Structure of spastin bound to a glutamate-rich peptide implies a hand-over-hand mechanism of substrate translocation.

REL 2020-03-04 2021-01-29 2021-02-04 Structure of spastin bound to a glutamate-rich peptide implies a hand-over-hand mechanism of substrate translocation. 0000-0003-1762-8390 Heidi Linda Schubert University of Utah, Department of Biochemistry 15 N Medical Drive East Salt Lake City Utah United States 84112 0000-0001-6796-7740 Christopher P Hill University of Utah, Department of Biochemistry 15 N Medical Drive East Salt Lake City Utah United States 84112 Han H Schubert HL Purdy MD Yeager M Sundquist WI Hill CP 1.8 10.6019/EMPIAR-10382 EMDB EMD-20327 Han H Schubert HL McCullough J Monroe N Purdy MD Yeager M Sundquist WI Hill CP Structure of spastin bound to a glutamate-rich peptide implies a hand-over-hand mechanism of substrate translocation. Journal of Biological Chemistry J Biol Chem United States 295 2 435 443 2019 English 10.1074/jbc.AC119.009890 31767681

Many members of the AAA+ ATPase family function as hexamers that unfold their protein substrates. These AAA unfoldases include spastin, which plays a critical role in the architecture of eukaryotic cells by driving the remodeling and severing of microtubules, which are cytoskeletal polymers of tubulin subunits. Here, we demonstrate that a human spastin binds weakly to unmodified peptides from the C-terminal segment of human tubulin α1A/B. A peptide comprising alternating glutamate and tyrosine residues binds more tightly, which is consistent with the known importance of glutamylation for spastin microtubule severing activity. A cryo-EM structure of the spastin-peptide complex at 4.2 Å resolution revealed an asymmetric hexamer in which five spastin subunits adopt a helical, spiral staircase configuration that binds the peptide within the central pore, whereas the sixth subunit of the hexamer is displaced from the peptide/substrate, as if transitioning from one end of the helix to the other. This configuration differs from a recently published structure of spastin from Drosophila melanogaster, which forms a six-subunit spiral without a transitioning subunit. Our structure resembles other recently reported AAA unfoldases, including the meiotic clade relative Vps4, and supports a model in which spastin utilizes a hand-over-hand mechanism of tubulin translocation and microtubule remodeling.

Unaligned multi-frame movies of ADPBeFx-bound Spastin /data micrographs - multiframe MRC MRC 1200 49 2 49 SIGNED 16 BIT INTEGER 4096 1.056 4096 1.056

Unaligned multi-frame movies of ADPBeFx-bound Spastin

data/FoilHole_16483990_Data_16480075_16480076_20181118_1720_Fractions.mrc data/FoilHole_16483990_Data_16480075_16480076_20181118_1720_Fractions.mrc data