Multiscale Computational Modeling of Tubulin-Tubulin Lateral Interaction

Abstract

Microtubules are multistranded polymers in eukaryotic cells that support key cellular functions such as chromosome segregation, motor-based cargo transport, and maintenance of cell polarity. Microtubules self-assemble via ''dynamic instability,'' in which the dynamic plus ends switch stochastically between alternating phases of polymerization and depolymerization. A key question in the field is what are the atomistic origins of this switching, i.e., what is different between the GTP-and GDP-tubulin states that enables microtubule growth and shortening, respectively? More generally, a major challenge in biology is how to connect theoretical frameworks across length-and timescales, from atoms to cellular behavior. In this study, we describe a multiscale model by linking atomistic molecular dynamics (MD), molecular Brownian dynamics (BD), and cellularlevel thermokinetic modeling of microtubules. Here, we investigated the underlying interaction energy when tubulin dimers associate laterally by performing all-atom MD simulations. We found that the lateral potential energy is not significantly different among three nucleotide states of tubulin, GTP, GDP, and GMPCPP and is estimated to be y À11 k B T. Furthermore, using MD potential energy in our BD simulations of tubulin dimers confirms that the lateral bond is weak on its own, with a mean lifetime of $0.1 ms, implying that the longitudinal bond is required for microtubule assembly. We conclude that nucleotide-dependent lateral-bond strength is not the key mediator microtubule dynamic instability, implying that GTP acts elsewhere to exert its stabilizing influence on microtubule polymer. Furthermore, the estimated lateral-bond strength (DG 0 lat y À5 k B T) is well-aligned with earlier estimates based on thermokinetic modeling and light microscopy measurements. Thus, we have computationally connected atomistic-level structural information, obtained by cryo-electron microscopy, to cellular-scale microtubule assembly dynamics using a combination of MD, BD, and thermokinetic models to bridge from Å ngstroms to micrometers and from femtoseconds to minutes.

Poisson poisoning as the mechanism of action of the microtubule-targeting agent colchicine

“…To test this hypothesis, we measured the intradimer bending angles of the dimers (Fig. 8G, and S1E), decomposed in radial (θx), tangential (θy) and twist (θz) bending directions (65). Both dimers in solution moved toward a more bent conformation, as predicted by previous MD studies, in both radial and tangential directions with GTP-and GDP-TC complexes being more confined in bending motions compared to the unliganded tubulin dimer.…”

“…Hence, in contrast to vinblastine (57), colchicine does not promote oligomerization, consistent with previous studies (64,96), and did not strengthen the longitudinal bond. Note that lateral oligomers cannot be detected in the solution due to the lateral bond being weak with a short lifetime (65,97). These results altogether provide evidence that colchicine stabilizes dynamics by strengthening the lateral bond in the MT thereby slowing off-rates of tubulin bound to colchicine in the MT lattice.…”

“…A conformational change can also be reflected as a change in bending preference of the TC complex relative to the free tubulin. Based on previous MD studies (65,98,99), free tubulin conformation is bent and moves toward a more bent conformation if simulated long enough. To test this hypothesis, we measured the intradimer bending angles of the dimers (Fig.…”

“…Rootmean-square deviation (RMSD) and root-mean-square fluctuation (RMSF) values were calculated based on tcl scripts written for VMD. BSASA and bending angles were calculated based on the methodology previously described (65).…”

“…Finally, we used the same method to determine the likelihood that colchicine is able to reach the active site of tubulin via transport along the identified channels. The last 200ns of the MD simulation trajectories for TC complex and unliganded tubulin in GDP-and GTP-states, along with 200ns trajectory of straight tubulin in solution in both nucleotide states from our previous study (65), were used for this analysis.…”