1ap2
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SINGLE CHAIN FV OF C219
OverviewOverview
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.
About this StructureAbout this Structure
1AP2 is a Protein complex structure of sequences from Mus musculus. Full crystallographic information is available from OCA.
ReferenceReference
A single chain Fv fragment of P-glycoprotein-specific monoclonal antibody C219. Design, expression, and crystal structure at 2.4 A resolution., Hoedemaeker FJ, Signorelli T, Johns K, Kuntz DA, Rose DR, J Biol Chem. 1997 Nov 21;272(47):29784-9. PMID:9368049
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