Determination of the net exchange rate of tubulin dimer in steady-state microtubules by fluorescence correlation spectroscopy

The microtubule cytoskeleton plays an important role in eukaryotic cells, e , g,, in cell movement or morphogenesis. Microtubules, formed by assembly o f tubulin dimers, are dynamic polymers changing randomly between periods of growing and shortening, a property known as dynamic instability. Another p rocess characterizing the dynamic behaviour is the so-called treadmilling d ue to different binding constants of tubulin at both microtubule ends. In t his study, we used tetramethyl rhodamine (TMR)-labeled tubulin added to mic rotubule suspensions to determine the net exchange rate (NER) of tubulin di mers by fluorescence correlation spectroscopy (FGS) as a measure for microt ubule dynamics. This approach, which seems to be suitable as a screening sy stem to detect compounds influencing the NER of tubulin dimers into microtu bules at steady-state, showed that taxol, nocodazole, colchicine, and vinbl astine affect microtubule dynamics at concentrations as low as 10(-9)-10-(1 0) M.