EMD-42793
Aquaporin Z with ALFA tag and bound to nanobody
EMD-42793
Single-particle1.9 Å
Deposition: 13/11/2023Map released: 25/09/2024
Last modified: 16/10/2024
Sample Organism:
Escherichia coli,
Vicugna pacos
Sample: Aquaporin Z with ALFA tag bound to nanobody
Fitted models: 8uy6 (Avg. Q-score: 0.68)
Deposition Authors: Stover L, Bahramimoghaddam H, Wang L, Zhou M, Laganowsky A
Sample: Aquaporin Z with ALFA tag bound to nanobody
Fitted models: 8uy6 (Avg. Q-score: 0.68)
Deposition Authors: Stover L, Bahramimoghaddam H, Wang L, Zhou M, Laganowsky A
Grafting the ALFA tag for structural studies of aquaporin Z.
Stover L,
Bahramimoghaddam H,
Wang L,
Schrecke S,
Yadav GP,
Zhou M,
Laganowsky A
(2024) J Struct Biol X , 9 , 100097 - 100097
(2024) J Struct Biol X , 9 , 100097 - 100097
Abstract:
Aquaporin Z (AqpZ), a bacterial water channel, forms a tetrameric complex and, like many other membrane proteins, activity is regulated by lipids. Various methods have been developed to facilitate structure determination of membrane proteins, such as the use of antibodies. Here, we graft onto AqpZ the ALFA tag (AqpZ-ALFA), an alpha helical epitope, to make use of the high-affinity anti-ALFA nanobody (nB). Native mass spectrometry reveals the AqpZ-ALFA fusion forms a stable, 1:1 complex with nB. Single-particle cryogenic electron microscopy studies reveal the octameric (AqpZ-ALFA)4(nB)4 complex forms a dimeric assembly and the structure was determined to 1.9 Å resolution. Dimerization of the octamer is mediated through stacking of the symmetrically bound nBs. Tube-like density is also observed, revealing a potential cardiolipin binding site. Grafting of the ALFA tag, or other epitope, along with binding and association of nBs to promote larger complexes will have applications in structural studies and protein engineering.
Aquaporin Z (AqpZ), a bacterial water channel, forms a tetrameric complex and, like many other membrane proteins, activity is regulated by lipids. Various methods have been developed to facilitate structure determination of membrane proteins, such as the use of antibodies. Here, we graft onto AqpZ the ALFA tag (AqpZ-ALFA), an alpha helical epitope, to make use of the high-affinity anti-ALFA nanobody (nB). Native mass spectrometry reveals the AqpZ-ALFA fusion forms a stable, 1:1 complex with nB. Single-particle cryogenic electron microscopy studies reveal the octameric (AqpZ-ALFA)4(nB)4 complex forms a dimeric assembly and the structure was determined to 1.9 Å resolution. Dimerization of the octamer is mediated through stacking of the symmetrically bound nBs. Tube-like density is also observed, revealing a potential cardiolipin binding site. Grafting of the ALFA tag, or other epitope, along with binding and association of nBs to promote larger complexes will have applications in structural studies and protein engineering.