gpcp  (Jena Bioscience)

Journal: eLife

Article Title: Structural model for differential cap maturation at growing microtubule ends

doi: 10.7554/eLife.50155

Figure Lengend Snippet: GDP-BeF 3 - -MT (blue), GDP-AlF x -MT (red), GMPPCP-MT (salmon), GMPCP-MT (yellow), GMPCPP-MT (orange), GDP-Tx-MT (brown) and GDP-MT (gray). ( A ) Top ; representative image (GMPCPP-MTs) of meridional diffraction displaying the meridional plane from l = 1 (4nm) layer line and related harmonics (l = 2 to 4) for longitudinal metric calculations. Bottom ; meridional intensity patterns, where arrows indicate the 4 nm and 8 nm peaks. The inset shows the best fit of 1 nm band experimental intensities in a Lorentzian normal distribution, highlighting positional differences between all tested MT growing conditions (peaks maxima, arrows). ( B ) Top ; representative image (GMPCPP-MTs) of equatorial diffraction highlighting the equatorial plane (l = 0) for lateral metric calculations. Bottom ; equatorial intensity patterns showing the corresponding Bessel functions from J 01 to J 04 +J N1 . The inset shows the J N1 , calculated as described in M and M, displaying the differences in peak maxima (arrows) that occur under various nucleotide polymerization conditions. The red dash line on ( A ) and ( B ) top images shows planes used for intensity line plotting in q x space and further metric calculations. ( C ) Estimation of the number of PFs per MT and percentage of each subpopulation within the solution from fiber diffraction experiments ( left ) and cryo-EM images ( right ).

Article Snippet: Chemical compound, nucleotide , GMPCP , Jena Bioscience , Jena Bioscience:GpCp NU-414 , .

Techniques: Cryo-EM Sample Prep

Journal: eLife

Article Title: Structural model for differential cap maturation at growing microtubule ends

doi: 10.7554/eLife.50155

Figure Lengend Snippet: GTP-bound state (BeF 3 - , blue), transition state (AlF x , red), expanded state (GMPCPP, GMPCP, GDP-Tx, orange), and GDP-bound state (gray). ( A ) Schematic GTPase related conformational changes within the MT lattice. Tubulin activation upon GTP binding (T A ) induces polymerization. During assembly (MT A ), the formation of lateral contacts favors tubulin straightening, which allows GTP hydrolysis. GTPase activity drives MT through a transitional state (MT T ), where the P i is at the nucleotide-binding site before it is released. Expansion (orange) may be an intermediate transient step between GDP•P i and GDP states, which may facilitates P i release (MT E ) and would be blocked in the presence of taxol. GDP-MT (MT M ) shrinks through a ‘peeling-off’ disassembly in which tubulin reverts to the curved conformation, which is inactive (T I ) in the GDP-bound state. ( B ) MT model illustrating specific lattice features of the GTPase cycle. This mosaic structure shows that: (i) the GTP-bound tip (blue) contains curved PFs/sheets that come together into a straight lattice due to the formation of lateral contacts, (ii) the post-hydrolysis GDP•P i lattice (red) retains overall MT structure, (iii) hypothetically, lattice undergoes an energy-consuming expansion phase (orange) that contributes to P i release, and (iv) in the GDP state (gray) subtle changes on the PF skew distinguish the metastable compact lattice or, (v) lattice reverts into its previous lower energy state (compaction), preventing the structure from returning to the cap architecture.

Article Snippet: Chemical compound, nucleotide , GMPCP , Jena Bioscience , Jena Bioscience:GpCp NU-414 , .

Techniques: Activation Assay, Binding Assay, Activity Assay

Journal: eLife

Article Title: Structural model for differential cap maturation at growing microtubule ends

doi: 10.7554/eLife.50155

Figure Lengend Snippet:

Article Snippet: Chemical compound, nucleotide , GMPCP , Jena Bioscience , Jena Bioscience:GpCp NU-414 , .

Techniques: Purification, Over Expression, Software