Cryo-EM analysis of a domain antibody bound rotary ATPase complex

Roberta B Davies; Callum Smits; Andrew S W Wong; Daniela Stock; Mary Christie; Sara Sandin; Alastair G Stewart

doi:10.1016/j.jsb.2017.01.002

Cryo-EM analysis of a domain antibody bound rotary ATPase complex

J Struct Biol. 2017 Mar;197(3):350-353. doi: 10.1016/j.jsb.2017.01.002. Epub 2017 Jan 20.

Authors

Roberta B Davies¹, Callum Smits¹, Andrew S W Wong², Daniela Stock³, Mary Christie³, Sara Sandin⁴, Alastair G Stewart⁵

Affiliations

¹ Molecular, Structural and Computational Biology Division, The Victor Chang Cardiac Research Institute, Darlinghurst, NSW 2010, Australia.
² NTU Institute of Structural Biology, Nanyang Technological University, 636921, Singapore.
³ Molecular, Structural and Computational Biology Division, The Victor Chang Cardiac Research Institute, Darlinghurst, NSW 2010, Australia; Faculty of Medicine, The University of New South Wales, Sydney, NSW 2052, Australia.
⁴ NTU Institute of Structural Biology, Nanyang Technological University, 636921, Singapore; School of Biological Sciences, Nanyang Technological University, 637551, Singapore.
⁵ Molecular, Structural and Computational Biology Division, The Victor Chang Cardiac Research Institute, Darlinghurst, NSW 2010, Australia; Faculty of Medicine, The University of New South Wales, Sydney, NSW 2052, Australia. Electronic address: [email protected].

PMID: 28115258
DOI: 10.1016/j.jsb.2017.01.002

Abstract

The bacterial A/V-type ATPase/synthase rotary motor couples ATP hydrolysis/synthesis with proton translocation across biological membranes. The A/V-type ATPase/synthase from Thermus thermophilus has been extensively studied both structurally and functionally for many years. Here we provide an 8.7Å resolution cryo-electron microscopy 3D reconstruction of this complex bound to single-domain antibody fragments, small monomeric antibodies containing just the variable heavy domain. Docking of known structures into the density revealed the molecular orientation of the domain antibodies, suggesting that structure determination of co-domain antibody:protein complexes could be a useful avenue for unstable or smaller proteins. Although previous studies suggested that the presence of fluoroaluminate in this complex could change the rotary state of this enzyme, we observed no gross structural rearrangements under these conditions.

Keywords: ATPase; Antibodies; Cryo-EM; Membrane protein.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Adenosine Triphosphatases / chemistry
Adenosine Triphosphatases / metabolism*
Antibodies / metabolism*
Cryoelectron Microscopy / methods*
Membrane Proteins / metabolism
Protein Structure, Secondary
Thermus thermophilus / enzymology

Substances

Antibodies
Membrane Proteins
Adenosine Triphosphatases