PDBsum entry 3fgo

Hydrolase

PDB id: 3fgo

Contents

Protein chains

994 a.a. *

Ligands

MF4 ×2

CZA ×2

ACP ×2

ACT ×4

Metals

_MG ×2

__K ×2

_MN ×3

Waters ×465

* Residue conservation analysis

PDB id:

3fgo

Name:

Hydrolase

Title:

Crystal structure of the e2 magnesium fluoride complex of the (sr) ca2+-atpase with bound cpa and amppcp

Structure:

Sarcoplasmic/endoplasmic reticulum calcium atpase 1. Chain: a, b. Synonym: serca1, calcium pump 1, calcium-transporting atpase sarcoplasmic reticulum type, fast twitch skeletal muscle isoform, sr ca(2+)-atpase 1, endoplasmic reticulum class 1/2 ca(2+) atpase. Ec: 3.6.3.8

Source:

Oryctolagus cuniculus. Rabbit. Organism_taxid: 9986

Resolution:

2.50Å R-factor: 0.176 R-free: 0.217

Authors:

M.Laursen,M.Bublitz,K.Moncoq,C.Olesen,J.V.Moller,H.S.Young,P.Nissen, J.P.Morth

Key ref:

M.Laursen et al. (2009). Cyclopiazonic Acid Is Complexed to a Divalent Metal Ion When Bound to the Sarcoplasmic Reticulum Ca2+-ATPase. J Biol Chem, 284, 13513-13518. PubMed id: 19289472 DOI: 10.1074/jbc.C900031200

Date:

08-Dec-08 Release date: 07-Apr-09

Protein chains

P04191  (AT2A1_RABIT) -  Sarcoplasmic/endoplasmic reticulum calcium ATPase 1 from Oryctolagus cuniculus

Seq: 1001 a.a.

Struc: 994 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.7.2.2.10 - P-type Ca(2+) transporter.
• Reaction: Ca²⁺(in) + ATP + H2O = Ca²⁺(out) + ADP + phosphate + H⁺

Ca(2+)(in) + ATP + H2O = Ca(2+)(out) (Bound ligand (Het Group name = ACP) matches with 81.25% similarity) + ADP + phosphate + H(+)

• Cofactor: Mg(2+)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1074/jbc.C900031200

J Biol Chem 284:13513-13518 (2009)

PubMed id: 19289472

Cyclopiazonic Acid Is Complexed to a Divalent Metal Ion When Bound to the Sarcoplasmic Reticulum Ca2+-ATPase.

M.Laursen, M.Bublitz, K.Moncoq, C.Olesen, J.V.Møller, H.S.Young, P.Nissen, J.P.Morth.

ABSTRACT

We have determined the structure of the sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA) in an E2.P(i)-like form stabilized as a complex with MgF(4)(2-), an ATP analog, adenosine 5'-(beta,gamma-methylene)triphosphate (AMPPCP), and cyclopiazonic acid (CPA). The structure determined at 2.5A resolution leads to a significantly revised model of CPA binding when compared with earlier reports. It shows that a divalent metal ion is required for CPA binding through coordination of the tetramic acid moiety at a characteristic kink of the M1 helix found in all P-type ATPase structures, which is expected to be part of the cytoplasmic cation access pathway. Our model is consistent with the biochemical data on CPA function and provides new measures in structure-based drug design targeting Ca(2+)-ATPases, e.g. from pathogens. We also present an extended structural basis of ATP modulation pinpointing key residues at or near the ATP binding site. A structural comparison to the Na(+),K(+)-ATPase reveals that the Phe(93) side chain occupies the equivalent binding pocket of the CPA site in SERCA, suggesting an important role of this residue in stabilization of the potassium-occluded E2 state of Na(+),K(+)-ATPase.

Selected figure(s)

Figure 1.
Interpretation of the SERCA·CPA complex. A, overall structure of SERCA showing the N-domain (red), A-domain (yellow), P-domain (purple), and transmembrane region (light blue). A 2F[o] - F[c] electron density map, contoured at 1σ, shows electron density both for AMPPCP between the N- and A-domains and for the CPA binding site in the Ca^2+ entry channel. The Mn^2+ ion is shown by a van der Waals sphere representation (orange) both in the AMPPCP site and in the CPA site. B, the AMPPCP binding site. The residues Asn^628, Arg^678, and Lys^205 are within interacting distance of the γ-phosphate in AMPPCP, and Asp^203 stabilizes Arg^678. Hydrogen bonding to a water molecule (W4) is colored blue and shown as dashed lines, whereas other hydrogen-bonding networks are colored black. Arg^174 and Glu^439 make a salt bridge linking the A- and N-domains. The 2F[o] - F[c] map is contoured at 1σ (blue mesh), covering the AMPPCPC molecule. C, transmembrane domain of SERCA with bound CPA. Three residues in the transmembrane segment of SERCA are involved in polar interactions with CPA: Gln^56, Asp^59, and Asn^101. Both Asn^101 and Gln^56 contribute with side chain and backbone atoms to the coordination of the tetramic acid part of CPA. The side chain oxygen of Gln^56 participates in the coordination sphere of the manganese atom. The anomalous difference Fourier map (orange mesh) contoured at 10 σ identifies the Mn^2+ ion coordinated at the CPA-SERCA binding interface. D, SERCA structures with TG-Boc12-ADT (green) (PDB ID 2BY4) and BHQ (blue) (PDB ID 2AGV) are superimposed onto transmembrane helices 1–5 of our new structure (PDB ID 3FGO). TG-Boc12-ADT, BHQ, and CPA·Mn^2+ (yellow) are shown in stick representation. The drug pocket is viewed from the cytoplasmic side of the SR membrane. E, a structural alignment between the Na^+, K^+-ATPase (purple) and SERCA (light blue), both stabilized with . The M1 kink region of the Na^+, K^+-ATPase exhibits a significant structural difference to SERCA. The intruding M1 loop is stabilized by Phe^93 in the Na^+,K^+-ATPase, occupying the same binding pocket as the indole moiety of CPA and the aromatic moiety of BHQ in SERCA.

Figure 2.
Binding pocket of Cyclopiazonic acid. A, topology of the CPA binding site. Conjugated doublebond systems of the CPA molecule are high lighted in green. Atom numbering in CPA is according to International Union of Pure and Applied Chemistry (IUPAC) nomenclature. B, fragment-based view of the CPA binding pocket. The drug binding pocket can be subdivided into three regions, each accommodating drug moieties of distinct chemical character. The polar region (designated A, highlighted in yellow) coordinates the acyl-tetramic acid moiety, with Gln^56, Asp^59, and Asn^101 as main interacting residues. The coordinated M^2+ occupies a central position in this interaction. The center of the pocket displays a preference for conjugated π-systems (B, highlighted in blue). BHQ binds to this part, superposing with CPA in this “B-pocket.” The relatively wide hydrophobic region of the pocket (C, highlighted in green) accommodates bulky, non-polar moieties. The decomposed analysis of this site proposes functional extensions or modifications to yield tailored high affinity drugs. For instance, an aliphatic chain at site D, bridging toward the TG binding site, and a suitable hydrogen-bonding partner to Asp^254 (region E), may further improve drug affinity.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2009, 284, 13513-13518) copyright 2009.