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PDBsum entry 1k9n

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protein Protein-protein interface(s) links
Membrane protein PDB id
1k9n
Jmol
Contents
Protein chains
18 a.a.
Theoretical model
PDB id:
1k9n
Name: Membrane protein
Title: Model of the pentameric transmembrane domain of phospholamban, a regulator of the calcium atpase serca2a
Structure: Cardiac phospholamban. Chain: a, b, c, d, e. Fragment: putative membrane spanning domain residues 35-52. Synonym: plb
Source: Homo sapiens. Human. Organ: heart. Tissue: sarcoplasmic reticulum
Authors: M.G.Paterlini,J.Karim,C.B.Karim,D.D.Thomas
Key ref:
C.B.Karim et al. (1998). Cysteine reactivity and oligomeric structures of phospholamban and its mutants. Biochemistry, 37, 12074-12081. PubMed id: 9724519 DOI: 10.1021/bi980642n
Date:
29-Oct-01     Release date:   14-Nov-01    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chains
Pfam   ArchSchema ?
P26678  (PPLA_HUMAN) -  Cardiac phospholamban
Seq:
Struc:
52 a.a.
18 a.a.
Key:    PfamA domain  Secondary structure

 

 
DOI no: 10.1021/bi980642n Biochemistry 37:12074-12081 (1998)
PubMed id: 9724519  
 
 
Cysteine reactivity and oligomeric structures of phospholamban and its mutants.
C.B.Karim, J.D.Stamm, J.Karim, L.R.Jones, D.D.Thomas.
 
  ABSTRACT  
 
To test models for the pentameric structure of phospholamban (PLB) and study its structure and molecular dynamics in SDS solution, we characterized recombinant PLB and several of its mutants by (a) reactivity of cysteine residues toward DTNB [5, 5'-dithiobis(2-nitrobenzoic acid)] and a thiol-reactive spin label, (b) oligomeric state on SDS-PAGE, and (c) EPR of the spin-labeled proteins. WT-PLB has three cysteine residues (36, 41, and 46), all located in the hydrophobic C-terminal transmembrane region. In SDS at pH 7.5, exhaustive reaction with either sulfhydryl reagent resulted in essentially 2 mol of cysteine reacted/mol of WT-PLB, with only slight destabilization of the native pentameric structure. When WT-PLB was denatured in guanidine at pH 8.1, all three cysteines reacted, disrupting the pentamer, which was restored upon cleavage of the disulfide bonds with DTT. In the tetrameric mutant C41L-PLB, the two remaining cysteine residues reacted, reversibly destabilizing the tetramer. In the monomeric mutant L37A-PLB, all three cysteines reacted. The pentameric double cysteine replacement mutant C36,46A-PLB showed negligible reactivity. We conclude that Cys-41 is the unreactive cysteine in PLB and is located at a crucial site for the maintenance of the pentameric structure. EPR spectra in SDS of spin-labeled WT-PLB and mutants correlate with the oligomeric state on SDS-PAGE; oligomeric proteins show decreased spin-label mobility compared with monomers. Molecular dynamics calculations were used to construct an atomic model for the transmembrane region of the PLB pentamer, constrained by previous mutagenesis results and the results of the present study. We conclude that (a) the mobilities of spin-labels attached to PLB and its mutants are sensitive to oligomeric state and (b) the pattern of cysteine reactivity, spin-label mobility, and oligomeric state supports a structural model for the PLB pentamer in which interactions between each pair of subunits are stabilized by a leucine-isoleucine zipper.
 

Literature references that cite this PDB file's key reference

  PubMed id Reference
21235348 J.M.Fukuto, and S.J.Carrington (2011).
HNO signaling mechanisms.
  Antioxid Redox Signal, 14, 1649-1657.  
21282613 K.N.Ha, L.R.Masterson, Z.Hou, R.Verardi, N.Walsh, G.Veglia, and S.L.Robia (2011).
Lethal Arg9Cys phospholamban mutation hinders Ca2+-ATPase regulation and phosphorylation by protein kinase A.
  Proc Natl Acad Sci U S A, 108, 2735-2740.  
19840770 S.Chu, A.T.Coey, and G.A.Lorigan (2010).
Solid-state (2)H and (15)N NMR studies of side-chain and backbone dynamics of phospholamban in lipid bilayers: investigation of the N27A mutation.
  Biochim Biophys Acta, 1798, 210-215.  
18611372 D.T.Moore, B.W.Berger, and W.F.DeGrado (2008).
Protein-protein interactions in the membrane: sequence, structural, and biological motifs.
  Structure, 16, 991.  
18287099 E.M.Kelly, Z.Hou, J.Bossuyt, D.M.Bers, and S.L.Robia (2008).
Phospholamban oligomerization, quaternary structure, and sarco(endo)plasmic reticulum calcium ATPase binding measured by fluorescence resonance energy transfer in living cells.
  J Biol Chem, 283, 12202-12211.  
18081313 N.J.Traaseth, K.N.Ha, R.Verardi, L.Shi, J.J.Buffy, L.R.Masterson, and G.Veglia (2008).
Structural and dynamic basis of phospholamban and sarcolipin inhibition of Ca(2+)-ATPase.
  Biochemistry, 47, 3.  
18247624 N.J.Traaseth, R.Verardi, and G.Veglia (2008).
Asymmetric methyl group labeling as a probe of membrane protein homo-oligomers by NMR spectroscopy.
  J Am Chem Soc, 130, 2400-2401.  
17908690 K.N.Ha, N.J.Traaseth, R.Verardi, J.Zamoon, A.Cembran, C.B.Karim, D.D.Thomas, and G.Veglia (2007).
Controlling the Inhibition of the Sarcoplasmic Ca2+-ATPase by Tuning Phospholamban Structural Dynamics.
  J Biol Chem, 282, 37205-37214.  
17996192 W.Liu, J.Z.Fei, T.Kawakami, and S.O.Smith (2007).
Structural constraints on the transmembrane and juxtamembrane regions of the phospholamban pentamer in membrane bilayers: Gln29 and Leu52.
  Biochim Biophys Acta, 1768, 2971-2978.  
15654870 A.D.van Dijk, R.Boelens, and A.M.Bonvin (2005).
Data-driven docking for the study of biomolecular complexes.
  FEBS J, 272, 293-312.  
15787961 A.M.Slovic, J.D.Lear, and W.F.DeGrado (2005).
De novo design of a pentameric coiled-coil: decoding the motif for tetramer versus pentamer formation in water-soluble phospholamban.
  J Pept Res, 65, 312-321.  
16043693 K.Oxenoid, and J.J.Chou (2005).
The structure of phospholamban pentamer reveals a channel-like architecture in membranes.
  Proc Natl Acad Sci U S A, 102, 10870-10875.
PDB code: 1zll
15298923 E.E.Metcalfe, J.Zamoon, D.D.Thomas, and G.Veglia (2004).
(1)H/(15)N heteronuclear NMR spectroscopy shows four dynamic domains for phospholamban reconstituted in dodecylphosphocholine micelles.
  Biophys J, 87, 1205-1214.  
15382237 Y.Park, M.Elsner, R.Staritzbichler, and V.Helms (2004).
Novel scoring function for modeling structures of oligomers of transmembrane alpha-helices.
  Proteins, 57, 577-585.  
14507721 J.Zamoon, A.Mascioni, D.D.Thomas, and G.Veglia (2003).
NMR solution structure and topological orientation of monomeric phospholamban in dodecylphosphocholine micelles.
  Biophys J, 85, 2589-2598.
PDB code: 1n7l
12833255 N.A.Lockwood, R.S.Tu, Z.Zhang, M.V.Tirrell, D.D.Thomas, and C.B.Karim (2003).
Structure and function of integral membrane protein domains resolved by peptide-amphiphiles: application to phospholamban.
  Biopolymers, 69, 283-292.  
12649422 W.F.DeGrado, H.Gratkowski, and J.D.Lear (2003).
How do helix-helix interactions help determine the folds of membrane proteins? Perspectives from the study of homo-oligomeric helical bundles.
  Protein Sci, 12, 647-665.  
12023229 J.Torres, J.A.Briggs, and I.T.Arkin (2002).
Contribution of energy values to the analysis of global searching molecular dynamics simulations of transmembrane helical bundles.
  Biophys J, 82, 3063-3071.  
11477077 C.B.Karim, M.G.Paterlini, L.G.Reddy, G.W.Hunter, G.Barany, and D.D.Thomas (2001).
Role of cysteine residues in structural stability and function of a transmembrane helix bundle.
  J Biol Chem, 276, 38814-38819.
PDB code: 1kch
11606281 J.Torres, A.Kukol, and I.T.Arkin (2001).
Mapping the energy surface of transmembrane helix-helix interactions.
  Biophys J, 81, 2681-2692.  
11371203 Q.Yao, L.T.Chen, J.Li, K.Brungardt, T.C.Squier, and D.J.Bigelow (2001).
Oligomeric interactions between phospholamban molecules regulate Ca-ATPase activity in functionally reconstituted membranes.
  Biochemistry, 40, 6406-6413.  
10978176 C.B.Karim, C.G.Marquardt, J.D.Stamm, G.Barany, and D.D.Thomas (2000).
Synthetic null-cysteine phospholamban analogue and the corresponding transmembrane domain inhibit the Ca-ATPase.
  Biochemistry, 39, 10892-10897.  
10679347 K.G.Fleming (2000).
Riding the wave: structural and energetic principles of helical membrane proteins.
  Curr Opin Biotechnol, 11, 67-71.  
10841762 S.Frank, R.A.Kammerer, S.Hellstern, S.Pegoraro, J.Stetefeld, A.Lustig, L.Moroder, and J.Engel (2000).
Toward a high-resolution structure of phospholamban: design of soluble transmembrane domain mutants.
  Biochemistry, 39, 6825-6831.  
10075652 L.G.Reddy, J.M.Autry, L.R.Jones, and D.D.Thomas (1999).
Co-reconstitution of phospholamban mutants with the Ca-ATPase reveals dependence of inhibitory function on phospholamban structure.
  J Biol Chem, 274, 7649-7655.  
10096878 P.Pollesello, A.Annila, and M.Ovaska (1999).
Structure of the 1-36 amino-terminal fragment of human phospholamban by nuclear magnetic resonance and modeling of the phospholamban pentamer.
  Biophys J, 76, 1784-1795.  
9837953 G.Chu, L.Li, Y.Sato, J.M.Harrer, V.J.Kadambi, B.D.Hoit, D.M.Bers, and E.G.Kranias (1998).
Pentameric assembly of phospholamban facilitates inhibition of cardiac function in vivo.
  J Biol Chem, 273, 33674-33680.  
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 code is shown on the right.