Limits...
Mechanistic insights into the recycling machine of the SNARE complex.

Zhao M, Wu S, Zhou Q, Vivona S, Cipriano DJ, Cheng Y, Brunger AT - Nature (2015)

Bottom Line: The 20S supercomplex exhibits broken symmetry, transitioning from six-fold symmetry of the NSF ATPase domains to pseudo four-fold symmetry of the SNARE complex.SNAPs interact with the SNARE complex with an opposite structural twist, suggesting an unwinding mechanism.The interfaces between NSF, SNAPs, and SNAREs exhibit characteristic electrostatic patterns, suggesting how one NSF/SNAP species can act on many different SNARE complexes.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Physiology, Howard Hughes Medical Institute, Stanford University, Stanford, California 94305, USA.

ABSTRACT
Evolutionarily conserved SNARE (soluble N-ethylmaleimide sensitive factor attachment protein receptors) proteins form a complex that drives membrane fusion in eukaryotes. The ATPase NSF (N-ethylmaleimide sensitive factor), together with SNAPs (soluble NSF attachment protein), disassembles the SNARE complex into its protein components, making individual SNAREs available for subsequent rounds of fusion. Here we report structures of ATP- and ADP-bound NSF, and the NSF/SNAP/SNARE (20S) supercomplex determined by single-particle electron cryomicroscopy at near-atomic to sub-nanometre resolution without imposing symmetry. Large, potentially force-generating, conformational differences exist between ATP- and ADP-bound NSF. The 20S supercomplex exhibits broken symmetry, transitioning from six-fold symmetry of the NSF ATPase domains to pseudo four-fold symmetry of the SNARE complex. SNAPs interact with the SNARE complex with an opposite structural twist, suggesting an unwinding mechanism. The interfaces between NSF, SNAPs, and SNAREs exhibit characteristic electrostatic patterns, suggesting how one NSF/SNAP species can act on many different SNARE complexes.

Show MeSH

Related in: MedlinePlus

Structures of ATP- and ADP-bound NSFa, Side-view of ATP-bound NSF. b, Side-view ofADP-bound NSF. The six protomer chains are rainbow colored counterclockwise based on therelative positions of the D1 domains to the D2 ring in the ATP-bound NSF model; the chainwith the closest distance between D1 and D2 domains is named Chain A (red). Nucleotidesare shown as grey surfaces. See Methods for generation and refinement of the atomicmodels. c, A schematic diagram showing the topology of ATPase rings of ATP-and ADP-bound NSF, respectively. D1 rings are colored according to the models shown inpanel a and b.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4320033&req=5

Figure 2: Structures of ATP- and ADP-bound NSFa, Side-view of ATP-bound NSF. b, Side-view ofADP-bound NSF. The six protomer chains are rainbow colored counterclockwise based on therelative positions of the D1 domains to the D2 ring in the ATP-bound NSF model; the chainwith the closest distance between D1 and D2 domains is named Chain A (red). Nucleotidesare shown as grey surfaces. See Methods for generation and refinement of the atomicmodels. c, A schematic diagram showing the topology of ATPase rings of ATP-and ADP-bound NSF, respectively. D1 rings are colored according to the models shown inpanel a and b.

Mentions: Both the ATP- and ADP-bound structures of NSF are organized into three layers: tworings consisting of six D2 domains and six D1 domains, respectively, and a layer of six(four) N domains for ATP (ADP)-bound NSF (Figs. 1c, e,and 2a, b). For ADP-bound NSF, the remaining two Ndomains are flipped along the sides of the ATPase rings with well resolved densitiescompared to the N domains atop the D1 ring, leaving little doubt as regards the identity ofthese two densities (Fig. 1e and Extended Data Figs. 4c, e and 7c).


Mechanistic insights into the recycling machine of the SNARE complex.

Zhao M, Wu S, Zhou Q, Vivona S, Cipriano DJ, Cheng Y, Brunger AT - Nature (2015)

Structures of ATP- and ADP-bound NSFa, Side-view of ATP-bound NSF. b, Side-view ofADP-bound NSF. The six protomer chains are rainbow colored counterclockwise based on therelative positions of the D1 domains to the D2 ring in the ATP-bound NSF model; the chainwith the closest distance between D1 and D2 domains is named Chain A (red). Nucleotidesare shown as grey surfaces. See Methods for generation and refinement of the atomicmodels. c, A schematic diagram showing the topology of ATPase rings of ATP-and ADP-bound NSF, respectively. D1 rings are colored according to the models shown inpanel a and b.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4320033&req=5

Figure 2: Structures of ATP- and ADP-bound NSFa, Side-view of ATP-bound NSF. b, Side-view ofADP-bound NSF. The six protomer chains are rainbow colored counterclockwise based on therelative positions of the D1 domains to the D2 ring in the ATP-bound NSF model; the chainwith the closest distance between D1 and D2 domains is named Chain A (red). Nucleotidesare shown as grey surfaces. See Methods for generation and refinement of the atomicmodels. c, A schematic diagram showing the topology of ATPase rings of ATP-and ADP-bound NSF, respectively. D1 rings are colored according to the models shown inpanel a and b.
Mentions: Both the ATP- and ADP-bound structures of NSF are organized into three layers: tworings consisting of six D2 domains and six D1 domains, respectively, and a layer of six(four) N domains for ATP (ADP)-bound NSF (Figs. 1c, e,and 2a, b). For ADP-bound NSF, the remaining two Ndomains are flipped along the sides of the ATPase rings with well resolved densitiescompared to the N domains atop the D1 ring, leaving little doubt as regards the identity ofthese two densities (Fig. 1e and Extended Data Figs. 4c, e and 7c).

Bottom Line: The 20S supercomplex exhibits broken symmetry, transitioning from six-fold symmetry of the NSF ATPase domains to pseudo four-fold symmetry of the SNARE complex.SNAPs interact with the SNARE complex with an opposite structural twist, suggesting an unwinding mechanism.The interfaces between NSF, SNAPs, and SNAREs exhibit characteristic electrostatic patterns, suggesting how one NSF/SNAP species can act on many different SNARE complexes.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Physiology, Howard Hughes Medical Institute, Stanford University, Stanford, California 94305, USA.

ABSTRACT
Evolutionarily conserved SNARE (soluble N-ethylmaleimide sensitive factor attachment protein receptors) proteins form a complex that drives membrane fusion in eukaryotes. The ATPase NSF (N-ethylmaleimide sensitive factor), together with SNAPs (soluble NSF attachment protein), disassembles the SNARE complex into its protein components, making individual SNAREs available for subsequent rounds of fusion. Here we report structures of ATP- and ADP-bound NSF, and the NSF/SNAP/SNARE (20S) supercomplex determined by single-particle electron cryomicroscopy at near-atomic to sub-nanometre resolution without imposing symmetry. Large, potentially force-generating, conformational differences exist between ATP- and ADP-bound NSF. The 20S supercomplex exhibits broken symmetry, transitioning from six-fold symmetry of the NSF ATPase domains to pseudo four-fold symmetry of the SNARE complex. SNAPs interact with the SNARE complex with an opposite structural twist, suggesting an unwinding mechanism. The interfaces between NSF, SNAPs, and SNAREs exhibit characteristic electrostatic patterns, suggesting how one NSF/SNAP species can act on many different SNARE complexes.

Show MeSH
Related in: MedlinePlus