Tubulin nucleation and microtubule (MT) assembly are frequent events in cells. To understand the fundamental mechanisms of MT assembly it is critical to resolve the early stages of the reaction, dictating the assembly pathway. Despite years of research, tubulin nucleation is still one of the least understood steps of MT dynamics, owing to the size of tubulin and its highly dynamic character. To determine the fundamental molecular-level mechanisms directing tubulin nucleation and the early assembly states, we will reconstitute in vitro a minimal model system that mimics the key elements of cytoskeleton MT assembly. We will use time-resolved solution small angle-X-ray scattering (TR-SAXS) and our advanced analysis tools to follow in real-time, the active self-organization, dynamic structures, and bioenergetics of tubulin proteins and their regulation by fluxes of chemical energy, associated with nucleotide hydrolysis reactions.