The Low Angle X-Ray Diffraction Pattern from Skinned Fibers of Rabbit Psoas Muscle: Effect of Changes in [Ca++] and [Orthophosphate

Abstract

Bundles of 3-5 fibers were activated isometrically at different pCa by a temperature jump from 1°C to 12°C using a mechanical apparatus (Linari et al., Biophys. J. 92:2476, 2007) modified to collect the X-ray diffraction pattern. The M3 meridional reflection from the axial repeat of the myosin heads was sampled by X-ray interference between half-sarcomeres. In relaxed fibers at 12°C, the M3 reflection had a major peak at 14.56 nm and a minor peak at 14.37 nm. The ratio of peak intensities (RM3) was 0.43±0.06 and the spacing (SM3) was 14.49±0.01 nm. In relaxed fibers the intensity of the main peak reduced with increasing temperature, so that at 36°C (the physiological temperature) the 14.37 nm peak was dominant, with small satellite peaks on either side, as in resting intact fibers from frog muscle. During activation at 12°C at saturating [Ca++], pCa 4.5, the intensity of the M3 reflection (IM3) increased to 1.9±0.4 times the relaxed value with major and minor peaks at 14.68 nm and 14.46 nm; RM3 was 0.62±0.03 and SM3 was 14.59±0.01 nm. Activation at pCa 5.5 or at pCa 4.5 with addition of 10 mM orthophosphate (Pi) had similar effects: force was reduced to 0.34±0.10 the control value and IM3 to 0.56±0.03; RM3 was 0.46±0.07 and SM3 was 14.55±0.02 nm. These results give structural support to the conclusion from mechanical experiments (Linari et al., 2007) that both decreasing [Ca++] and increasing [Pi] reduce isometric force by a decrease in the number of force generating myosin heads with no change in force per head.Supported by FIRB-Futuro in Ricerca and Ente Cassa di Risparmio di Firenze (Italy), MRC (UK).