3AOU

Structure of the Na+ unbound rotor ring modified with N,N f-Dicyclohexylcarbodiimide of the Na+-transporting V-ATPase

Summary for 3AOU

• Entry DOI: 10.2210/pdb3aou/pdb
• Descriptor: V-type sodium ATPase subunit K, DICYCLOHEXYLUREA, UNDECYL-MALTOSIDE, ... (4 entities in total)
• Functional Keywords: sodium ion transport, v-atpase, dccd, membrane rotor ring, hydrolase
• Biological source: Enterococcus hirae
• Cellular location: Cell membrane ; Multi-pass membrane protein : P43457
• Total number of polymer chains: 10
• Total formula weight: 163675.78

Authors

Mizutani, K.,Yamamoto, M.,Yamato, I.,Kakinuma, Y.,Shirouzu, M.,Yokoyama, S.,Iwata, S.,Murata, T. (deposition date: 2010-10-06, release date: 2011-08-17, Last modification date: 2024-10-23)

Primary citation

Mizutani, K.,Yamamoto, M.,Suzuki, K.,Yamato, I.,Kakinuma, Y.,Shirouzu, M.,Walker, J.E.,Yokoyama, S.,Iwata, S.,Murata, T.
Structure of the rotor ring modified with N,N'-dicyclohexylcarbodiimide of the Na+-transporting vacuolar ATPase.
Proc.Natl.Acad.Sci.USA, 108:13474-13479, 2011

PubMed Abstract: The prokaryotic V-ATPase of Enterococcus hirae, closely related to the eukaryotic enzymes, provides a unique opportunity to study the ion-translocation mechanism because it transports Na(+), which can be detected by radioisotope (22Na(+)) experiments and X-ray crystallography. In this study, we demonstrated that the binding affinity of the rotor ring (K ring) for 22Na(+) decreased approximately 30-fold by reaction with N,N(')-dicyclohexylcarbodiimide (DCCD), and determined the crystal structures of Na(+)-bound and Na(+)-unbound K rings modified with DCCD at 2.4- and 3.1-Å resolutions, respectively. Overall these structures were similar, indicating that there is no global conformational change associated with release of Na(+) from the DCCD-K ring. A conserved glutamate residue (E139) within all 10 ion-binding pockets of the K ring was neutralized by modification with DCCD, and formed an "open" conformation by losing hydrogen bonds with the Y68 and T64 side chains, resulting in low affinity for Na(+). This open conformation is likely to be comparable to that of neutralized E139 forming a salt bridge with the conserved arginine of the stator during the ion-translocation process. Based on these findings, we proposed the ion-translocation model that the binding affinity for Na(+) decreases due to the neutralization of E139, thus releasing bound Na(+), and that the structures of Na(+)-bound and Na(+)-unbound DCCD-K rings are corresponding to intermediate states before and after release of Na(+) during rotational catalysis of V-ATPase, respectively.

PubMed: 21813759
DOI: 10.1073/pnas.1103287108

Experimental method

X-RAY DIFFRACTION (3.14 Å)