title: Modulation and Functional Role of the Orientations of the N- and P-Domains of Cu+ -Transporting ATPase along the Ion Transport Cycle.
abstract: Ion transport of different P-type ATPases is regulated similarly through the interplay of multiple protein domains. In the presence of ATP, binding of a cation to the ion binding site in the transmembrane helices leads to the phosphorylation of the P-domain, allowing ion transfer across the membrane. The details of the mechanism, however, are not clear. Here, we report the modulation of the orientation between the N- and P-domains of Cu(+)-transporting ATPase along the ion transport cycle using high-resolution nuclear magnetic resonance spectroscopy in solution. On the basis of residual dipolar coupling measurements, it is found that the interdomain orientation (relative openness) of the N- and P-domains is distinctly modulated depending on the specific state of the N- and P-domains along the ion translocation cycle. The two domains' relative position in the apo state is semiopen, whereas it becomes closed upon binding of ATP to the N-domain. After phosphorylation of the P-domain and the release of ADP, the opening, however, becomes the widest among all the states. We reason such wide opening resulting from the departure of ADP prepares the N- and P-domains to accommodate the A-domain for interaction and, hence, promote ion transport and allow dephosphorylation of the P-domain. Such wide interdomain opening is abolished when an Asn to Asp mutation is introduced into the conserved DXXK motif located in the hinge region of the N- and P-domains of Cu(+)-ATPase, suggesting the indispensible role of the N- and P-interdomain orientation during ion transportation. Our results shed new light on the structural and mechanistic details of P-type ATPase function at large.
authors.forename: Dan
authors.initials: D
authors.lastname: Meng
authors.forename: Lei
authors.initials: L
authors.lastname: Bruschweiler-Li
authors.affiliation: ‡National High Magnetic Field Laboratory, Tallahassee, Florida 32310, United States.
authors.forename: Fengli
authors.initials: F
authors.lastname: Zhang
authors.affiliation: ‡National High Magnetic Field Laboratory, Tallahassee, Florida 32310, United States.
authors.forename: Rafael
authors.initials: R
authors.lastname: Brüschweiler
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020816:Amino Acid Motifs; D019943:Amino Acid Substitution; D019843:Archaeal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019611:Archaeoglobus fulgidus / Q000235:genetics / Q000378:metabolism; D027682:Cation Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D017124:Conserved Sequence; D003300:Copper / Q000378:metabolism*; D000073840:Copper-Transporting ATPases; D019766:Genes, Archaeal; D017136:Ion Transport; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D019906:Nuclear Magnetic Resonance, Biomolecular; D011487:Protein Conformation; D054730:Protein Interaction Domains and Motifs; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D019843:Archaeal Proteins; D027682:Cation Transport Proteins; D011994:Recombinant Proteins; D003300:Copper; D000251:Adenosine Triphosphatases; D000073840:Copper-Transporting ATPases