Biomedical subjects

S Chaen

Publications and source records attributed to S Chaen.

27 records · Page 2Linked to original sources

PubMed1991

Measurement of work done by ATP-induced sliding between rabbit muscle myosin and algal cell actin cables in vitro.

1. The basic properties of the ATP-dependent actin-myosin interaction responsible for muscle contraction were studied using an in vitro force-movement assay system, in which a glass microneedle coated with rabbit skeletal muscle myosin was made to slide on the actin filament arrays (actin cables) in the internodal cell of an alga Nitellopsis obtusa with ionophoretic application of ATP. 2. In response to an ATP current pulse (intensity, 5-85 nA; duration, 0.5-10 s), the myosin-coated needle moved for a distance and eventually stopped, indicating reformation of rigor actin-myosin linkages to prevent elastic recoil of the bent needle. A subsequent ATP current pulse again produced the needle movement starting from the baseline force attained by the preceding needle movement. 3. With a constant amount of ATP application, the amount of work done by the ATP-induced actin-myosin sliding first increased with increasing baseline force from zero to 0.4-0.6P0, and then decreased with further increasing baseline force, thus giving a bell-shaped work versus baseline force relation. 4. With increasing amount of ATP application, the amount of work done by the actin-myosin sliding increased more steeply as the baseline force was increased from zero to 0.4-0.6P0. 5. These results are discussed in connection with the basic properties of the actin-myosin sliding in muscle contraction.

Actins↗

PubMed1990

Steady-state force-velocity relation in the ATP-dependent sliding movement of myosin-coated beads on actin cables in vitro studied with a centrifuge microscope.

To eliminate the gap between the biochemistry of actomyosin in solution and the physiology of contracting muscle, we developed an in vitro force-movement assay system in which the steady-state force-velocity relation in the actin-myosin interaction can be studied. The assay system consists of the internodal cells of an alga, Nitellopsis obtusa, containing well-organized actin filament arrays (actin cables); tosyl-activated polystyrene beads (diameter, 2.8 microns; specific gravity, 1.3) coated with skeletal muscle myosin; and a centrifuge microscope equipped with a stroboscopic light source and a video system. The internodal cell preparation was mounted on the rotor of the centrifuge microscope, so that centrifugal forces were applied to the myosin-coated beads moving along the actin cables in the presence of ATP. Under constant centrifugal forces directed opposite to the bead movement ("positive" loads), the beads continued to move with constant velocities, which decreased with increasing centrifugal forces. The steady-state force-velocity curve thus obtained was analogous to the double-hyperbolic force-velocity curve of single muscle fibers. The unloaded velocity of bead movement was 1.6-3.6 microns/s (20-23 degrees C), while the maximum "isometric" force generated by the myosin molecules on the bead was 1.9-39 pN. If, on the other hand, the beads were subjected to constant centrifugal forces in the direction of bead movement ("negative" loads), the bead also moved with constant velocities. Unexpectedly, the velocity of bead movement did not increase with increasing negative loads but first decreased by 20-60% and then increased towards the initial unloaded velocity until the beads were eventually detached from the actin cables.

Actins↗

PubMed1989

Simultaneous recordings of force and sliding movement between a myosin-coated glass microneedle and actin cables in vitro.

To elucidate the molecular mechanism of muscle contraction resulting from the ATP-dependent actin-myosin interaction, we constructed an assay system with which both the force and the movement produced by the actin-myosin interaction in vitro can be simultaneously recorded and analyzed. The assay system consisted of the giant internodal cells of an alga, Nitellopsis obtusa, which contain well-organized arrays of actin filaments (actin cables) running along the cell long axis, and a glass microneedle (tip diameter, approximately 7 microns; elastic coefficient, approximately 40 pN/microns), which was coated with skeletal muscle myosin at the tip and extended from a micromanipulator at right angles with the actin cables. When the myosin-coated tip of the microneedle was brought into contact with the exposed surface of the actin cables, it exhibited ATP-dependent movement along the actin cables over a distance of 20-150 microns in 20-200 s (20-23 degrees C) and eventually stopped due to a balance between forces generated by the actin-myosin interaction (800-6000 pN) and by the bent microneedle. Since the load on the force-generating myosin molecules increased with the bending displacement of the microneedle (auxotonic condition), the relation between the load and the sliding velocity of the myosin heads past the actin cables was determined from the time course of the microneedle movement recorded with a video system. The shape of the force-velocity curve thus obtained was convex upwards, similar to that of the force-velocity curve of intact frog muscle fibers obtained under the auxotonic condition.

Actins↗

PubMed1988

Cooperative interactions of myosin two heads in muscle force generation.

To investigate the possibility of cooperative interactions between the two myosin heads in muscle contraction, Ca2+-activated force development, K+-EDTA- and Mg2+-ATPase activities, muscle fiber stiffness, and the velocity of unloaded shortening were measured on partially p-PDM treated glycerinated muscle fibers, which contained a mixture of myosin molecules with zero, one and two of their heads inactivated. It was found that the magnitude of the Ca2+-activated isometric force development was proportional to the square of both K+-EDTA- and Mg2+-ATPase activities and also to the square of muscle fiber stiffness. If the two myosin heads in the glycerinated fibers are assumed to react independently with p-PDM, the above results strongly suggest that (i) each myosin molecule in the thick filaments can generate force only when its two heads do not react with p-PDM, (ii) muscle fiber stiffness is determined by the total number of native heads, and (iii) there is no cooperative interaction between the two myosin heads in catalyzing ATP hydrolysis.

Adenosine Triphosphatases↗

PubMed1986

Evidence for cooperative interactions of myosin heads with thin filament in the force generation of vertebrate skeletal muscle fibers.

To examine the possibility of cooperative interactions between the two myosin heads in muscle contraction, Ca2+-activated force development, K+-EDTA-and Mg2+-ATPase activities, muscle fiber stiffness, and the velocity of unloaded shortening were measured on partially p-phenylenedimaleimide (p-PDM)-treated glycerinated muscle fibers, which contained a mixture of myosin molecules with zero, one, and two of their heads inactivated, and the relationships among these values (expressed relative to the control values) were studied. It was found that the magnitude of the Ca2+-activated isometric force development was proportional to the square of both K+-EDTA- and Mg2+-ATPase activities and also to the square of muscle fiber stiffness. If the two myosin heads in the glycerinated fibers are assumed to react independently with p-PDM, the above results strongly suggest that each myosin molecule in the thick filaments can generate force only when its two heads do not react with p-PDM, muscle fiber stiffness is determined by the total number of native heads, and there is no cooperative interaction between the two myosin heads in catalyzing ATP hydrolysis.

Adenosine Triphosphatases↗

PubMed1985

Fluorescence properties and contraction characteristics of ANM (N-(1-anilinonaphthyl-4)maleimide)-labeled rabbit psoas muscle fibers.

Fluorescence spectra of ANM-labeled, glycerinated rabbit psoas muscle fibers were recorded in relaxed, contracted, and rigor states. SDS polyacrylamide gel electrophoresis of the ANM-labeled muscle fibers indicated that proteins labeled with ANM were myosin heavy chain, C protein, and actin. In a relaxed state in the presence of ATP, myosin heavy chain was mainly labeled. During the transition from rigor to the relaxed or contracted state, there was a blue shift (about 5 nm) of the ANM emission spectrum. Similar experiments with FAM (N-(3-fluoranthyl)-maleimide)-labeled muscle fibers showed that these fluorescence changes were not artifacts due to the movement of muscle fibers. The fibers labeled in the ATP relaxing solution showed a marked decrease in both isometric force and unloaded shortening velocity (Vo), while in the fibers labeled in the rigor solution isometric tension was not markedly suppressed, though Vo decreased to the same extent as in the fibers labeled in the ATP relaxing solution. Fluorescence spectra of ANM-labeled HMM in different states were also measured. A fluorescence enhancement and a blue shift (about 5 nm) of the emission maximum were observed in HMM + MgATP or HMM + MgATP + F-actin in comparison with HMM + F-actin. These results suggest that the fluorescence spectra of the ANM-labeled muscle fibers reflect their conformational changes between the rigor state (in the absence of MgATP) and the relaxed or contracted state (in the presence of MgATP).

Actomyosin↗

PubMed1984

Substrate-concentration dependence of contraction parameters in glycerinated insect flight muscle fibers from Lethocerus derollei.

The contraction characteristics of the dorsal longitudinal muscle of Lethocerus derollei were investigated by applying small sinusoidal length changes (+/- 1% of resting length) to glycerinated muscle bundles and studying the effect of varying the frequency from 0.1 to 10 Hz and the concentration of MgATP from 35 microM to 2.3 mM. The maximum work done by the muscle per cycle increased as the MgATP concentration was decreased from 2.3 mM to 52 microM. Between 52 and 35 microM, the maximum work suddenly changed from a positive to a negative value. The optimal frequency for maximal work shifted from low to high values with increase in the MgATP concentration. As the temperature was increased, the optimal work frequency in 2.3 mM MgATP solution shifted to a higher value. As the MgATP concentration was increased, the optimal frequency for maximal power increased. The maximal value of the power was an increasing function of the MgATP concentration, reaching a plateau above 52 microM MgATP. The muscle stiffness was a decreasing function of the MgATP concentration, and above 52 microM MgATP it reached a minimum of about 22% of that in the rigor solution. These results are discussed in relation to the crossbridge kinetics.

Adenosine Triphosphate↗

PubMed1981

Substrate-concentration of dependences of tension, shortening velocity and ATPase activity of glycerinated single muscle fibers.

Tension P0 and ATPase activity J0 of glycerinated single muscle fibers under isometric concentration as well as the velocity V0 of unloaded shortening were measured as a function of substrate concentration [S]. The stiffness of fibers with sinusoidal length changes at 1 kHz was used as a qualitative measure of the amount of rigor complex. P0 is an increasing function of [S] at low substrate concentrations and has a broad maximum at about 10-40 micrometers MgATP. In this concentration range, 10-40 micrometers, V0 still has a very small value. Then it increases and finally reaches at a plateau at about 1 mM MgATP. J0 increases as P0 does. However, it reaches at a saturated level at about the same concentration as V0. Either 0.5 mM 8-BrATP or 1 mM PPi was added to the substrate solutions to reduce the amount of rigor complex at low substrate concentrations. The addition of PPi of 8-BrATP decreases P0 dominantly at low concentrations of substrate and shifts the maximum to about 100 micrometers MgATP. 8-BrATP considerably increases V0 at low substrate concentrations while V0 is decreased by added PPi. The temperature coefficients, Q10 values were obtained for P0, J0, and V0. The values are essentially constant, 2.1-2.4, in the cases of P0 and J0, and about the same values were found for V0 at very low substrate concentrations. However, they become about 3.3 in the concentration range from 34 micrometers and 2.3 mM. The P-V relation was obtained at 11 micrometers and 2.3 micrometers MgATP. The normalized P-V relation at 11 micrometers was unchanged when 8-BrATP was added. The results are discussed in connection with the mechanism of actomyosin-ATPase activity as well as that of the elementary cycle of the motive force generation.

Adenosine Triphosphatases↗

PubMed1977

The three-state model for the elementary process of energy conversion in muscle.

A three-state model for the elementary process of energy conversion in striated muscle is analysed; in two of the three states, the crossbridge is attached to an actin site, while the third is a detached state. This model accounts for the mechanical properties of steady shortening and lengthening processes as well as those of isometric and isotonic transient processes in a quantitative way. Moreover, qualitative agreement is obtained for the total energy liberation from muscle. Biochemical properties are also computed for transient processes. Comparisions are made with other models with "three states".

Adenosine Triphosphatases↗