1AP2

SINGLE CHAIN FV OF C219

Summary for 1AP2

• Entry DOI: 10.2210/pdb1ap2/pdb
• Descriptor: MONOCLONAL ANTIBODY C219 (3 entities in total)
• Functional Keywords: single chain fv, monoclonal antibody, c219, p-glycoprotein, immunoglobulin
• Biological source: Mus musculus (house mouse); More
• Total number of polymer chains: 4
• Total formula weight: 51739.21

Authors

Hoedemaeker, P.J.,Rose, D.R. (deposition date: 1997-07-23, release date: 1997-12-24, Last modification date: 2024-10-16)

Primary citation

Hoedemaeker, F.J.,Signorelli, T.,Johns, K.,Kuntz, D.A.,Rose, D.R.
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, 1997

PubMed 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.

PubMed: 9368049
DOI: 10.1074/jbc.272.47.29784

Experimental method

X-RAY DIFFRACTION (2.36 Å)