Because microtubules are the structural elements of cells, it is essential to study the mechanical properties of single microtubules under physiological conditions. Previously, we measured the effect of temperature on the flexural rigidity of a single taxol-stabilized microtubule and found that the flexural rigidity is 2.5 x 10(-24) Nm(2), independent of temperature in the 20-35 degrees C range. Employing the same technique here, we have measured the flexural rigidity of microtubules polymerized in the presence of guanylyl-(a,b)-methylene-diphosphonate (GMPCPP, the slowly hydrolyzable GTP analogue) and in the presence of GTP only; both of the states were taxol-free. The obtained values were approximately 5-fold (for GMPCPP) and three- to 4-fold (for GTP) greater than those of taxol-stabilized microtubules. Interestingly, rigidity decreased as temperature increased, that is, temperature dependence was only observed in taxol-free microtubules. Length dependence was also observed. These results indicate that the transition of microtubule's rigidity is associated with the tubulin conformation change from a GTP-bound state to a GDP-bound state in the alpha/beta subunit. We discuss the relationship of the regulation mechanism of the microtubules in the cell body to the changes in rigidity through hydrolysis. (C) 2010 Elsevier Inc. All rights reserved.