Stales and transitions during forced unfolding of a single spectrin repeat

Spectrin is a vital and abundant protein of the cytoskeleton. It has an elo ngated structure that is made by a chain of so-called spectrin repeats. Eac h repeat contains three antiparallel alpha-helices that form a coiled-coil structure. Spectrin forms an oligomeric structure that is able to cross-lin k actin filaments. In red cells, the spectrin/actin meshwork underlying cel l membrane is thought to be responsible for special elastic properties of t he cell. In order to determine mechanical unfolding properties of the spect rin repeat, rye have used single molecule force spectroscopy to study the s tates of unfolding of an engineered polymeric protein consisting of identic al spectrin domains. We demonstrate that the unfolding of spectrin domains can occur in a stepwise fashion during stretching. The force-extension patt erns exhibit features that are compatible with the existence of at least on e intermediate between the folded and the completely unfolded conformation. Only those polypeptides that still contain multiple intact repeats display intermediates, indicating a stabilisation effect. Precise force spectrosco py measurements on single molecules using engineered protein constructs rev eal states and transitions during the mechanical unfolding of spectrin. Sin gle molecule force spectroscopy appears to open a new window for the analys is of transition probabilities between different conformational states, (C) 2000 Federation of European Biochemical Societies. Published by Elsevier S cience B.V. All rights reserved.