Researchers of Sechenov University together with their colleagues from several Russian institutes studied the dynamics of microtubules that form the basis of the cytoskeleton and take part in the transfer of particles within a cell and its division. The computer model they developed describes the mechanical properties of protofilaments (longitudinal fibers that compose microtubules) and suggests how they assemble and disassemble. All the details of the study can be found in PLOS Computational Biology.
Microtubules are long hollow cylinders with walls consisting of tubulin molecules arranged helix-wise. Each cycle contains 13 pairs of α- and β-tubulin, so a microtubule is composed of 13 longitudinal fibers, protofilaments. Microtubules grow by addition of tubulin from the cytoplasm. The protein binds more actively to one end (plus-end) of the microtubule and dissociates from the other one (minus-end) quicker. Both processes take place simultaneously, but their rate changes: when the concentration of tubulin is sufficient, microtubules grow faster than degrade, and when the concentration is low - vice versa. Between the phases of growth and shrinking there is a period of stability, but it is very short.
Though the mechanisms of microtubule's growth and shortening are well-studied, there are still quite a few questions about how the structure and properties of tubulin change. It is known that both molecules of tubulin (α- and β-tubulin) are connected to a molecule of guanosine triphosphate (GTP). GTP of β-tubulin can hydrolyse and turn into guanosine diphosphate (GDP) that causes the double tubulin molecule (dimer) to dissociate from a microtubule. The authors of the paper tried to understand how the properties of tubulin dimers and protofilaments depend on GTP hydrolysis and what provides the difference between plus- and minus-ends of microtubules. Above all, microtubules take part in cell division, and studies of these mechanisms will contribute to the search for yet unknown ways to suppress the replication of cancer cells. In particular, microtubules serve as molecular targets for an important anti-tumour drug, paclitaxel, that inhibits microtubule disassembly.
Existing studies offer several models of possible changes in protein structure taking place upon GTP hydrolysis: a slight curving of tubulin dimers or weakening of longitudinal bonds between dimers without significant changes in their shape. Some researchers also suggest that hydrolysis may affect interactions between neighbouring protofilaments. According to the authors, it was impossible to prove or refute any of these claims for a long time because of the lack of precise experimental data. In this research they verified the first hypothesis and computed the 'behaviour' of molecules using the latest of available experimental structures obtained by cryo-electron tomography. They examined bonds between dimers as well as between α- and β-tubulin within them.
Scientists modelled the bending of the tubulin dimer and the whole protofilament, with GTP and GDP bound to them, throughout one millisecond, watching the angle and direction of the curvature and assessing the strength of bonds within and between dimers. The results showed that protofilaments with GTP and GDP-bound tubulin were bent almost to the same extent, so the first hypothesis was disproved. But it turned out that GTP influences the flexibility of the bonds between dimers: protofilaments made of tubulin connected with GTP were much more bendable compared with those containing GDP.
Using the revealed difference in bond rigidity between GTP and GDP-connected protofilaments, the authors concluded that more flexible bonds ease the straightening of protofilaments and thus facilitate the assembly of the microtubule.
'Based on simulations, we developed a simple model of dynamic instability of microtubules, i.e. their assembly and disassembly. A deeper understanding of this process on the molecular level would enable a targeted development of medicines able to affect the stability of microtubules and thus prevent the reproduction of tumour cells', said Philipp Orekhov, co-author of the paper and senior scientist at the Institute for Personalized Medicine, Sechenov University.
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Journal
PLoS Computational Biology