Structural Biology

Biography

Biography: John H. Miller

Abstract

    Tubulin    , a 50 kDa protein, associates into α, β-heterodimers which polymerize in to the    microtubule   , a maj or cytoskeletal component of all eukaryote cells. Micr otubules consist of 13 protofilaments with the α,β-dimers stacked head-to-tail. Numerous molecules, both    endogenous    and   exogenous  , bind to tubulin and affect it s ability to  polymerize  and  depolymer ize . Insights into the s   tructure of tubulin and the binding sites of different  ligands  were first obtained from       electron crystallography       of Zn- and paclitaxel-stabilize d, antiparallel sheets.      Paclitaxel      is the first known micr otubule-stabilizing agent. More recent studies have used a      T2R complex      consisting of a        stathmin-like domain      bound to two     heterodimers     linked in a head-to- tail fashion. Stathmin, by inducing a     curve    d dimer structure,  prevents its assembly into polymers. This    T2R complex   , often in combination with the enzyme tubu lin tyrosine ligase has allowed detailed structural mapping by    X- ray crystallography    to less than 2.3 Å resolution. There are two known   microtubule-destabilizing sites   on  β-tubulin , the colchicine site and the  vinca alkaloid site, and two sta   bilizing sites, the taxoid site and the laulimalide/peloruside site. Recent    X-ray crystallography    analysis has confirmed the location of the stabilizing sites and shown that ligands that bind the two different sites can bind simultaneously . Adding both types of ligands together can lead to synergistic effects on the dynamicity of the   microtubules  , both in solution and inside cells; however, the mechanisms of stabilization and synergy between  ligands are not fully understood. In collaboration with Eva Nogales at Berkeley, high resolution   cryo-EM   studies are currently underway on the structural association between  peloruside  and tubulin.