Macromolecular Crowding Modulates Cross-Bridge Performance

Abstract

Cross-bridge kinetics is usually studied in diluted solution that does not reflect in vivo conditions of cell cytoplasm. In cells, myosin and actin work in dense environment of small and large macromolecules, affecting cross-bridge kinetics. We used high molecular weight sucrose polymer Ficoll70 to examine the effect of crowding on kinetics of acto-myosin interaction and cross-bridge cycle. Dictyostelium discoideum myosin II S1 and rabbit skeletal actin were used in the study. We observed two-fold increase of the rate of acto-myosin cycle in the presence of 25% w/v Ficoll70 in the reaction solution. Transient kinetics studies showed that virtually every kinetic step of the cross-bridge cycle was affected by Ficoll70. Ficoll70 decreases ADP affinity to acto-myosin, and therefore increases the overall rate of the cycle, because ADP release is the rate limiting step in D.discoideum myosin II actin activated ATPase activity. We conclude that macromolecular crowding modulates cross-bridge performance in cells.