animal origin including cattle, lamb, swine, poultry, and fish. Muscle is a complex matrix with a pH of 5.7, composed of muscle fibers, various types of connective tissue, adipose tissue, cartilage, and bones. Sarcoplasmic proteins such as myoglobin, and glycolytic enzymes are soluble in water while the myofibrillar proteins such as myosin and actin are soluble in concentrated salt solutions (14). The connective tissue proteins, collagen and elastin, are insoluble in both solvents. 

The myofibrillar proteins make up 50-60% of the total protein of muscle cells. Insoluble at low ionic strengths, these proteins dissolve when the ionic strength exceeds -0.3 and can be extracted with salt solutions. Analysis of isolated mammalian myofibrils86 shows that nine proteins account for 96% or more of the protein myosin, which constitutes the bulk of the thick filaments, accounts for 43% and actin, the principal component of the thin filaments, 22%. 

Actomyosin. At high salt concentrations ( . . 0.6 M KC1), actin and myosin combine to form actomyosin filaments giving a highly viscous solution. Actomyosin retains the ATPase activity of myosin and demonstrates "super-precipitation" on the addition of ATP (24,34). As expected, there are differences between actomyosins of rabbit and fish with respect to solubility (10,22,35,36), viscosity (46) and ultracentrifugal behavior (477. Since actomyosin is the most readily available form of myofibrillar proteins from fish muscle, its behavior relative to deterioration during frozen storage has been most frequently studied. 

Among the above hypotheses, effects of lipids (4-17,59-62, 69-71,155-159), formaldehyde (160-166), and gas-solid interface TMJ appear to be very important in Gadoid fishes. Denaturation of myofibrillar proteins caused by free fatty acids and/or lipid peroxides must occur during frozen storage. To prove this, Jarenback and Liljemark have shown by electron microscopy that, in muscle stored frozen with added linoleic and linolenic hydroperoxides, myosin became resistant to extraction with salt solution (168). 

The major regulatory proteins located on an actin filament are troponin and tropomyosin, each occupying 5% of the total myofibrillar proteins. Both proteins confer calcium sensitivity on the ATP-actin-myosin interactions (see Section II). There are minor regulatory proteins that modify the fine structures of myosin and actin filaments and also of Z lines. 

Before summarizing the reactions generally occurring in proteins irradiated at low temperatures, it is instructive to review some of the major observations that have been made for certain representative proteins. Those selected for illustration are myoglobin, ribonuclease (RNase), the myofibrillar proteins myosin and actomysin, and gelatin. Wherever possible, comparisons will be made between results for fluid and frozen systems. 

Effects of linoleic acid and linoleic acid hydroperoxides on the myofibrils and the solutions of myofibrillar proteins of cod muscle have been proved using the electron microscopy (80). Linoleic acid hydroperoxides were ten times more effective than linoleic acid in reducing the amount of the protein in KCl-extracts from the myofibrils incubated with the acid or its hydroperoxides. Linoleic acid seemed to prevent the dissolution of the myofibril frame work but appeared not to impair the extraction of myosin while hydroperoxides appeared to cause a retention of A-bands (myosin) in the myofibrils. 

Protein ( histidine ) Methyltransferase. An enzyme which methylates histidine in proteins to give primarily 3-methylhistidine residues has been observed in myofibrillar protein and in the sarcoplasmic fraction of muscle homogenates (218). S-Adenosyl-L-methionine serves as the methyl donor for the enzyme. The enzyme has not been solubilized and purified. Very little is known about the substrate specificity of protein-(histidine) methyltransferase. Actins from a wide variety of species consistently contain one 3-N-methylhistidine residue per molecule (191, 219). It appears that myosin from white muscle contains two residues of 3-N-methylhistidine (one residue per heavy chain), whereas myosin from red muscle contains no 3-N-methylhistidine (220). The amino acid sequence around the methylated residue of rabbit skeletal muscle is (221) ... 

The myofibril is comprised of relatively few proteins compared with the 200 or more different proteins that exist in a muscle cell. Table V lists the known myofibril proteins of vertebrate skeletal muscle and the approximate amount of each protein in the myofibril. Note that myosin and actin comprise up to 75% of the myofibrillar proteins and together are necessary and sufficient for in vitro contraction. Contraction of... 

Sotelo, C.G., Pineiro, C., Perez-Martin, R.I., and Gallardo, J.M. 2000. Analysis of fish and squid myofibrillar proteins by capillary sodium dodecyl sulfate gel electrophoresis Actin and myosin quantification. Ear. Food Res. Technol. 211, 443-448. 

FIG. 2 Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of pork and chicken myofibrillar proteins treated with purified actinidin at various pH values. MHC, myosin heavy chain A, actin. Reprinted with permission from Nishiyama (2001, Figs. 2 and 5A). 

Actin and myosin are the principal contractile proteins of the three different t es of vertebrate muscle skeletal (or striated) muscle, cardiac muscle and smooth (non-striated) muscle. Muscle protein makes up approximately 40% of body protein. Actin comprises about 25% and myosin about 50% of myofibrillar protein. In striated muscles, myosin is the principal protein of the thick filaments, and actin the thin filaments. Shortening of the myofibrils is achieved by sliding the thick and thin filaments to increase the overlap between them (see Rayment Holdea 1994). Many of the other proteins assist in this process (see Harold, 1986). Actin and myosin also occur in non-muscle cells, but their spatial organisation is more variable and less regular and... 

In a muscle tissue (Fig lb), the fiber cells are thin and elongated as opposed to the polygonal cells in plants. The major myofibrillar proteins, myosin and actin, form the myofilament bundles in the sarcoplasm of the fiber cell. Bundles of fiber cells form the muscle tissue. Connective tissue distributed between individual (endomysium) and bundles of fiber cells (perimysium) as well as around the whole muscle (epimysium) holds the cells and the muscle tissue together. Majority of lipids is located in the adipose tissue depots associated with the connective tissue between the bundles of fiber cells in poultry and red meat as well as fish muscle [2]. 

The increased gel strength of beef surimi compared with beef MSM was thought to be due to the increased protein content. In addition, since the centrifugation stage also removed water-soluble proteins, the remaining protein is made up predominantly of the myofibrillar proteins actin and myosin. These salt-soluble proteins form a strong, highly elastic gel when heated. 

Lawrie (1985) classified meat proteins into three types salt-soluble (myofibrillar) proteins, water-soluble (sarcoplasmic) proteins, and insoluble connective tissue proteins. The myofibrillar proteins actin, myosin and actomyosin are the major proteins responsible for determining the heat stability of comminuted meat emulsions. 

By selecting specific muscle types the authors were able to separate different myofibrillar protein types slow-contracting, fast-contracting and heart. Since each muscle fibre type contains its own specific myosin chain, the gelation properties of fractions prepared from them vary. In fact, most muscles contain a mixture of both types of myosin, as well as a third (fast-contracting) type. Salt was required to produce gels however, once... 

---