Mechanism of actin polymerization revealed by cryo-EM structures of actin filaments with three different bound nucleotides

Proc Natl Acad Sci U S A. 2019 Mar 5;116(10):4265-4274. doi: 10.1073/pnas.1807028115. Epub 2019 Feb 13.

Abstract

We used cryo-electron microscopy (cryo-EM) to reconstruct actin filaments with bound AMPPNP (β,γ-imidoadenosine 5'-triphosphate, an ATP analog, resolution 3.1 Å), ADP-Pi (ADP with inorganic phosphate, resolution 3.1 Å), or ADP (resolution 3.6 Å). Subunits in the three filaments have similar backbone conformations, so assembly rather than ATP hydrolysis or phosphate dissociation is responsible for their flattened conformation in filaments. Polymerization increases the rate of ATP hydrolysis by changing the positions of the side chains of Q137 and H161 in the active site. Flattening during assembly also promotes interactions along both the long-pitch and short-pitch helices. In particular, conformational changes in subdomain 3 open up multiple favorable interactions with the DNase-I binding loop in subdomain 2 of the adjacent subunit. Subunits at the barbed end of the filament are likely to be in this favorable conformation, while monomers are not. This difference explains why filaments grow faster at the barbed end than the pointed end. When phosphate dissociates from ADP-Pi-actin through a backdoor channel, the conformation of the C terminus changes so it distorts the DNase binding loop, which allows cofilin binding, and a network of interactions among S14, H73, G74, N111, R177, and G158 rearranges to open the phosphate release site.

Keywords: ATPase; actin; cryo-electron microscopy; polymerization.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Actin Cytoskeleton / chemistry*
  • Actin Cytoskeleton / ultrastructure
  • Actins / chemistry*
  • Adenylyl Imidodiphosphate / chemistry*
  • Animals
  • Avian Proteins / chemistry*
  • Catalytic Domain
  • Chickens
  • Cryoelectron Microscopy
  • Polymerization*

Substances

  • Actins
  • Avian Proteins
  • Adenylyl Imidodiphosphate

Associated data

  • PDB/6DJM
  • PDB/6DJN
  • PDB/6DJO