PDBsum entry 2p46

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protein metals Protein-protein interface(s) links
Hydrolase/immune system PDB id
Jmol PyMol
Protein chains
120 a.a. *
121 a.a. *
_ZN ×11
Waters ×83
* Residue conservation analysis
PDB id:
Name: Hydrolase/immune system
Title: Complex of a camelid single-domain vhh antibody fragment wit at 2.5a resolution: se5b-ortho-2 crystal form with five se- (l4m, m34, m51, f68m, m83) in vhh scaffold.
Structure: Ribonuclease pancreatic. Chain: a, c. Synonym: rnase 1, rnase a. Antibody cab-rn05. Chain: b, d. Engineered: yes
Source: Bos taurus. Cattle. Organism_taxid: 9913. Camelus dromedarius. Arabian camel. Organism_taxid: 9838. Expressed in: escherichia coli. Expression_system_taxid: 562
2.50Å     R-factor:   0.225     R-free:   0.299
Authors: V.Tereshko,S.Uysal,A.Koide,K.Margalef,S.Koide,A.A.Kossiakoff
Key ref:
V.Tereshko et al. (2008). Toward chaperone-assisted crystallography: protein engineering enhancement of crystal packing and X-ray phasing capabilities of a camelid single-domain antibody (VHH) scaffold. Protein Sci, 17, 1175-1187. PubMed id: 18445622 DOI: 10.1110/ps.034892.108
11-Mar-07     Release date:   11-Mar-08    
Go to PROCHECK summary

Protein chains
Pfam   ArchSchema ?
P61823  (RNAS1_BOVIN) -  Ribonuclease pancreatic
150 a.a.
120 a.a.
Protein chains
No UniProt id for this chain
Struc: 121 a.a.
Key:    PfamA domain  Secondary structure  CATH domain

 Enzyme reactions 
   Enzyme class: Chains A, C: E.C.  - Pancreatic ribonuclease.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: Endonucleolytic cleavage to nucleoside 3'-phosphates and 3'-phosphooligonucleotides ending in C-P or U-P with 2',3'-cyclic phosphate intermediates.
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     extracellular region   1 term 
  Biological process     nucleic acid phosphodiester bond hydrolysis   3 terms 
  Biochemical function     nucleic acid binding     7 terms  


DOI no: 10.1110/ps.034892.108 Protein Sci 17:1175-1187 (2008)
PubMed id: 18445622  
Toward chaperone-assisted crystallography: protein engineering enhancement of crystal packing and X-ray phasing capabilities of a camelid single-domain antibody (VHH) scaffold.
V.Tereshko, S.Uysal, A.Koide, K.Margalef, S.Koide, A.A.Kossiakoff.
A crystallization chaperone is an auxiliary protein that binds to a target of interest, enhances and modulates crystal packing, and provides high-quality phasing information. We critically evaluated the effectiveness of a camelid single-domain antibody (V(H)H) as a crystallization chaperone. By using a yeast surface display system for V(H)H, we successfully introduced additional Met residues in the core of the V(H)H scaffold. We identified a set of SeMet-labeled V(H)H variants that collectively produced six new crystal forms as the complex with the model antigen, RNase A. The crystals exhibited monoclinic, orthorhombic, triclinic, and tetragonal symmetry and have one or two complexes in the asymmetric unit, some of which diffracted to an atomic resolution. The phasing power of the Met-enriched V(H)H chaperone allowed for auto-building the entire complex using single-anomalous dispersion technique (SAD) without the need for introducing SeMet into the target protein. We show that phases produced by combining SAD and V(H)H model-based phases are accurate enough to easily solve structures of the size reported here, eliminating the need to collect multiple wavelength multiple-anomalous dispersion (MAD) data. Together with the presence of high-throughput selection systems (e.g., phage display libraries) for V(H)H, the enhanced V(H)H domain described here will be an excellent scaffold for producing effective crystallization chaperones.
  Selected figure(s)  
Figure 3.
Figure 3. Schematics of the packing modes found in the crystals of the cAb-RN05 V[H]H–RNase A complexes. The unit cell is indicated in red. The orientation of the crystal axes is shown in black. The cyan arrow indicates the view shown in Supplemental Figure S2. The V[H]H and RNase A molecules are represented as white cylinders and cyan spheres, respectively. This representation is based on the calculation of molecular ellipsoids performed in MOLREP. (A) Monoclinic Mono-1 crystal form with one complex in ASU. This form was observed for the SE3 and SE5a complexes, crystallized isomorphously. (B) Monoclinic SE3-Mono-2 crystal form with two complexes in ASU. (C) Orthorhombic SE5b-Ortho-1 crystal form with one complex in ASU. (D) Orthorhombic SE5b-Ortho-2 crystal form with two complexes in ASU. (E) Trigonal SE5b-Tri crystal form with one complex in ASU. (F) Tetragonal SE5b-Tetra crystal form with one complex in ASU. (G) Original 1BZQ crystal structure crystallized in triclinic space group with four complexes in ASU.
Figure 4.
Figure 4. Favorable crystal contacts formed by dimerization of V[H]H through H-bonding in N-terminal β-strands. (A) V[H]H–V[H]H alignment in the SE5b dimer. Two V[H]H domains interact via their N-terminal βA'- and βA''-strands with the residues E6 (βA'), G8, G9, and L11 (βA'') being involved in the H-bonds (2.6–3.3 Å range). The SE5b-Tetra structure was selected for presentation. (Inset) Four H-bonding pairs found in SE5b dimers. The main-chain N- and O-atoms involved in H-bonds are shown as small circles colored in blue and red, respectively. There are four H-bonds in Se5b-Tri and Se5b-Tetra crystals (shown as filled circles). In Se5b-Ortho-2, the residues E6 and L11 contribute both N- and O-atoms in intermolecular H-bonding network, resulting in two additional H-bonds (shown as open circles). (B). V[H]H–V[H]H alignment in the 1BZQ dimer. Two V[H]H domains interact via their N-terminal βA''-strands with the residues G9, L11, and Q13 being involved in the H-bonds. (Inset) Three H-bonding pairs found in 1BZQ. L11 contributes both N- and O-atoms in H-bonding network, resulting in a total of four H-bonds.
  The above figures are reprinted by permission from the Protein Society: Protein Sci (2008, 17, 1175-1187) copyright 2008.  
  Figures were selected by an automated process.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
22743979 A.D.Lawson (2012).
Antibody-enabled small-molecule drug discovery.
  Nat Rev Drug Discov, 11, 519-525.  
21217145 J.C.Pai, J.A.Culver, J.E.Drury, R.S.Motani, R.L.Lieberman, and J.A.Maynard (2011).
Conversion of scFv peptide-binding specificity for crystal chaperone development.
  Protein Eng Des Sel, 24, 419-428.  
20969962 M.Wu, Y.J.Park, E.Pardon, S.Turley, A.Hayhurst, J.Deng, J.Steyaert, and W.G.Hol (2011).
Structures of a key interaction protein from the Trypanosoma brucei editosome in complex with single domain antibodies.
  J Struct Biol, 174, 124-136.
PDB codes: 3k7u 3k80 3k81
21151117 Y.Koldobskaya, E.M.Duguid, D.M.Shechner, N.B.Suslov, J.Ye, S.S.Sidhu, D.P.Bartel, S.Koide, A.A.Kossiakoff, and J.A.Piccirilli (2011).
A portable RNA sequence whose recognition by a synthetic antibody facilitates structural determination.
  Nat Struct Mol Biol, 18, 100-106.  
20458517 D.Smolarek, C.Hattab, G.Hassanzadeh-Ghassabeh, S.Cochet, C.Gutiérrez, Brevern, R.Udomsangpetch, J.Picot, M.Grodecka, K.Wasniowska, S.Muyldermans, Y.Colin, C.Le Van Kim, M.Czerwinski, and O.Bertrand (2010).
A recombinant dromedary antibody fragment (VHH or nanobody) directed against human Duffy antigen receptor for chemokines.
  Cell Mol Life Sci, 67, 3371-3387.  
20739007 J.K.Lee, and R.M.Stroud (2010).
Unlocking the eukaryotic membrane protein structural proteome.
  Curr Opin Struct Biol, 20, 464-470.  
20945358 M.H.Kubala, O.Kovtun, K.Alexandrov, and B.M.Collins (2010).
Structural and thermodynamic analysis of the GFP:GFP-nanobody complex.
  Protein Sci, 19, 2389-2401.
PDB code: 3ogo
20382990 T.Beck, T.Gruene, and G.M.Sheldrick (2010).
The magic triangle goes MAD: experimental phasing with a bromine derivative.
  Acta Crystallogr D Biol Crystallogr, 66, 374-380.
PDB codes: 3gt3 3gt4
22477774 F.Gorrec (2009).
The MORPHEUS protein crystallization screen.
  J Appl Crystallogr, 42, 1035-1042.  
20004165 R.Ganesan, C.Eigenbrot, Y.Wu, W.C.Liang, S.Shia, M.T.Lipari, and D.Kirchhofer (2009).
Unraveling the allosteric mechanism of serine protease inhibition by an antibody.
  Structure, 17, 1614-1624.
PDB codes: 2wub 2wuc 3k2u
19477632 S.Koide (2009).
Engineering of recombinant crystallization chaperones.
  Curr Opin Struct Biol, 19, 449-457.  
19536805 T.M.Blois, and J.U.Bowie (2009).
G-protein-coupled receptor structures were not built in a day.
  Protein Sci, 18, 1335-1342.  
18854190 A.S.Chervin, D.H.Aggen, J.M.Raseman, and D.M.Kranz (2008).
Engineering higher affinity T cell receptors using a T cell display system.
  J Immunol Methods, 339, 175-184.  
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.