MOLECULAR BASIS OF TITIN ELASTICITY

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DESCRIPTION (Adapted from abstract): Passive elastic properties play a critical role in muscle physiology. They determine the filling volume of a ventricle during a heartbeat and the amount of mechanical energy stored during the flap of a wing. Titin is a rod-like modular protein that stretches between the M and Z lines of a sarcomere. The region of titin that overlaps with the sarcomeric I band is responsible for the passive elastic properties of the sarcomere. However, the molecular mechanisms that allow titin to extend under force are not known. I band titin is composed of tandem repeats of immunoglobulin (Ig) like domains and a polypeptide region rich in amino acids, P,E,V, and K (the PEVK region). It has been proposed that the elastic properties of I band titin result from the unfolding/refolding reactions of its individual modules and the elasticity of the PEVK region. The main objective of this proposal is to combine biophysical and molecular biological techniques to examine the molecular basis of the elasticity of the I band titin from human cardiac muscle. Towards this aim the investigators will exploit recently developed atomic force microscopy (AFM) techniques that are capable of measuring the mechanical properties of single proteins. They will use AFM to measure the mechanical stability and kinetics of individual I band titin modules, and will use mutagenesis of engineered tandem repeats of these modules to examine the determinants of unfolding and refolding. Tandem modular proteins will be engineered composed of mixtures of modules, to examine the elasticity that results from combining mechanical units of different mechanical stability and kinetics. The measurements will examine the conformational changes that allow titin modules to reversibly extend under an applied force. They studies are aimed at elucidating the mechanical architecture of a modular elastic protein and the molecular basis of the extensibility of Ig and FNIII modules. The mechanical properties of these modules may also find wider significance since they are typical building blocks of a large number of other proteins.