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Protein crosstalking through osmotic work : the free energy of formation of the MgADP-myosin complexes at the muscle protein osmotic pressure
No full textA method is presented to determine the energy of formation of the myosin-ADP complexes at the muscle protein osmotic pressure. It is found that, at 1.8 x 10^5 dynes/cm^2, the putative protein osmotic pressure in skeletal muscle, the increase of MgADP from 0.05 to 2 mmol, increases the free energy of myosin- ADP and myosin-(ADP)2 by 7.56 x 10^9 and by 9.85 x 10^10 erg/mol, respectively, and decreases the free energy of myosin by 8.34 x 10^10 erg/mol. It is pointed out that the local changes of water chemical potential, induced by the binding of MgADP to myosin, can be sensed by other structures of the contractile machinery, wich per se may even be insensitive to MgADP. Crosstalking between macromolecules can thus be achieved by changes of the water chemical point
Myofibrils of skeletal muscle : the activity coefficient of orthophosphate
No full textIn the kinetic model of the cross-bridge cycle the rate limiting steps are dependent on the number of cross-bridges in the strongly bound. To analyze the orthophosphate concentration impact we studied the free energy of hydrolysis of ATP in muscle. We determined that at the physiological protein osmotic pressure the orthophosphate showed a classic activity coefficient (0.85) in the myofibrils of skeletal muscle Under the same conditions and at saturation, 2.67 mumoles of orthophosphate are bound per gram of dry myofibrils, with a dissociation constant of 7 x 10(-5) molal. Work is in progress to determine the activity coefficients of adenine nucleotide analogues
Mechanic and elastic properties of F-actin
No full textThe mechanic and elastic properties of F-actin were extensively investigated, but, surprising, no attention was paid to the influence of critical concentration on tensile strength. Formally, the critical concentration is the main determinant of the free energy of the monomer-monomer interaction, thus of tensile strength. We have investigated this aspect, the influence of critical concentration on tensile strength, by stretching rhodamnine phalloidin actin filaments by a laser trap. We found that by increasing ionic strength from 3 to 19 mM, critical concentration decreased fron 146 to 36 nM and the yield strength increased from 5.6 to 28.6 pN. At the ionic strength of 12-13 mM, the elastic modulus increased by 330-430 kP/nm up to break point, were it was 38-44.2 MP. Our finding poses the question of how actin filament bears the forces, ie how thin filament supports muscle contraction
Is nebulin truly a component of the thin filament
No full textNebulin is a protein coextensive with thin filament in skeletal muscle. Several evidences seem to indicate that nebulin may be a component of the thin filament.
We prepared thin filaments from rabbit and beef skeletal muscle by extraction of myofibrils at high and at low ionic strength. At the end of the purification procedure, nebulin was always found in the fraction containing the thick filaments and was absent from the fraction containing the thin filaments. We propose therefore that, even if nebulin binds to thin filaments, the binding is very week
Fructose diphosphatase from rabbit liver. VIII. The involvement of tyrosine residues in the catalytic activity
No full textFructose diphosphatase from rabbit liver. IX. Isolation and kinetic properties of the enzyme-substrate complex
No full textActomyosin interaction and mechanical performance modulation in skeletal muscle
No full textMuscle contraction involves the sliding of the myosin-containing thick filaments past the actin-containing thin filaments and the ATP hydrolysis by this actomyosin complex provides energy for contractile process. The force actively generated during contraction is transmitted to the end of the contractile unit through thin filaments, the contribution of titin, a sarcomere scaffold protein, is unclear. Our attention was focused on elastic and mechanic properties of single myofilaments in rigor condition, absence of ATP. Although earlier observations provided strong evidence that filament are rigid, recent results conflict with this proposal: thin and thick filaments are extensible during contraction. Thus, the thin filament properties are crucial parameters for the mechanism contraction. We used laser trap to pull actin filaments and S1-(myosin subfragment 1) decorated actin filaments. Since acto-myosin overlap region length is sliding related, and only a fraction of myosin heads is attached to actin, we tested several S1-decoration percentages, from 20 to 100%. We observed the strain due to the applied stress and calculated Young's modulus and filament elongation. The elastic responses to applied forces decrease when S1-decoration % increase. The maxima elongations decrease from 4.5% for F-actin to 3.4%, 1.9% and 0.5% for 20% 40% and 100% S1-decorated F-actin. We also measured the maximum strain supported by filaments and found a lineard relationship between S1-decoration % and applied tensile strength: forces are twice for 40% and more than 4 times for 100%. These results prove that crossbridges formations influence actin filaments stability, by strengthening the monomer-monomer interaction, and contribute to unload contraction force. we are aware that our observations are not exaustive, and our future aim is to understand the role played by actin associated protein
Fructose 1,6-diphosphatase from rabbit liver. XIII. The number of Mn2+ binding sites measured with 54Mn2+
No full textRhodamine phalloidin F-actin : critical concentration versus tensile strength
No full textA knowledge of actin's mechanical properties is crucial to understanding its role in motile and cytoskeletal system. It has long been assumed that thin filaments are rigid, however, this view has been challenged in recent years by accurate measurements of filament compliance. We have previously determined tensile strength, elastic modulus and work required to break the filament of phalloidin rhodamine actin be stretching filaments by a laser trap. these experiments have proved that mechanic and elastic properties and actin critical concentration are inversely related and posed the question of how thin filament bears the forces developed in muscle contraction. A solution migth be found in the interaction itself of the cross bridge with the thin filament. We have investigated this aspect by determining the yield strength of thin filament, we have found that myosin subfragment S1 delays the polimerization of F-actin. It is therefore likely that crossbridge attachment decreases the critical concentration of thin filament, by stregthening the interactions of the neighbouring actin monomers
Fructose 1,6-diphosphatase from rabbit liver. VI. Functional tyrosyl residues in the active center
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