X-ray diffraction studies of the structural state of crossbridges in skinned frog sartorius muscle at low ionic strength

Abstract

Low-angle X-ray diffraction diagrams were obtained from chemically skinned frog sartorius muscles under low ionic strength relaxing conditions. Experiments on single muscle fibres from rabbit muscle and on muscle proteins in solution have suggested the presence of a ‘low ionic strength attached state’ of the myosin crossbridges to actin, in which the overall ATP splitting and force-generating cycle is still blocked. This opened up the possibility that structural information about one of the intermediate states in the crossbridge cycle might be obtained under these conditions. Using synchrotron radiation as a high intensity X-ray source we were able to record the appropriate diffraction diagrams with short exposure times and were able to compare the same muscles at normal and at low ionic strength. Changes in the intensities of the equatorial reflections are consistent with the existence of some actin-attached crossbridges at low ionic strength in a relaxing medium and an increase in the 143 Å meridional intensity can be interpreted in a similar way. However, these attached bridges do not give rise to changes in the actin-based layer line reflections, nor is their presence associated with a weakening of the myosin layer line pattern. These results provide further evidence for the existence of bound states of crossbridges, in which their orientation relative to actin is not sharply defined.