| MWexperimental | 259 | kDa |
| MWexpected | 266 | kDa |
| VPorod | 374 | nm3 |
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log I(s)
2.94×10-2
2.94×10-3
2.94×10-4
2.94×10-5
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s, nm-1
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Ternary complex of Kif2A-bound tandem tubulin heterodimers represents a kinesin-13-mediated microtubule depolymerization reaction intermediate.
Trofimova D, Paydar M, Zara A, Talje L, Kwok BH, Allingham JS, Nat Commun 9(1):2628 (2018) Europe PMCSASDCR9 – Kif2A-tubulin-DARP complex in the presence of AMP-PNP
Designed Ankyrin Repeat Protein D1Kinesin-like protein KIF2A
Tubulin alpha-1B chain
Tubulin beta-2B chain
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Fits and models
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Synchrotron SAXS data from solutions of Kif2A-tubulin-DARPin complex in presence of AMP-PNP in 20 mM HEPES, 1 mM MgCl2, 150 mM NaCl, pH 7.2 were collected at the BioSAXS G1 beam line at the Cornell High Energy Synchrotron Source (CHESS; Ithaca, NY, USA) using a CCD Finger Lakes CCD detector at a sample-detector distance of 1.5 m and at a wavelength of λ = 0.1267 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). 1200 successive 2 second frames were collected using size exclusion chromatography SAXS (SEC-SAXS; 3.70 mg/ml sample injection at 22°C).
For the SEC-SAXS analysis, a mixture of Kif2A, tubulin and DARPin was passed continuously through an X-ray sample cell via an in-line size exclusion column (GE Superdex 200 10/300 GL) at a flow rate of 1 ml/min. The column was pre-equilibrated with running buffer consisting of 20 mM HEPES, 150 mM NaCl, 1 mM MgCl2 pH 7.2. The protein sample was prepared at 25 μM tubulin, 27 μM DARP in and 20 μM Kif2A and injected into a 100 μL loop. Approximately 1200 two-second exposures were collected per sample, and 100 buffer profiles preceding the elution peaks were averaged and used for background subtraction. |
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