Article Information

• PDF (784 kb)

PubMed

• Articles by B. Brenner
• Articles by L.C. Yu

Related Articles

• A Model of Cross-Bridge Attachment to Actin in the A·M·ATP S...

  A Model of Cross-Bridge Attachment to Actin in the A·M·ATP State Based on X-Ray Diffraction from Permeabilized Rabbit Psoas Muscle Biophysical Journal, Volume 82, Issue 4, 1 April 2002, Pages 2123-2133 Jin Gu, Sengen Xu and Leepo C. Yu Abstract A model of cross-bridges binding to actin in the weak binding A·M·ATP state is presented. The modeling was based on the x-ray diffraction patterns from the relaxed skinned rabbit psoas muscle fibers where ATP hydrolysis was inhibited by N-phenylmaleimide treatment (S. Xu, J. Gu, G. Melvin, L. C. Yu. 2002. 82:2111–2122). Calculations included both the myosin filaments and the actin filaments of the muscle cells, and the binding to actin was assumed to be single headed. To achieve a good fit, considerable flexibility in the orientation of the myosin head and the position of the S1-S2 junction is necessary, such that the myosin head can bind to a nearby actin whereas the tail end was kept in the proximity of the helical track of the myosin filament. Hence, the best-fit model shows that the head binds to actin in a wide range of orientations, and the tail end deviates substantially from its lattice position in the radial direction (∼60Å). Surprisingly, the best fit model reveals that the detached head, whose location thus far has remained undetected, seems to be located close to the surface of the myosin filament. Another significant requirement of the best-fit model is that the binding site on actin is near the N terminus of the actin subunit, a position distinct from the putative rigor-binding site. The results support the idea that the essential role played by the weak binding states M·ATP ↔ A·M·ATP for force generation lies in its flexibility, because the probability of attachment is greatly increased, compared with the weak binding M·ADP·P ↔ A·M·ADP·P states. Abstract | Full Text | PDF (455 kb)

• Structural Features of Cross-Bridges in Isometrically Contra...

  Structural Features of Cross-Bridges in Isometrically Contracting Skeletal Muscle Biophysical Journal, Volume 82, Issue 5, 1 May 2002, Pages 2536-2547 Theresia Kraft, Thomas Mattei, Ante Radocaj, Birgit Piep, Christoph Nocula, Markus Furch and Bernhard Brenner Abstract Two-dimensional x-ray diffraction was used to investigate structural features of cross-bridges that generate force in isometrically contracting skeletal muscle. Diffraction patterns were recorded from arrays of single, chemically skinned rabbit psoas muscle fibers during isometric force generation, under relaxation, and in rigor. In isometric contraction, a rather prominent intensification of the actin layer lines at 5.9 and 5.1nm and of the first actin layer line at 37nm was found compared with those under relaxing conditions. Surprisingly, during isometric contraction, the intensity profile of the 5.9-nm actin layer line was shifted toward the meridian, but the resulting intensity profile was different from that observed in rigor. We particularly addressed the question whether the differences seen between rigor and active contraction might be due to a rigor-like configuration of both myosin heads in the absence of nucleotide (rigor), whereas during active contraction only one head of each myosin molecule is in a rigor-like configuration and the second head is weakly bound. To investigate this question, we created different mixtures of weak binding myosin heads and rigor-like actomyosin complexes by titrating MgATPS at saturating [Ca] into arrays of single muscle fibers. The resulting diffraction patterns were different in several respects from patterns recorded under isometric contraction, particularly in the intensity distribution along the 5.9-nm actin layer line. This result indicates that cross-bridges present during isometric force generation are not simply a mixture of weakly bound and single-headed rigor-like complexes but are rather distinctly different from the rigor-like cross-bridge. Experiments with myosin-S1 and truncated S1 (motor domain) support the idea that for a force generating cross-bridge, disorder due to elastic distortion might involve a larger part of the myosin head than for a nucleotide free, rigor cross-bridge. Abstract | Full Text | PDF (387 kb)

• The Effect of Thin Filament Activation on the Attachment of ...

  The Effect of Thin Filament Activation on the Attachment of Weak Binding Cross-Bridges: A Two-Dimensional X-Ray Diffraction Study on Single Muscle Fibers Biophysical Journal, Volume 76, Issue 3, 1 March 1999, Pages 1494-1513 T. Kraft, S. Xu, B. Brenner and L.C. Yu Abstract To study possible structural changes in weak cross-bridge attachment to actin upon activation of the thin filament, two-dimensional (2D) x-ray diffraction patterns of skinned fibers from rabbit psoas muscle were recorded at low and high calcium concentration in the presence of saturating concentrations of MgATPS, a nucleotide analog for weak binding states. We also studied 2D x-ray diffraction patterns recorded under relaxing conditions at an ionic strength above and below 50mM, because it had been proposed from solution studies that reducing ionic strength below 50mM also induces activation of the thin filament. For this project a novel preparation had to be established that allows recording of 2D x-ray diffraction patterns from single muscle fibers instead of natural fiber bundles. This was required to minimize substrate depletion or product accumulation within the fibers. When the calcium concentration was raised, the diffraction patterns recorded with MgATPS revealed small changes in meridional reflections and layer line intensities that could be attributed in part to the effects of calcium binding to the thin filament (increase in , decrease in first actin layer line intensity, increase in ) and in part to small structural changes of weakly attached cross-bridges (e.g., increase in and ). Calcium-induced small-scale structural rearrangements of cross-bridges weakly attached to actin in the presence of MgATPS are consistent with our previous observation of reduced rate constants for attachment and detachment of cross-bridges with MgATPS at high calcium. Yet, no evidence was found that weakly attached cross-bridges change their mode of attachment toward a stereospecific conformation when the actin filament is activated by adding calcium. Similarly, reducing ionic strength to less than 50mM does not induce a transition from nonstereospecific to stereospecific attachment. Abstract | Full Text | PDF (600 kb)

• …more

Abstract

Radial equilibrium lengths of the weakly attached, force-generating, and rigor cross-bridges are determined by recording their resistance to osmotic compression. Radial equilibrium length is the surface-to-surface distance between myosin and actin filaments at which attached cross-bridges are, on average, radially undistorted. We previously proposed that differences in the radial equilibrium length represent differences in the structure of the actomyosin cross-bridge. Until now the radial equilibrium length had only been determined for various strongly attached cross-bridge states and was found to be distinct for each state examined. In the present work, we demonstrate that weakly attached cross-bridges, in spite of their low affinity for actin, also exert elastic forces opposing osmotic compression, and they are characterized by a distinct radial equilibrium length (12.0 nm vs. 10.5 nm for force-generating and 13.0 nm for rigor cross-bridge). This suggests significant differences in the molecular structure of the attached cross-bridges under these conditions, e.g., differences in the shape of the myosin head or in the docking of the myosin to actin. Thus, the present finding supports our earlier conclusion that there is a structural change in the attached cross-bridge associated with the transition from a weakly bound configuration to the force-generating configuration. The implications for imposing spatial constraints on modeling actomyosin interaction in the filament lattice are discussed.