Kif2C Minimal Functional Domain Has Unusual Nucleotide Binding Properties That Are Adapted to Microtubule Depolymerization

Abstract

Background: Kinesin-13 proteins, such as Kif2C, share a conserved motor domain with motile kinesins, but they depolymerize microtubules. Results: Kif2C is mostly ATP-bound, and Kif2C-ATP has a strong binding preference for curved tubulin. Conclusion: Kif2C-ATP starts depolymerization by favoring a curved tubulin conformation at microtubule ends. Significance: Kinesin-13 proteins have evolved unique nucleotide binding properties to fulfill their microtubule disassembly activity.

Prime movers: the mechanochemistry of mitotic kinesins

“…showing that the Mg•ADP form of KIF2C binds more tightly to microtubule tips than to free tubulin (whereas the Mg•ATP form prefers free tubulin 177 ). Recycling of the KIF2 enzyme occurs via ADP release, which only takes a few seconds even in the absence of tubulin or microtubules 25 .…”

A designed ankyrin repeat protein selected to bind to tubulin caps the microtubule plus end

“…The N3C DARPin library (40) was used to select tubulin binders, using four rounds of ribosome display (18,19). In these experiments, the target (tubulin) was covalently coupled via a disulphide bond to a biotinylated stathmin-derived peptide (41) and immobilized via neutravidin or streptavidin coated on microtiter plates. Details of the selection are described in SI Methods.…”

Structural Insights into the Unusually Strong ATPase Activity of the AAA Domain of the Caenorhabditis elegans Fidgetin-like 1 (FIGL-1) Protein

“…ATPase Activity Assay-The steady-state ATPase activity of 1 M CeFIGL-1-AAA was measured using an enzyme-coupled assay in 50 mM Tris-HCl (pH 8.2), 75 mM NaCl, 1 mM DTT, 2 mM MgCl 2 , 1 mM EGTA, and various concentrations of ATP at 25°C as described previously (22). The turnover numbers per subunit were calculated.…”