Myosin—characteristics of the semi-tendinosus and extensor digitorum longus muscle of new-born and adult rabbits

• 1.1. Ca²⁺-ATPase activity of myosin, prepared from the semi-tendinosus muscle of new-born rabbits is higher than that of adult ones, and in contrast to that of adult slow semi-tendinosus muscle, myosin is stable on exposure to pH 9.4.
• 2.2. Myosin from the semi-tendinosus muscle of new-born rabbits reveals light chains of both fast and slow types, that from adult animals however reveals light chains of the slow type only.
• 3.3. Based on the high ATPase activity, stability to alkaline pH and the unchanged electrophoretic pattern of light chains, it is concluded that in contrast to the semi-tendinosus muscle, fast extensor digitorum longus contains both at birth and in the adult rabbit myosin of the fast type only.

     

Myofibrillar proteins in skeletal muscles of parr,smolt and adult atlantic salmon (Salmo salarl.). Comparison with another salmonid,the arctic charr Salvelinus alpinus (l.)

The heavy and light subunits of myosin from white and red muscles of Atlantic salmon parr, smolt and adult individuals were analyzed by SDS-PAGE and two-dimensional electrophoresis. Tropomyosin was identified by comigration with rat tropomyosins in two-dimensional gels in the presence and absence of urea. These myofibrillar proteins were compared to those of Arctic charr.

• 1.1. The myosin heavy chain from Atlantic salmon red muscles was associated with two types of light chain, 1S and 2S, that comigrated with the light chains 1S and 2S of Arctic charr.
• 2.2. As in the Arctic charr, four myosin light chain spots were detected in white muscles: two fast myosin light chains type 1, one of which comigrated with its analogous in the Arctic charr; one fast myosin light chain type 2, differing slightly in isoelectric point from that of Arctic charr; and one fast myosin light chain type 3 with higher electrophoretic mobility than that of Arctic charr.
• 3.3. Three tropomyosin spots were detected. White muscles contained only one type of β-tropomyosin and red muscles two types of α-tropomyosin. These three tropomyosin spots comigrated with those of Arctic charr.
• 4.4. Two myosin heavy chain bands were observed in red muscles of salmon parrs but only one in the rest of the red muscles analyzed.
• 5.5. Only one myosin heavy chain band was detected in white muscles by SDS-glycerol-polyacrylamide gel electrophoresis. Alfa-chymotryptic peptide mapping of these white myosin heavy chain bands revealed differences attributed to the presence of a new type of myosin heavy chain first detected several months after smoltification.

     

Diversity of native myosin and myosin heavy chain in fish skeletal muscles

• 1.1. Polymorphism of native myosin and myosin heavy chain (MHC) of fish skeletal muscles was analysed by pyrophosphate and SDS-gel electrophoreses.
• 2.2. Depending on the species, three or four myosin isoforms were detected in the white muscle, one or two isoforms in the pure red muscle, and four isomyosins were found in the red muscle composed of red and pink (intermediate) fibres.
• 3.3. It is suggested that all main types of fish muscle fibre (red, intermediate and white) differ in myosin isoform content.
• 4.4. Myosin heavy chain of the red muscle is a distinct protein from that of the white muscle. However, structural differences between these proteins vary among species.

     

Thermal stability of fish myosin

• 1.1. Thermal stability of fish myosin has been studied by using differential scanning calorimetry (DSC) and circular dichroism (CD).
• 2.2. The temperature range of the sharp decrease in α-helical content agreed very closely with that of the endothermic peaks.
• 3.3. There was a high correlation between the enthalpy of denaturation (ΔH) and the decreasing quantity in α-helicity (Δh).
• 4.4. The structure of fish myosins was much more unstable than that of rabbit.
• 5.5. The instability of fish myosins was reflected in its rod moiety.

     

Electrophoretic comparison of myosin light chains and parvalbumins of trunk and head muscles from two barbel (Barbus barbus) populations

• 1.1. Myosin light chains and parvalbumins have been compared in several trunk and head muscles from small and large size barbels (Barbus barbus) living in river or reared in hatchery.
• 2.2. These proteins isolated from white, red and ventricle fibres exhibit identical electrophoretic characteristics in the four batches.
• 3.3. The slow fibre content and the parvalbumin distribution are generally similar in river and hatchery barbels of the same size but differ between small and large barbels within a population.
• 4.4. Alterations of the mode of fertilization and breeding conditions do not modify the differentiation of myosin and parvalbumins in barbels.

     

Myosin polymorphism in urodelan amphibian dorsal skeletal muscle

• 1.1. Myosin isoforms were analyzed in the dorsal skeletal muscle of four urodelan amphibian species using a modified pyrophosphate gel electrophoresis which allowed better discrimination than classical methods.
• 2.2. The three fast and the intermediate isomyosins were characterized by a specific heavy chain, respectively HCf and HCi, associated with different combinations of the fast light chains LC1f, LC2f and LC3f.
• 3.3. Slow myosin was characterized by one (P. waltlii, T. palmatus, S. maculata) or two (T. alpestris) isoforms, combining a specific slow myosin heavy chain (HCs) with slow light chains only in the case of P. waltlii, or with slow and fast light chains in the other species.

     

In situ phosphorylation of contractile proteins of a molluscan Mytilus edulis catch muscle in different functional states

• 1.1. The incorporation of ³²P into the contractile proteins of the anterior byssus retractor muscle of Mytilus edilus L. was analyzed during the different stages of a contraction-catch-relaxatin cycle.
• 2.2. The experiments were performed with saponin-skinned fibers preincubated with γ-³²P-ATP.
• 3.3. The total amount of ³²P incorporated into the fiber proteins was anlyzed by measuring the label of TCA-insoluble protein in a scintillation counter.
• 4.4. The dose incorporated was about twice as high during Ca²⁺ induced contraction and serotonin induced accelerated relaxation as during test and catch.
• 5.5. The molecular mass of the phosphorylated proteins was analyzed by autoradiography of the proteins separated by SDS-PAGE.
• 6.6. Up to 26 protein spots of different molecular masses were labelled, including such well characterized protein spe+cies as myosin heavy and light chains, paramyosin and tropomyosin.

     

Lactate dehydrogenase of goldfish red and white muscle in relation to thermal accumulation

• 1.1. Activities of the red and white muscle LDH from 8°C-acclimated goldfish were about three times higher than those acclimated to 28°C.
• 2.2. Isozyme composition and some kinetic properties of the red muscle LDH differed from those of the white muscle enzyme.
• 3.3. The amount of red muscle as well as LDH activity tended to increase during cold acclimation.

     

The effect of pyridoxine deficiency on aminotransferase activity in liver and white muscle of rainbow trout (Salmo gairdneri Richardson)

• 1.1. Aspartate aminotransferase activity in the white muscle of rainbow trout dropped significantly after the animals had received a pyridoxine-deficient diet for 7 days.
• 2.2. Alanine aminotransferase activity in the white muscle of rainbow trout dropped significantly after the animals had been fed on a pyridoxine-deficient diet for 21 days.
• 3.3. Alanine and aspartate aminotransferase activity in the liver of rainbow trout did not drop significantly until the animals had received a pyridoxine-deficient diet for 28 days.
• 4.4. After rainbow trout have received a pyridoxine-deficient diet for 35 days, feeding with complete diet for 7 days is sufficient to restore the aminotransferase activities to the levels observed in control animals.

     

Proteolysis of skeletal muscle in yellowfin tuna (Thunnus albacares): Evidence of calpain activation

• 1.1. White muscle of yellowfin tuna is subject to a form of deterioration known as “burnt tuna”.
• 2.2. TEM and SDS-PAGE were used to quantify cellular differences in deteriorated white muscle of yellowfin tuna.
• 3.3. Electron micrographs showed a significant loss of Z-disc integrity and an increase in intracellular edema in burnt tuna.
• 4.4. Electrophoresis established that a specific doublet of proteins, 42 kD and 46 kD was lost.
• 5.5. Proteolysis of isolated myofibrils incubated in calpain (EC 3.4.22.17) was greatest at pH 7.5 and was selective for intermediate molecular weight proteins.
• 6.6. This evidence suggests that burnt tuna is a specific and limited proteolysis of myofibrillar structural proteins characteristic of calpain proteolysis.

     

The enzymatic properties of smooth muscle myosin B from dog colon

• 1.1. Smooth myosin B and myosin A were prepared from dog colon and their enzymatic properties were studied.
• 2.2. Colonic myosin B with two light chain corresponding to L₂ and L₃ in skeletal myosin showed much lower ATPase activities than rabbit skeletal myosin B.
• 3.3. The Mg²⁺-ATPase of myosin B was activated at high magnesium concentrations with the maximum activation between 10⁻³ and 10⁻²M and showed only a slight dependence on KCl concentration. On the other hand, Mg²⁺-ATPase activity of myosin A decreased with decreasing KCI concentration, suggesting the activation by actin of colonic myosin ATPase as much as skeletal myosin ATPase.
• 4.4. The pH dependence of Ca²⁺-ATPase showed a U-shaped curve although above pH 8.5 the activity was suppressed rapidly. The activity-ionic strength curve indicated that Ca²⁺- and ethylenediamine-tetraacetic acid (EDTA)-ATPase activities increased with increasing KCI concentration.
• 5.5. Mg²⁺-ATPase was fairly stable to urea treatment, whereas EDTA- and Ca²⁺-ATPase were activated by a low concentration of urea, followed by an inhibition.
• 6.6. These results were discussed as compared with those of skeletal myosin B.

     

The effect of changes in external salinity on the free amino acids and two aminotransferases of white muscle from fasted Salmo gairdneri Richardson

• 1.1. Aspartic acid. glutamic acid and serine concentrations in the white muscle of starved rainbow trout kept in diluted sea water (600 mOsm/l) for 8 days were significantly higher than in control animals kept in fresh water.
• 2.2. After 24 days the levels of all amino acids investigated (aspartic acid, glutamic acid, serine, glycine. alanine, threonine and lysine) in the white muscle of starved rainbow trout kept in diluted sea water were higher than in the white muscle of animals kept in fresh water without food.
• 3.3. Alanine aminotransferase activity in starved rainbow trout kept in diluted sea water for 24 days was higher than in the control animals kept in fresh water.
• 4.4. There is a significant correlation between alanine concentration and alanine aminotransferase activity in the white muscle of rainbow trout.

     

In vitro non-enzymatic glycosylation of myofibrillar proteins

• 1.1. Glycation is non-enzymatic modification of proteins by sugars in which not only structural but also biological properties of proteins are altered.
• 2.2. Our in vitro experiments show that incubation of myofibrillar proteins with ribose results in sugar attachment to proteins and at the same time myofibrillar ATPase activity is lowered.
• 3.3. DETAPAC, aminoguanidine and 2-mercaptoethanol all partially block myofibrillar protein glycation and ATPase activity is less inactivated.
• 4.4. The dependence of ATPase activity of myofibrils incubated with ribose on the amount of 2-mercaptoethanol present suggests that also modification of SH groups is involved in enzyme inactivation.
• 5.5. Electrophoretic studies revealed that heavy chains of myosin, actin, and tropomyosins are proteins which are mainly glycated in vitro.

     

Phylogenetic relationships between cyprinidae parvalbumins—I. Characterization of chub (Leuciscus cephalus L.) components

• 1.1. Three parvalbumins, the components II, IV and V, have been isolated from the white muscle of chub.
• 2.2. They bind two Ca²⁺ per mole and have the usual properties of parvalbumins, i.e. typical amino acid composition, molecular weight and u.v. spectra.
• 3.3. They can be divided in two chemically and immunologically distinct groups.

     

Purification and characterization of lipoprotein lipase from the white adipose,skeletal muscle,cardiac muscle,mammary gland and lung tissues of the rat

• 1.1. Lipoprotein lipase (LPL) was isolated from five rat tissues: white adipose, skeletal muscle, cardiac muscle, mammary gland and lung.
• 2.2. Specific activity of the preparations varied from 75 U/mg for skeletal muscle and 720 U/mg for adipose.
• 3.3. The preparations were further analysed using SDS-PAGE and a single component identified. The mol. wt of 61,000 Da of this component was consistent for all five of the tissue sources.
• 4.4. Significant differences in the values of the isoelectric points of the enzyme species were revealed. The values varied from 7.23 (SEM 0.022) for cardiac and lung to 7.51 (SEM 0.037) for mammary.
• 5.5. Two-dimensional electrophoresis, using isoelectric focusing in the first dimension and SDS-PAGE in the second revealed differences in the patterns of stained material derived from the five tissue sources.

     

Fluorescence studies on the interaction of muscle M-line proteins,creatine kinase and the 165,000 dalton component,with each other and with myosin and myosin subfragments

• 1.1. The binding of 8-anilino-l-naphthalene sulfonate (ANS) by M-line creatine kinase (CPK) and the 165,000 dalton protein component was studied by fluorescence.
• 2.2. One mole of ANS binds to a mole of each M-protein and the binding site on these two M-proteins is hydrophobic in nature.
• 3.3. M-line proteins labeled with ANS were used to demonstrate their interaction with myosin and myosin subfragments 1 and 2.
• 4.4. A unique finding of this study, that labeled M-line CPK binds to subfragment 2 of myosin, is of significance from a structural view point, since only the rod portions of myosin molecules are exposed at the M-line and therefore able to interact with CPK.
• 5.5. The use of a sulfhydryl fluorescent probe, N-(2-iodoacetyl)-N-(5-sulfo-l-napthyl) ethylene diamine, (1,5-AEDANS), has shown that the essential sulfhydryl group on CPK is very important for its interaction with both myosin and the 165,000 dalton M-line protein component.

     

Changes in myosin heavy chain isoform expression of overloaded rat skeletal muscles

• 1.1. The effect of functional overload produced by tenotomy of synergistic gastrocnemius muscle on the expression of myosin heavy chain (MHC) isoforms in the plantaris and soleus muscles of the rat was studied using gradient sodium dodecyl sulfate-acrylamide gel electrophoresis.
• 2.2. Five weeks tenotomy, the plantaris and soleus muscle weights induced by tenotomy of the gastrocnemius muscle were 44.3% (P < 0.005) and 37.4% (P < 0.005), respectively, heavier than the contralateral control muscles.
• 3.3. Although four types of MHC isoforms were observed in both control and experimental plantaris, the percentage of MHC isoforms in the control and experimental muscles differed; the hypertrophied plantaris muscle contained more HCI (P < 0.05), HCIIa and HCIId (P < 0.05) and less HCIIb (P < 0.05) than the control muscle.
• 4.4. The control soleus muscle contained two MHC isofonns, HCI and HCIIa. However, there was only a single HCI isoform in the hypertrophied soleus muscle.
• 5.5. These results indicate that overloading a skeletal muscle by removing its synergists produces not only the muscle hypertrophy but also the changes in the expression of MHC isofonns.

     

Comparative study on AMP deaminase in gill,muscle and blood of fish

• 1.1. High AMP deaminase activities were determined in the gill of one selachian, Scyliorhinus caniculus, and five teleosts, Anguilla anguilla, Cyprinus carpio, Salmo gairdneri, Perca fluviatilis and Esox lucius.
• 2.2. The highest activity was generally found in skeletal white muscle, except in A. anguilla and S. caniculus.
• 3.3. In s. caniculus a very high AMP deaminase activity was found in the blood where it was shown to be tightly regulated by inorganic phosphate.
• 4.4. Seasonal variations were observed for AMP deaminase activity in gill and white muscle, but also for blood Hb and protein concentration in the three tissues examined.

     

Partial purification and characterization of cysteine proteinase from metamorphosing tadpole tails of Rana catesbeiana

• 1.1. A leupeptin-sensitive proteinase was partially purified from regressing tadpole tails by acetone factionation and column chromatography on S-Sepharose.
• 2.2. The enzyme degraded hemoglobin and myoglobin at pH 3.0. The enzyme also hydrolyzed Z-Phe-Arg-MCA and Boc-Val-Leu-Lys-MCA at pH 4.0.
• 3.3. The enzyme activity was inhibited by leupeptin, egg cystatin, E-64 and monoiodoacetic acid and was activated by l-cysteine.
• 4.4. The enzyme degraded myosin and actin in myofibrils of tadpole tails.
• 5.5. The enzyme belongs to the cysteine proteinase and is possibly involved in tail degradation during the metamorphosis of tadpoles.

     

Mitochondrial and peroxisomal fractions derived from human white adipocytes

• 1.1. A procedure is described for the separation of intact peroxisomes from human white adipocytes using a linear metrizamide gradient (20–50% w/v).
• 2.2. Peroxisomes were found in the high density region of the gradient in an intact form.
• 3.3. Mitochondria were distributed in the high density and low density regions of the gradient.
• 4.4. Lysosomes separated well from the peroxisomes, occurring only in the low density region of the gradient.
• 5.5. Low levels of glyoxylate cycle enzyme activities (isocitrate lyase and malate synthase) were detected within the light and heavy mitochondrial pellet fractions.