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Immunoglobulin PDB id
1ap2
Jmol
Contents
Protein chains
112 a.a. *
120 a.a. *
Waters ×102
* Residue conservation analysis
PDB id:
1ap2
Name: Immunoglobulin
Title: Single chain fv of c219
Structure: Monoclonal antibody c219. Chain: a, c. Fragment: fv. Synonym: variable domain. Engineered: yes. Other_details: light and heavy chains linked with a synthetic (ggggs)3 linker. Monoclonal antibody c219. Chain: b, d.
Source: Mus musculus. House mouse. Organism_taxid: 10090. Gene: cdna. Expressed in: escherichia coli. Expression_system_taxid: 562.
Biol. unit: Dimer (from PQS)
Resolution:
2.36Å     R-factor:   0.204     R-free:   0.284
Authors: P.J.Hoedemaeker,D.R.Rose
Key ref:
F.J.Hoedemaeker et al. (1997). A single chain Fv fragment of P-glycoprotein-specific monoclonal antibody C219. Design, expression, and crystal structure at 2.4 A resolution. J Biol Chem, 272, 29784-29789. PubMed id: 9368049 DOI: 10.1074/jbc.272.47.29784
Date:
23-Jul-97     Release date:   24-Dec-97    
PROCHECK
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 Headers
 References

 

 
DOI no: 10.1074/jbc.272.47.29784 J Biol Chem 272:29784-29789 (1997)
PubMed id: 9368049  
 
 
A single chain Fv fragment of P-glycoprotein-specific monoclonal antibody C219. Design, expression, and crystal structure at 2.4 A resolution.
F.J.Hoedemaeker, T.Signorelli, K.Johns, D.A.Kuntz, D.R.Rose.
 
  ABSTRACT  
 
A construct encoding a single chain variable fragment of the anti-P-glycoprotein monoclonal antibody C219 was made by combining the coding sequences for the heavy and light chain variable domains with a sequence encoding the flexible linker (GGGGS)3, an OmpA signal sequence, a c-myc identification tag, and a five-histidine purification tag. The construct was expressed in Escherichia coli and purified from the periplasmic fraction using a nickel chelate column and ion exchange chromatography. Three-step Western blot analysis showed that the construct retains binding affinity for P-glycoprotein. Crystals of 1.0 x 0.2 x 0.2 mm were grown in 100 mM citrate, pH 4.5, 21% polyethylene glycol 6000 in the presence of low concentrations of subtilisin, resulting in proteolytic removal of the linker and purification tags. The structure was solved to a resolution of 2.4 A with an R factor of 20.6, an Rfree of 28.5, and good stereochemistry. This result could lead to a clinically useful product based on antibody C219 for the diagnosis of P-glycoprotein-mediated multidrug resistance. The molecule will also be useful in biophysical studies of functional domains of P-glycoprotein, as well as studies of the intact molecule.
 
  Selected figure(s)  
 
Figure 4.
Fig. 4. Detail of the electron density at the V[L]-V[H] interface, contoured at 1 . View is "up" toward the CDRs along the pseudo-2-fold^ axis. Two short-range hydrogen bonds between glutamines in the^ light and heavy chains are indicated in green. Figs. 4 and 5A^ were produced with SETOR (49). 		Figure 6.
Fig. 6. Helical wheel plot of the peptide epitope. Hydrophobic residues are indicated with boldface lettering; residues that are potentially important for binding are boxed (13). The epitope^ sequence is extended by one helix turn (VVQEALDKARE) to show the^ continuous amphiphilicity.
 
  The above figures are reprinted by permission from the ASBMB: J Biol Chem (1997, 272, 29784-29789) copyright 1997.  
  Figures were selected by the author.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
21329716 Q.Xie, N.Tang, R.Wan, Y.Qi, X.Lin, and J.Lin (2011).
Recombinant snake venom cystatin inhibits the growth, invasion and metastasis of B16F10 cells and MHCC97H cells in vitro and in vivo.
  Toxicon, 57, 704-711.  
20731718 E.Crowley, M.L.O'Mara, I.D.Kerr, and R.Callaghan (2010).
Transmembrane helix 12 plays a pivotal role in coupling energy provision and drug binding in ABCB1.
  FEBS J, 277, 3974-3985.  
19816775 X.Gu, X.Jia, J.Feng, B.Shen, Y.Huang, S.Geng, Y.Sun, Y.Wang, Y.Li, and M.Long (2010).
Molecular modeling and affinity determination of scFv antibody: proper linker peptide enhances its activity.
  Ann Biomed Eng, 38, 537-549.  
19456124 E.Crowley, M.L.O'Mara, C.Reynolds, D.P.Tieleman, J.Storm, I.D.Kerr, and R.Callaghan (2009).
Transmembrane helix 12 modulates progression of the ATP catalytic cycle in ABCB1.
  Biochemistry, 48, 6249-6258.  
18415095 P.Lu, and M.G.Feng (2008).
Bifunctional enhancement of a beta-glucanase-xylanase fusion enzyme by optimization of peptide linkers.
  Appl Microbiol Biotechnol, 79, 579-587.  
16219697 S.Matsuzawa, M.Cuddy, T.Fukushima, and J.C.Reed (2005).
Method for targeting protein destruction by using a ubiquitin-independent, proteasome-mediated degradation pathway.
  Proc Natl Acad Sci U S A, 102, 14982-14987.  
15103154 R.K.Gaur, M.B.Kupper, R.Fischer, and K.M.Hoffmann (2004).
Preliminary X-ray analysis of a human V(H) fragment at 1.8 A resolution.
  Acta Crystallogr D Biol Crystallogr, 60, 965-967.  
11745490 R.Niv, Y.G.Assaraf, D.Segal, E.Pirak, and Y.Reiter (2001).
Targeting multidrug resistant tumor cells with a recombinant single-chain FV fragment directed to P-glycoprotein.
  Int J Cancer, 94, 864-872.  
11279614 Y.Heike, K.Kasono, C.Kunisaki, S.Hama, N.Saijo, T.Tsuruo, D.A.Kuntz, D.R.Rose, and D.T.Curiel (2001).
Overcoming multi-drug resistance using an intracellular anti-MDR1 sFv.
  Int J Cancer, 92, 115-122.  
11099853 A.Goel, D.Colcher, J.S.Koo, B.J.Booth, G.Pavlinkova, and S.K.Batra (2000).
Relative position of the hexahistidine tag effects binding properties of a tumor-associated single-chain Fv construct.
  Biochim Biophys Acta, 1523, 13-20.  
10570132 J.M.van Den Elsen, D.A.Kuntz, F.J.Hoedemaeker, and D.R.Rose (1999).
Antibody C219 recognizes an alpha-helical epitope on P-glycoprotein.
  Proc Natl Acad Sci U S A, 96, 13679-13684.
PDB code: 2ap2
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