title: The effect of temperature on mechanical resistance of the native and intermediate states of I27.
abstract: We investigated the effect of temperature on the mechanical unfolding of I27 from human cardiac titin, employing a custom-built temperature control device for single-molecule atomic force microscopy measurement. A sawtooth pattern was observed in the force curves where each force peak reports on the unfolding of an I27 domain. In early unfolding events, we observed a "hump-like" deviation due to the detachment of beta-strand A from each I27 domain. The force at which the humps appear was approximately 130 pN and showed no temperature dependence, at least in the temperature range of 2 degrees C-30 degrees C. The hump structure was successfully analyzed with a two-state worm-like chain model, and the Gibbs free energy difference of the detachment reaction was estimated to be 11.6 +/- 0.58 kcal/mol and found to be temperature independent. By contrast, upon lowering the temperature, the mean unfolding force from the partly unfolded intermediate state was found to markedly increase and the unfolding force distribution to broaden significantly, suggesting that the distance (x(u)) between the folded and transition states in the energy landscape along the pulling direction is decreased. These results suggest that the local structure of beta-strand A are stabilized by topologically simple local hydrogen-bond network and that the temperature does not affect the detachment reaction thermodynamically and kinetically, whereas the interaction between the beta-strands A' and G, which is a critical region for its mechanical stability, is strongly dependent on the temperature.
authors.affiliation: School of Materials Science, Japan Advanced Institute of Science and Technology, Nomi, Ishikawa, Japan.
authors.forename: Yukinori
authors.initials: Y
authors.lastname: Taniguchi
authors.forename: David J
authors.initials: DJ
authors.lastname: Brockwell
authors.forename: Masaru
authors.initials: M
authors.lastname: Kawakami
mesh_terms: D064211:Connectin; D018625:Microscopy, Atomic Force; D009124:Muscle Proteins / Q000737:chemistry*; D011489:Protein Denaturation; D011494:Protein Kinases / Q000737:chemistry*; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D013314:Stress, Mechanical; D013696:Temperature; D013816:Thermodynamics
chemical_list: D064211:Connectin; D009124:Muscle Proteins; C577285:TTN protein, human; D011494:Protein Kinases
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