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PDBsum entry 2w2p

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protein metals Protein-protein interface(s) links
Hydrolase/receptor PDB id
2w2p
Jmol
Contents
Protein chains
274 a.a. *
48 a.a. *
93 a.a. *
Metals
_CA
Waters ×31
* Residue conservation analysis
PDB id:
2w2p
Name: Hydrolase/receptor
Title: Pcsk9-deltac d374a mutant bound to wt egf-a of ldlr
Structure: Proprotein convertase subtilisin/kexin type 9. Chain: p. Fragment: prodomain, residues 53-152. Synonym: proprotein convertase pc9, subtilisin/kexin-like p pc9, neural apoptosis-regulated convertase 1, narc-1, pcsk engineered: yes. Proprotein convertase subtilisin/kexin type 9. Chain: a. Fragment: catalytic domain, residues 153-451.
Source: Homo sapiens. Human. Organism_taxid: 9606. Expressed in: escherichia coli. Expression_system_taxid: 469008.
Resolution:
2.62Å     R-factor:   0.228     R-free:   0.270
Authors: M.J.Bottomley,A.Cirillo,L.Orsatti,L.Ruggeri,T.S.Fisher,J.C.S R.T.Cummings,R.M.Cubbon,P.Lo Surdo,A.Calzetta,A.Noto,J.Bays M.Mattu,F.Talamo,R.De Francesco,C.P.Sparrow,A.Sitlani,A.Car
Key ref:
M.J.Bottomley et al. (2009). Structural and Biochemical Characterization of the Wild Type PCSK9-EGF(AB) Complex and Natural Familial Hypercholesterolemia Mutants. J Biol Chem, 284, 1313-1323. PubMed id: 19001363 DOI: 10.1074/jbc.M808363200
Date:
03-Nov-08     Release date:   18-Nov-08    
PROCHECK
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 Headers
 References

Protein chain
Pfam   ArchSchema ?
Q8NBP7  (PCSK9_HUMAN) -  Proprotein convertase subtilisin/kexin type 9
Seq:
Struc:
 
Seq:
Struc:
692 a.a.
274 a.a.*
Protein chain
Pfam   ArchSchema ?
P01130  (LDLR_HUMAN) -  Low-density lipoprotein receptor
Seq:
Struc:
 
Seq:
Struc:
860 a.a.
48 a.a.*
Protein chain
Pfam   ArchSchema ?
Q8NBP7  (PCSK9_HUMAN) -  Proprotein convertase subtilisin/kexin type 9
Seq:
Struc:
 
Seq:
Struc:
692 a.a.
93 a.a.
Key:    PfamA domain  PfamB domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 9 residue positions (black crosses)

 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     proteolysis   1 term 
  Biochemical function     calcium ion binding     2 terms  

 

 
DOI no: 10.1074/jbc.M808363200 J Biol Chem 284:1313-1323 (2009)
PubMed id: 19001363  
 
 
Structural and Biochemical Characterization of the Wild Type PCSK9-EGF(AB) Complex and Natural Familial Hypercholesterolemia Mutants.
M.J.Bottomley, A.Cirillo, L.Orsatti, L.Ruggeri, T.S.Fisher, J.C.Santoro, R.T.Cummings, R.M.Cubbon, P.Lo Surdo, A.Calzetta, A.Noto, J.Baysarowich, M.Mattu, F.Talamo, R.De Francesco, C.P.Sparrow, A.Sitlani, A.Carfí.
 
  ABSTRACT  
 
PCSK9 regulates low density lipoprotein receptor (LDLR) levels and consequently is a target for the prevention of atherosclerosis and coronary heart disease. Here we studied the interaction, of LDLR EGF(A/AB) repeats with PCSK9. We show that PCSK9 binds the EGF(AB) repeats in a pH-dependent manner. Although the PCSK9 C-terminal domain is not involved in LDLR binding, PCSK9 autocleavage is required. Moreover, we report the x-ray structure of the PCSK9DeltaC-EGF(AB) complex at neutral pH. Compared with the low pH PCSK9-EGF(A) structure, the new structure revealed rearrangement of the EGF(A) His-306 side chain and disruption of the salt bridge with PCSK9 Asp-374, thus suggesting the basis for enhanced interaction at low pH. In addition, the structure of PCSK9DeltaC bound to EGF(AB)(H306Y), a mutant associated with familial hypercholesterolemia (FH), reveals that the Tyr-306 side chain forms a hydrogen bond with PCSK9 Asp-374, thus mimicking His-306 in the low pH conformation. Consistently, Tyr-306 confers increased affinity for PCSK9. Importantly, we found that although the EGF(AB)(H306Y)-PCSK9 interaction is pH-independent, LDLR(H306Y) binds PCSK9 50-fold better at low pH, suggesting that factors other than His-306 contribute to the pH dependence of PCSK9-LDLR binding. Further, we determined the structures of EGF(AB) bound to PCSK9DeltaC containing the FH-associated D374Y and D374H mutations, revealing additional interactions with EGF(A) mediated by Tyr-374/His-374 and providing a rationale for their disease phenotypes. Finally, we report the inhibitory properties of EGF repeats in a cellular assay measuring LDL uptake.
 
  Selected figure(s)  
 
Figure 1.
TR-FRET analyses of the PCSK9-LDLR interaction and its inhibition. A, in a TR-FRET assay at neutral pH, EGF(A) and EGF(AB) compete with the LDLR ectodomain for binding to PCSK9 with low/submicromolar IC[50] values. B, in the TR-FRET assay, full-length WT PCSK9 and WT PCSK9ΔC are equipotent at disrupting the interaction of labeled WT PCSK9 with labeled LDLR, demonstrating that the C-terminal domain of PCSK9 is not required for binding. In contrast with WT PCSK9ΔC, an unprocessed form of PCSK9ΔC (S386A) was unable to disrupt the interaction.
Figure 3.
Crystal structure of the PCSK9ΔC-EGF(AB) complex. Ribbon (A) and surface (B) representations, in the same orientation, of the complex formed between PCSK9ΔC and EGF(A) (yellow). EGF(A) contacts PCSK9 via the catalytic domain (red) but not the prodomain (blue). The P′ helix, which is part of the catalytic domain, is shown in pink. Dotted lines represent PCSK9 residues for which no electron density was observed. C, transparent surface representation of PCSK9ΔC; blue-labeled patches indicate residues contacted by EGF(A) (yellow ribbon; the yellow sphere is the Ca^2+ ion). Contacts are all less than 4 Å. Asp-374 (PCSK9) is at the tip of a β-hairpin loop. EGF(A) His-306 adopts a conformation that does not allow interaction with Asp-374 at neutral pH. Ser-153 is the N terminus of the PCSK9 catalytic domain generated upon autocleavage. Hydrogen bonds are established between side chain atoms of the following pairs of residues from PCSK9 and EGF(A): Arg-194 to Asp-310 and Asn-295, Asp-238 to Asn-295, Thr-377 to Asn-309 and Asp-310, and between backbone atoms for Thr-377 to Asp-310 and Phe-379 to Cys-308. D, transparent surface representation of EGF(A); yellow-labeled patches indicate residues contacted by PCSK9ΔC. The protein is oriented as if removing PCSK9ΔC from the complex shown in panel C followed by a 180° y axis rotation.
 
  The above figures are reprinted by permission from the ASBMB: J Biol Chem (2009, 284, 1313-1323) copyright 2009.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
19196236 C.J.Duff, M.J.Scott, I.T.Kirby, S.E.Hutchinson, S.L.Martin, and N.M.Hooper (2009).
Antibody-mediated disruption of the interaction between PCSK9 and the low-density lipoprotein receptor.
  Biochem J, 419, 577-584.  
  19020338 J.D.Horton, J.C.Cohen, and H.H.Hobbs (2009).
PCSK9: a convertase that coordinates LDL catabolism.
  J Lipid Res, 50, S172-S177.  
19224862 M.C.McNutt, H.J.Kwon, C.Chen, J.R.Chen, J.D.Horton, and T.A.Lagace (2009).
Antagonism of Secreted PCSK9 Increases Low Density Lipoprotein Receptor Expression in HepG2 Cells.
  J Biol Chem, 284, 10561-10570.
PDB codes: 3gcw 3gcx
19635789 S.Poirier, G.Mayer, V.Poupon, P.S.McPherson, R.Desjardins, K.Ly, M.C.Asselin, R.Day, F.J.Duclos, M.Witmer, R.Parker, A.Prat, and N.G.Seidah (2009).
Dissection of the endogenous cellular pathways of PCSK9-induced low density lipoprotein receptor degradation: evidence for an intracellular route.
  J Biol Chem, 284, 28856-28864.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB codes are shown on the right.