Vinblastine binding site

Bruggemann EP, Currier SJ, Gottesman MM, et al. Characterization of the azido-pine and vinblastine binding site of P-glycoprotein. J Biol Chem 1992 267 (29) 21020-21026. 

Fig. 4 Left, location of the vinblastine and colchicine binding sites in tubulin (1Z2B structure), a- and (3-tubulin are represented as yellow and magenta ribbons, respectively vinblastine and DAMA-colchicine are represented as green and cyan spheres, respectively GTP and GDP are represented as pink sticks. Vinblastine binds at the interdimer interface, whereas colchicine binds at the intradimer interface. Right, zoom on the vinblastine binding site. Secondary structure elements contacting vinblastine are colored blue...

The exact binding site of vinca alkaloids remained unknown until 2005, when the crystal structure of vinblastine bound to tubulin complexed with colchicine and with the stathmin-like domain of RB3 was determined (PDB entry 1Z2B) [3], The structure revealed that vinblastine binds to curved tubulin at the interface between two a/p-tubulin heterodimers (interdimer interface, Fig. 4), introducing a wedge that interferes with tubulin assembly. The vinblastine binding site is defined by loop T7, helix H10 and strand S9 in the a subunit of the first heterodimer, and by helix H6 and loops T5 and H6-H7 in the p subunit of the second heterodimer. In microtubules, this region is located toward the inner lumen and is... 

Fig. 5 Interaction of antimitotic peptides and depsipeptides with the vinca domain with respect to the vinblastine binding site (1Z2B structure). Residues supposed to interact with the peptides are represented as blue sticks, vinblastine is represented as green sticks. Tubulin ribbons are colored as in Fig. 4...

Martin, C., Higgins, C.F., and Callaghan, R. (2001) The vinblastine binding site adopts high- and low-affinity conformations during a transport cyde of P-glycoprotein. Biochemistry, 40 (51), 15733-15742. 

Cryptophycin-52 (26a), a member of the cryptophycin family developed by Eli Lilly and produced by total chemical synthesis [76], was selected from diverse synthetic analogues displaying superior potency, stability and amenability of clinical formulation [77,76]. Cryptophycin-52 in vitro binds non-covalently to tubulin at a single high affinity site, which presumably overlaps with the Vinblastine binding site [75]. Proliferation of diverse tumour cell lines was inhibited with IC50 values of 13 pM to 232 pM, minimally affected by P-gp or MRP overexpression [78]. Data from phase I and II trials showed severe toxicities as long... 

Rai S, Wolff J. Localization of the vinblastine-binding site on p-tubulin. j Biol Chem 1996 271 14707-14711. 

The cause of the cell cycle specificity of the bisindole alkaloids may be associated with the ability of these compounds to interact with the protein tubulin and thereby inhibit the polymerization (and depolymerization) of microtubules (16,17). In this respect the cellular pharmacology of vinca alkaloids is similar to that of other cytotoxic natural products such as colchicine or podophyllotoxin. On closer inspection, however, Wilson determined that the specific binding site on tubulin occupied by vinblastine or vincristine is chemically distinct from the site occupied by the other natural products (18). Subsequent experiments have determined that the maytansinoids, a class of ansa-macrocycles structurally distinct from the bisindoles, may bind to tubulin at an adjacent (or overlapping) site (19). A partial correlation of the antimitotic activity of these compounds with their tubulin binding properties has been made, but discrepancies in cellular uptake probably preclude any quantitative relationship of these effects (20). 

Small molecules modulating microtubule assembly have played major roles as tools in microtubule research, in a manner closely related to their chemotherapeutic interest [1], Tubulin was first purified in the last century as the colchicine-binding protein proposed to be the subunit of cellular microtubules [2], More recently, a colchicine derivative was employed to help crystallization and determine the structure of tubulin by X-ray diffraction [3], The colchicine, vinblastine [4] and paclitaxel [5] sites are main drug binding sites of tubulin, to which many other substances bind. The discovery of microtubule stabilization by paclitaxel [6] prompted its clinical development [7] and a burst of research on new MSAs, as well as the generalized use of paclitaxel or docetaxel as convenient reagents to assemble (see Fig. 1), stabilize or detect microtubules in the laboratory. One example is the development... 

Fig. 4 Diagram of the crystal structure of the T2R complex showing the binding sites of MT-destabilizing drugs (PDB entry 1Z2B) [13]. Protein subunits are represented as ribbons. RB3-SLD is colored orange, a-tubulin is purple, and p-tubulin is green. Small-molecule ligands are represented as spheres (vinblastine orange, colchicine red, GTP yellow, and GDP magenta). Colchicine binds to the p-subunit at the intradimer interface. Vinblastine binds at the interdimer interface...

Specificity of binding is supported by the observation of protein-mediated interligand NOEs [75, 76, 112] between pairs of ligands known to compete for the PTX-binding site (epothilones, taxanes, discodermolide, and baccatins) but not between ligands known to bind to different sites in MT, such as EpoA/vinblastine [76],... 

They can be classified in two main classes (poly)macrocyclic molecules (e.g. vinblastine) and (mostly linear) pseudopeptides (e.g. dolastatin 10). Some ligands (e.g. phomopsin) have characteristics of both classes (Fig. 5). Whether the tubulin binding sites of these widely diverse molecules overlap or not remains to be established. Whatever the case, taken together these sites define the tubulin vinca domain [43]. 

The X-ray crystallographic analysis of T2R crystals soaked with vinblastine has revealed its binding site between the two tubulins of this short protofilament-like curved assembly (Fig. 6) [55]. It is equally contributed by the a-subunit of one tubulin and the [i-subuni I of the other. Vinblastine therefore belongs to the growing class of small molecules ligands known as interfacial inhibitors [58] these... 

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