PDBsum entry 2ap2

Immune system

PDB id: 2ap2

Contents

Protein chains

239 a.a. *

12 a.a. *

14 a.a. *

Waters ×64

* Residue conservation analysis

PDB id:

2ap2

Name:

Immune system

Title:

Single chain fv of c219 in complex with synthetic epitope peptide

Structure:

Single chain fv. Chain: a, c. Engineered: yes. P-glycoprotein. Chain: p, q. Fragment: atp-binding domain. Synonym: epitope peptide. Engineered: yes

Source:

Mus musculus. Mouse. Organism_taxid: 10090. Cellular_location: periplasmic. Expressed in: escherichia coli. Expression_system_taxid: 562. Synthetic: yes. Cricetulus griseus. Chinese hamster.

Biol. unit:

Tetramer (from PQS)

Resolution:

2.40Å R-factor: 0.221 R-free: 0.274

Authors:

J.M.H.Van Den Elsen,D.R.Rose

Key ref:

J.M.van Den Elsen et al. (1999). Antibody C219 recognizes an alpha-helical epitope on P-glycoprotein. Proc Natl Acad Sci U S A, 96, 13679-13684. PubMed id: 10570132 DOI: 10.1073/pnas.96.24.13679

Date:

22-Mar-99 Release date: 24-Nov-99

Protein chains

Q505N9  (Q505N9_MOUSE) -  Igh protein from Mus musculus

Seq: 468 a.a.

Struc: 239 a.a.*

Protein chains

Q6KB05  (Q6KB05_MOUSE) -  ScFv B8E5 protein (Fragment) from Mus musculus

Seq: 255 a.a.

Struc: 239 a.a.*

Protein chain

P21448  (MDR1_CRIGR) -  ATP-dependent translocase ABCB1 from Cricetulus griseus

Seq: 1276 a.a.

Struc: 12 a.a.

Protein chain

P21448  (MDR1_CRIGR) -  ATP-dependent translocase ABCB1 from Cricetulus griseus

Seq: 1276 a.a.

Struc: 14 a.a.

* PDB and UniProt seqs differ at 63 residue positions (black crosses)

Enzyme reactions

• Enzyme class 1: Chains P, Q: E.C.7.6.2.1 - P-type phospholipid transporter.
• Reaction: ATP + H2O + phospholipidSide 1 = ADP + phosphate + phospholipidSide 2
• Enzyme class 2: Chains P, Q: E.C.7.6.2.2 - ABC-type xenobiotic transporter.
• Reaction: ATP + H2O + xenobioticSide 1 = ADP + phosphate + xenobioticSide 2

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1073/pnas.96.24.13679

Proc Natl Acad Sci U S A 96:13679-13684 (1999)

PubMed id: 10570132

Antibody C219 recognizes an alpha-helical epitope on P-glycoprotein.

J.M.van Den Elsen, D.A.Kuntz, F.J.Hoedemaeker, D.R.Rose.

ABSTRACT

The ABC transporter, P-glycoprotein, is an integral membrane protein that mediates the ATP-driven efflux of drugs from multidrug-resistant cancer and HIV-infected cells. Anti-P-glycoprotein antibody C219 binds to both of the ATP-binding regions of P-glycoprotein and has been shown to inhibit its ATPase activity and drug binding capacity. C219 has been widely used in a clinical setting as a tumor marker, but recent observations of cross-reactivity with other proteins, including the c-erbB2 protein in breast cancer cells, impose potential limitations in detecting P-glycoprotein. We have determined the crystal structure at a resolution of 2.4 A of the variable fragment of C219 in complex with an epitope peptide derived from the nucleotide binding domain of P-glycoprotein. The 14-residue peptide adopts an amphipathic alpha-helical conformation, a secondary structure not previously observed in structures of antibody-peptide complexes. Together with available biochemical data, the crystal structure of the C219-peptide complex indicates the molecular basis of the cross-reactivity of C219 with non-multidrug resistance-associated proteins. Alignment of the C219 epitope with the recent crystal structure of the ATP-binding subunit of histidine permease suggests a structural basis for the inhibition of the ATP and drug binding capacity of P-glycoprotein by C219. The results provide a rationale for the development of C219 mutants with improved specificity and affinity that could be useful in antibody-based P-glycoprotein detection and therapy in multidrug resistant cancers.

Selected figure(s)

Figure 2.
Fig. 2. Molecular surface representation of the scFv C219 (molecule II) binding site with the bound -helical P-glycoprotein epitope peptide. The molecular surface is colored for electrostatic potential (red for negative charge, blue for positive charge). Peptide residues and the approximate locations of C219 heavy (H) and light chain (L) hypervariable loops are indicated. Fig. 2 was produced with the program GRASP (32).

Figure 3.
Fig. 3. Interactions between the -helical peptide and the C219 binding site. (A) Two-dimensional LIGPLOT (33) representation of the interactions between residues of the minimal NBD-epitope peptide (P), C219 heavy (H) and light chain (L) residues, and solvent molecules (S), as seen in molecule I. The residues that form van der Waals contacts with the peptide are depicted as labeled arcs with radial spokes pointing toward the peptide atoms with which they interact. C219 residues that form hydrogen bonds are shown in a ball-and-stick representation, and the hydrogen bonds are presented as dashed lines. Of all of the intrapeptide hydrogen bonds present in the structure, only the bonds between Gln 3P and Asp 7P are shown. (B) Stereoplot of the Fv-peptide interactions seen in molecule II. (C) Comparison of the bound NBD-epitope peptide in molecule I and II. In B and C, light (L) and heavy chain (H) residues and backbone positions of the scFv C219 are shown in green and magenta. Peptide backbone and side chains are shown in khaki for molecule I and in gold for molecule II. Positions of water molecules are indicated as red spheres. Different positions of binding site residues and water molecules in molecule I are also colored khaki. B and C were generated by using MOLSCRIPT (34) and RASTER3D (35).

Figures were selected by the author.