Forced unfolding of fibronectin type 3 modules: An analysis by biased molecular dynamics simulations

Titin, an important constituent of vertebrate muscles, is a protein of the order of a micrometer in length in the folded state. Atomic force microscop y and laser tweezer experiments have been used to stretch titin molecules t o more than ten times their folded lengths. To explain the observed relatio n between force and extension, it has been suggested that the immunoglobuli n and fibronectin domains unfold one at a time in an all-or-none fashion. W e use molecular dynamics simulations to study the forced unfolding of two d ifferent fibronectin type 3 domains (the ninth, (9)Fn3, and the tenth, (10) Fn3, from human fibronectin) and of their heterodimer of known structure. A n external biasing potential on the N to C distance is employed and the pro tein is treated in the polar hydrogen representation with an implicit solva tion model. The latter provides an adiabatic solvent response, which is imp ortant for the nanosecond unfolding simulation method used here. A series o f simulations is performed for each system to obtain meaningful results. Th e two different fibronectin domains are shown to unfold in the same way alo ng two possible pathways. These involve the partial separation of the "beta -sandwich", an essential structural element, and the unfolding of the indiv idual sheets in a stepwise fashion. The biasing potential results are confi rmed by constant force unfolding simulations. For the two connected domains , there is complete unfolding of one domain ((9)Fn3) before major unfolding of the second domain ((10)Fn3). Comparison of different models for the pot ential energy function demonstrates that the dominant cohesive element in b oth proteins is due to the attractive van der Waals interactions; electrost atic interactions play a structural role but appear to make only a small co ntribution to the stabilization of the domains, in agreement with other stu dies of beta-sheet stability. The unfolding forces found in the simulations are of the order of those observed experimentally, even though the speed o f the former is more than six orders of magnitude greater than that used in the latter. (C) 1999 Academic Press.