Separation of large mammalian ventricular myosin differing in ATPase activity

To investigate a possible heterogeneity of human ventricular myosin, papillary muscles of patients with valvular dysfunction were examined using a modified native gel electrophoresis. Myosin was separated into 2 components termed VA and VB, whereby the VA to VB proportion appeared to depend on the ventricular load. The proportion of the faster migrating band VA was correlated (P<0.05) with end-diastolic pressure and the aortic pressure-cardiac index product. The regression based on these variables accounted for 67% of the variation in VA (R2=0.67). The VA proportion was, however, not significantly correlated with cardiac norepinephrine concentration. The ATPase activity of the 2 components of myosin was assessed from the Ca3(PO4)2 precipitation by incubating the gel in the presence of ATP and CaCl2. The ATPase activity of VA was 60% of that of VB. The VA and VB forms were observed also in the cat (31.4% VA), dog (32.1% VA), pig (28.5% VA), wild pig (33.7% VA), and roe deer (30.5% VA). VA and VB were not detected in the rat exhibiting the 3 isoforms V1, V2, and V3, rabbit (100% V3), and hare (86% V1). The data demonstrate a heterogeneity of large mammalian ventricular myosin, whereby an increased cardiac load appeared to be associated with a higher myosin VA proportion that exhibited a reduced ATPase activity.     

Effect of calponin on actin-activated myosin ATPase activity

Calponin inhibited the actin-activated myosin MgATPase activity in a dose-dependent manner without affecting the phosphorylation level of myosin light chain. This inhibition was Ca2(+)-independent. The decrease in enzymatic activity of myosin was correlated with binding of calponin to actin-tropomyosin filaments. Caldesmon showed a further inhibition of the calponin-induced inhibition of MgATPase activity of the thiophosphorylated myosin. Calponin-induced inhibition of the myosin MgATPase activity was reversed by the addition of calmodulin only in the presence of Ca2+. These results suggest that calponin acts as an inhibitory component of smooth muscle thin filaments.     

Reaction of myosin with salicylaldehyde II. Effect of salicylalation on the ATPase activity of myosin

The effect of salicylalation on the biological properties of myosin was studied.

1. 1. The ATPase activity of myosin is affected by salicylalation if the treatment is carried out at higher pH than 6.5. The Mg²⁺-activated ATPase shows a maximal curve with 250–380% maximal activation when 25–70 moles of salicylaldehyde are bound per mole of myosin. The EDTA-activated ATPase decreases with increasing salicylalation. Ca²⁺-activated ATPase shows a small increase with increasing salicylalation.

2. 2. Less salicylaldehyde is bound if the treatment is carried out in the presence of ATP, while that of PP_i does not affect the degree of salicylalation. The enzymic properties of myosins salicylalated in the presence of ATP or PP_i are not different from those of the samples treated in their absence.

3. 3. Salicylalation decreases ATP sensitivity of ATPase and superprecipitation of actomyosins reconstituted from salicylalated myosins only if more than 50 moles of salicylaldehyde are bound per mole myosin.

Effect of medium viscosity on the activity of myosin ATPase

The influence of increased medium viscosity on the activity of myosin Ca2+-ATPase has been studied in 28, 45 and 60% water-sucrose solutions at 25 degrees C. In the wide range of viscosities (10 divided by 430 mp) the rate constant of ATP hydrolysis displays the negative power-law dependence on solution viscosity with an index approximately -0,5. The obtained data confirm an idea about the existence of direct connection between the low-frequency liquid relaxations and structural dynamics of proteins and enzymes.     

Effect of swimming training on cardiac function and myosin ATPase activity in SHR

Male spontaneously hypertensive rats (SHR) and Wistar-Kyoto normotensive rats (WKY) were subjected to swimming training 6 times/wk, commencing at 4 wk of age, to determine whether this type of endurance exercise might alter contractile proteins and cardiac function in young adult SHR. The total duration of exercise was 190 h. Myofibrillar adenosinetriphosphatase (ATPase) activity was assayed at various free [Ca2+] ranging from 10(-7) to 10(-5) M. Ca2+-stimulated ATPase activity of actomyosin and purified myosin was determined at various Ca2+ concentrations both in the low and high ionic strength buffers. Actin-activated myosin ATPase activity of purified myosin was assayed at several concentrations of actin purified from rabbit skeletal muscle. Under all these conditions the contractile protein ATPase activity was comparable between trained and untrained WKY and SHR. Analysis of myosin isoenzymes on pyrophosphate gels showed a single band corresponding to V1 isoenzyme, and there were no differences between swimming-trained and nontrained WKY and SHR. Ventricular performance was assessed by measuring cardiac output and stroke volume after rapid intravenous volume overloading. Both cardiac index and stroke index were comparable in nontrained WKY and SHR but were significantly increased in the trained groups compared with their respective nontrained controls. These results suggest that myosin ATPase activity and distribution of myosin isoenzymes are not altered in the moderately hypertrophied left ventricle whether the hypertrophy is due to genetic hypertension (SHR) or to exercise training (trained WKY). Moreover, the data indicate that SHR, despite the persistence of a pressure overload, undergo similar increases in left ventricular mass and peak cardiac index after training, as do normotensive WKY.     

Effect of trinitrophenylation on myosin ATPase

1. The enzymic and actin binding properties of myosins trinitrophenylated to different extents in the presence or absence of ATP have been studied.

2. The enzymic properties of myosin trinitrophenylated in the absence of ATP are different from those of myosin treated in the presence of ATP even on trinitrophenylating an equal number of lysyl residues. On trinitrophenylation in the absence of ATP the EDTA-(K⁺-)activated ATPase and Ca²⁺-activated ATPase decrease while the Mg²⁺-activated ATPase considerably increases. In the presence of ATP the enzymic properties of myosin are much less affected by trinitrophenylation.

3. The actin binding capacity of trinitrophenylated myosin does not change, although its enzymic properties may be greatly altered, and even if its property to be activated by actin is completely lost.     

Effect of graded doses of nicotine on ovarian and uterine activities in albino rats

Nicotine (2 and 4 mg/kg body weight, i.p.) administered to albino rats for 20 days decreased the number of healthy follicles and increased the number of regressing follicles in the ovary. Uterine weight, its diameter, thickness of myometrium and endometrium and height of epithelium were reduced. Increase in the ovarian cholesterol level and decrease in glycogen content in nicotine treated rats indicate the inhibition brought in the steroidogenesis which is dependent on pituitary gonadotrophins. Decreased protein content of the ovary and uterus may be due to their retarded growth. Reduced number of estrous cycle with prolonged metaestrus and diestrus also supports the decreased estrogen synthesis responsible for cornification of vaginal smear in nicotine treated rats.     

Effect of the filamentous structure of myosin on the actomyosin ATPase activity

The ATPase activities of acto-heavy meromyosin and of acto-myosin minifilaments have been compared under the same conditions at low ATP (0.1 mM) and at several KC1 concentrations. The activities, which are strongly salt-dependent in both systems, have been found to be similar at high ionic strength (about 0.16 M) but different at lower ionic strength (0.06-0.07 M). Under this last condition, the catalytic constants kcat and Km are lower for acto-myosin minifilaments than for acto-heavy meromyosin ATPase. In addition, at low ionic strength, any decrease in the concentration of any of the ionic species (ATP, citrate, etc.) induces an increase in the interaction strength between myosin and actin filaments, as revealed by the Km changes. The presence of the troponintropomyosin complex and of Ca2+ also enhances the strength of this interaction. On the other hand, the occurrence of particular interactions between F-actin and myosin minifilaments is further substantiated by the phenomenon of superprecipitation which occurs when the ATP concentration decreases. The favourable effect of the organized structure of the myosin minifilaments on the ATPase activity of actomyosin is discussed.     

The specific activity of aldolase in the livers of old and young rats.

The catalytic activity of liver fructose-biphosphate aldolase (EC 4.1.2.13) of young and old rats in units per manomole sequence has been calculated. No evidence of the accumulation with age of altered enzyme molecules of low catalytic activity was obtained. This is contrary to results obtained using mice and rabbits and indicates that the accumulation of altered enzymes may not always be associated with aging. The possibility that altered proteins are formed, but do not accumulate, cannot be ruled out.     

Effect of staphylococcus active substances on ATPase activity of smooth muscle actomyosin and myosin

The effect of staphylococcus active substances--protein A (PA) and peptidoglican (PG) at concentrations 10(-6)-10(-2) mg/ml on the ATPase activity of pig stomach natural actomyosin and myosin was studied. It was shown that PA and PG at direct contact with smooth muscle contractile proteins caused the activation and inhibition of ATPase activity, respectively. On the basis of this investigation it was assumed that staphylococcal active substances were able to modify of the ATPase activity smooth muscle contractile proteins perhaps via direct action on the myosin molecule, which could be accompanied by conformational changes of the active center of myosin ATPase.     

Regeneration and transplantation of muscles in old rats and between young and old rats.

In order to compare the regenerative ability of skeletal muscle between young (5 month) and old (26 month) rats, sliced or intact extensor digitorum longus muscles were freely autografted into young and old rats and also reciprocally grafted from young to old inbred animals and vice versa. Sixty days after grafting, the transplants were analyzed for contractile and histochemical properties. There was a relative similarity between the contraction times of both normal control muscles and of all groups of transplants, although the contraction time tended to be prolonged and histochemical fiber pattern was more often found to be uniform in grafts of senescent animals. All groups of transplants possessed histochemically heterogeneous fiber types at 60 days. The experiments demonstrate that skeletal muscle in old rats possesses a substantial degree of regenerative ability and that the free tranpllantation of entire muscles in old animals is feasible.     

Protein synthesis, myosin ATPase activity and myofibrillar protein composition in hearts from tumour-bearing rats and mice.

Growing rats and adult weight-stable mice bearing a transplantable methylcholanthrene-induced sarcoma were compared with animals with various states of malnutrition. Heart protein synthesis was measured in vivo. Myocardial RNA, myofibrillar protein composition and the Ca2+-activated ATPase activity in heavy chains of native myosin were measured. 'Fingerprints' were made from myosin by trypsin treatment to evaluate possible structural changes in the protein. Cardiac protein-synthesis rate was decreased by 20% in growing tumour-bearing rats, by 35% in protein-malnourished (rats) and by 47% in starved rats, compared with freely fed controls (P less than 0.05). Adult tumour-bearing mice showed no significant decrease in myocardial protein synthesis. Pair-weighed control mice had significantly depressed heart protein synthesis. Protein translational efficiency was maintained in both tumour-bearing rats and mice, but was decreased in several groups of malnourished control animals. The Ca2+-activated myosin ATPase activity was decreased in all groups of malnourished animals, including tumour-bearing mice and rats, without any evidence of a change in cardiac isomyosin composition. We conclude that loss of cardiac muscle mass in tumour disease is communicated by both depressed synthesis and increased degradation largely owing to anorexia and host malnutrition. Increased adrenergic sensitivity in hearts from tumour-bearing and malnourished animals is not communicated by increased Ca2+-activated ATPase activity. This may be down-regulated in all groups with malnutrition, without any observable alterations in the isomyosin profile.     

Correlation of myofibrillar ATPase activity and myosin heavy chain content in ventricular and atrial myocardium of fish heart

In the fish heart, ventricular and atrial muscles contain different isoforms of native myosin and myosin heavy chain (MyHC) but the significance of this diversity is still not known. We have analysed ventricular and atrial myocardium of six freshwater fish species (goldfish, roach, bream, rudd, perch and pike-perch) using histochemical staining for myofibrillar ATPase activity as well as non-denaturing and SDS gel electrophoreses for native myosin and MyHC content. In the range of fish species studied, the intensity of ATPase reaction was higher in the atrial myocardium than in the ventricular myocardium and the composition of native myosin isoforms differed between these two muscles. The MyHC content in the cardiac muscle showed some species-related differences. In the goldfish, both atrial and ventricular cardiac muscle contained electrophoretically similar MyHC. In the other fish species, however, the ventricular myocardium showed electrophoretically faster MyHC than that present in the atrial myocardium. These results indicate that there are consistent and characteristic species-related differences between the ventricular and atrial muscles at the level of ATPase staining and the type of MyHC expressed. The findings suggest that fish ventricular and atrial muscles may differ in their contractile properties.