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PDBsum entry 1jk1
Go to PDB code: 
protein dna_rna metals links
Transcription/DNA PDB id
1jk1

 

 

 

 

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Contents
Protein chain
85 a.a. *
DNA/RNA
Metals
_ZN ×3
Waters ×136
* Residue conservation analysis
PDB id:
1jk1
Name: Transcription/DNA
Title: Zif268 d20a mutant bound to wt DNA site
Structure: 5'-d( Ap Gp Cp Gp Tp Gp Gp Gp Cp Gp G)-3'. Chain: b. Engineered: yes. 5'-d( Tp Cp Cp Gp Cp Cp Cp Ap Cp Gp C)-3'. Chain: c. Engineered: yes. Zif268. Chain: a. Fragment: zinc fingers (residues 333-421).
Source: Synthetic: yes. Mus musculus. House mouse. Organism_taxid: 10090. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008.
Biol. unit: Trimer (from PQS)
Resolution:
1.90Å     R-factor:   0.214     R-free:   0.266
Authors: J.C.Miller,C.O.Pabo
Key ref:
J.C.Miller and C.O.Pabo (2001). Rearrangement of side-chains in a Zif268 mutant highlights the complexities of zinc finger-DNA recognition. J Mol Biol, 313, 309-315. PubMed id: 11800559 DOI: 10.1006/jmbi.2001.4975
Date:
11-Jul-01     Release date:   19-Oct-01    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chain
Pfam   ArchSchema ?
P08046  (EGR1_MOUSE) -  Early growth response protein 1 from Mus musculus
Seq:
Struc: 		 
Seq:
Struc: 		533 a.a.
85 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 1 residue position (black cross)

DNA/RNA chains
  A-G-C-G-T-G-G-G-C-G-G 11 bases
  T-C-C-G-C-C-C-A-C-G-C 11 bases

 

 
DOI no: 10.1006/jmbi.2001.4975 J Mol Biol 313:309-315 (2001)
PubMed id: 11800559  
 
 
Rearrangement of side-chains in a Zif268 mutant highlights the complexities of zinc finger-DNA recognition.
J.C.Miller, C.O.Pabo.
 
  ABSTRACT  
 
Structural and biochemical studies of Cys(2)His(2) zinc finger proteins initially led several groups to propose a "recognition code" involving a simple set of rules relating key amino acid residues in the zinc finger protein to bases in its DNA site. One recent study from our group, involving geometric analysis of protein-DNA interactions, has discussed limitations of this idea and has shown how the spatial relationship between the polypeptide backbone and the DNA helps to determine what contacts are possible at any given position in a protein-DNA complex. Here we report a study of a zinc finger variant that highlights yet another source of complexity inherent in protein-DNA recognition. In particular, we find that mutations can cause key side-chains to rearrange at the protein-DNA interface without fundamental changes in the spatial relationship between the polypeptide backbone and the DNA. This is clear from a simple analysis of the binding site preferences and co-crystal structures for the Asp20-->Ala point mutant of Zif268. This point mutation in finger one changes the specificity of the protein from GCG TGG GCG to GCG TGG GC(G/T), and we have solved crystal structures of the D20A mutant bound to both types of sites. The structure of the D20A mutant bound to the GCG site reveals that contacts from key residues in the recognition helix are coupled in complex ways. The structure of the complex with the GCT site also shows an important new water molecule at the protein-DNA interface. These side-chain/side-chain interactions, and resultant changes in hydration at the interface, affect binding specificity in ways that cannot be predicted either from a simple recognition code or from analysis of spatial relationships at the protein-DNA interface. Accurate computer modeling of protein-DNA interfaces remains a challenging problem and will require systematic strategies for modeling side-chain rearrangements and change in hydration.
 
  Selected figure(s)  
 
Figure 4.
Figure 4. Stereo representation of simulated-annealing F[o] -F[c] electron density of the D20A- Image structure with residues 18 and 21 omitted from the calculation. The map is contoured at 3 s, and was generated using the program O and rendered using the SwissPDBViewer and POV Ray. 		Figure 5.
Figure 5. Comparison of the D20A mutant bound to different DNA sites and comparison with wt Zif268 structure. (a) Region of the D20A-GCG structure focusing on the interacting residues. Broken lines indicate hydrogen bonds. Simple modeling indicates that Glu21 could not obtain this new conformation in the wild-type complex without making electrostatically unfavorable contacts with Asp20. Modeling also suggests that this new Glu21 conformation would collide with the thymine methyl group if the mutant was bound to a Image site and this observation fits nicely with the binding data for the two proteins at the Image and Image sites. (b) Same region of the wild-type Zif268 structure[16] shown in previous panel. (c) Comparison of the interaction between Glu21 and Arg18 in the D20A- Image structure with the interaction between Asp20 and Arg18 in the wild-type structure. Broken lines indicate hydrogen bonds and distances are given in Å. (d) Corresponding region of the D20A-GCT structure. Only the most relevant contact made by the secondary Arg18 conformation is shown for clarity. We explored the possibility that the electron density modeled as an ordered water was actually due to an alternate conformation of Arg18, but we were not able to fit the Arg18 guanidinium group into this density without severely distorting the side-chain geometry and generating several unacceptable steric clashes.
 
  The above figures are reprinted by permission from Elsevier: J Mol Biol (2001, 313, 309-315) copyright 2001.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20099003 F.Li, D.Fitz, D.G.Fraser, and B.M.Rode (2010).
Arginine in the salt-induced peptide formation reaction: enantioselectivity facilitated by glycine, L- and D-histidine.
  Amino Acids, 39, 579-585.  
19056825 J.D.Sander, P.Zaback, J.K.Joung, D.F.Voytas, and D.Dobbs (2009).
An affinity-based scoring scheme for predicting DNA-binding activities of modularly assembled zinc-finger proteins.
  Nucleic Acids Res, 37, 506-515.  
18586699 J.Liu, and G.D.Stormo (2008).
Context-dependent DNA recognition code for C2H2 zinc-finger transcription factors.
  Bioinformatics, 24, 1850-1857.  
18332042 M.B.Noyes, X.Meng, A.Wakabayashi, S.Sinha, M.H.Brodsky, and S.A.Wolfe (2008).
A systematic characterization of factors that regulate Drosophila segmentation via a bacterial one-hybrid system.
  Nucleic Acids Res, 36, 2547-2560.  
18585359 M.F.Berger, G.Badis, A.R.Gehrke, S.Talukder, A.A.Philippakis, L.Peña-Castillo, T.M.Alleyne, S.Mnaimneh, O.B.Botvinnik, E.T.Chan, F.Khalid, W.Zhang, D.Newburger, S.A.Jaeger, Q.D.Morris, M.L.Bulyk, and T.R.Hughes (2008).
Variation in homeodomain DNA binding revealed by high-resolution analysis of sequence preferences.
  Cell, 133, 1266-1276.  
18545224 T.Cathomen, and J.K.Joung (2008).
Zinc-finger nucleases: the next generation emerges.
  Mol Ther, 16, 1200-1207.  
18922190 V.E.Angarica, A.G.Pérez, A.T.Vasconcelos, J.Collado-Vides, and B.Contreras-Moreira (2008).
Prediction of TF target sites based on atomistic models of protein-DNA complexes.
  BMC Bioinformatics, 9, 436.  
17214883 F.Spyrakis, P.Cozzini, C.Bertoli, A.Marabotti, G.E.Kellogg, and A.Mozzarelli (2007).
Energetics of the protein-DNA-water interaction.
  BMC Struct Biol, 7, 4.  
17603475 J.C.Miller, M.C.Holmes, J.Wang, D.Y.Guschin, Y.L.Lee, I.Rupniewski, C.M.Beausejour, A.J.Waite, N.S.Wang, K.A.Kim, P.D.Gregory, C.O.Pabo, and E.J.Rebar (2007).
An improved zinc-finger nuclease architecture for highly specific genome editing.
  Nat Biotechnol, 25, 778-785.  
17264128 T.W.Siggers, and B.Honig (2007).
Structure-based prediction of C2H2 zinc-finger binding specificity: sensitivity to docking geometry.
  Nucleic Acids Res, 35, 1085-1097.  
16402211 J.Miertus, W.Borozdin, V.Frecer, G.Tonini, S.Bertok, A.Amoroso, S.Miertus, and J.Kohlhase (2006).
A SALL4 zinc finger missense mutation predicted to result in increased DNA binding affinity is associated with cranial midline defects and mild features of Okihiro syndrome.
  Hum Genet, 119, 154-161.  
16998473 M.F.Berger, A.A.Philippakis, A.M.Qureshi, F.S.He, P.W.Estep, and M.L.Bulyk (2006).
Compact, universal DNA microarrays to comprehensively determine transcription-factor binding site specificities.
  Nat Biotechnol, 24, 1429-1435.  
16606702 S.Huntley, D.M.Baggott, A.T.Hamilton, M.Tran-Gyamfi, S.Yang, J.Kim, L.Gordon, E.Branscomb, and L.Stubbs (2006).
A comprehensive catalog of human KRAB-associated zinc finger genes: insights into the evolutionary history of a large family of transcriptional repressors.
  Genome Res, 16, 669-677.  
16246914 A.V.Morozov, J.J.Havranek, D.Baker, and E.D.Siggia (2005).
Protein-DNA binding specificity predictions with structural models.
  Nucleic Acids Res, 33, 5781-5798.  
16014175 J.Liu, and G.D.Stormo (2005).
Quantitative analysis of EGR proteins binding to DNA: assessing additivity in both the binding site and the protein.
  BMC Bioinformatics, 6, 176.  
16186128 J.Liu, and G.D.Stormo (2005).
Combining SELEX with quantitative assays to rapidly obtain accurate models of protein-DNA interactions.
  Nucleic Acids Res, 33, e141.  
16244132 S.S.Houshmandi, and W.M.Olivas (2005).
Yeast Puf3 mutants reveal the complexity of Puf-RNA binding and identify a loop required for regulation of mRNA decay.
  RNA, 11, 1655-1666.  
15613596 G.Paillard, C.Deremble, and R.Lavery (2004).
Looking into DNA recognition: zinc finger binding specificity.
  Nucleic Acids Res, 32, 6673-6682.  
15247345 M.D.Simon, K.Sato, G.A.Weiss, and K.M.Shokat (2004).
A phage display selection of engrailed homeodomain mutants and the importance of residue Q50.
  Nucleic Acids Res, 32, 3623-3631.  
15338611 A.T.Hamilton, S.Huntley, J.Kim, E.Branscomb, and L.Stubbs (2003).
Lineage-specific expansion of KRAB zinc-finger transcription factor genes: implications for the evolution of vertebrate regulatory networks.
  Cold Spring Harb Symp Quant Biol, 68, 131-140.  
14602907 C.D.Carvin, R.D.Parr, and M.P.Kladde (2003).
Site-selective in vivo targeting of cytosine-5 DNA methylation by zinc-finger proteins.
  Nucleic Acids Res, 31, 6493-6501.  
14652076 E.P.Baldwin, S.S.Martin, J.Abel, K.A.Gelato, H.Kim, P.G.Schultz, and S.W.Santoro (2003).
A specificity switch in selected cre recombinase variants is mediated by macromolecular plasticity and water.
  Chem Biol, 10, 1085-1094.
PDB codes: 1pvp 1pvq 1pvr
12001270 P.V.Benos, A.S.Lapedes, and G.D.Stormo (2002).
Is there a code for protein-DNA recognition? Probab(ilistical)ly. . .
  Bioessays, 24, 466-475.  
12384591 P.V.Benos, M.L.Bulyk, and G.D.Stormo (2002).
Additivity in protein-DNA interactions: how good an approximation is it?
  Nucleic Acids Res, 30, 4442-4451.  
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.

 

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