PDBsum entry: 1cqk

Immune system

PDB id: 1cqk

Contents

Protein chains

101 a.a. *

Waters ×77

* Residue conservation analysis

PDB id:

1cqk

Name:

Immune system

Title:

Crystal structure of the ch3 domain from the mak33 antibody

Structure:

Ch3 domain of mak33 antibody. Chain: a, b. Fragment: ch3 domain of mak33 antibody. Mutation: yes

Source:

Mus musculus. House mouse. Organism_taxid: 10090

Biol. unit:

Dimer (from PQS)

Resolution:

2.20Å R-factor: 0.196 R-free: 0.249

Authors:

M.J.Thies,J.Mayer,J.G.Augustine,C.A.Frederick,H.Lilie, J.Buchner

Key ref:

M.J.Thies et al. (1999). Folding and association of the antibody domain CH3: prolyl isomerization preceeds dimerization. J Mol Biol, 293, 67-79. PubMed id: 10512716 DOI: 10.1006/jmbi.1999.3128

Date:

06-Aug-99 Release date: 11-Sep-99

Protein chain

No UniProt id for this chain

DOI no: 10.1006/jmbi.1999.3128

J Mol Biol 293:67-79 (1999)

PubMed id: 10512716

Folding and association of the antibody domain CH3: prolyl isomerization preceeds dimerization.

M.J.Thies, J.Mayer, J.G.Augustine, C.A.Frederick, H.Lilie, J.Buchner.

ABSTRACT

The simplest naturally occurring model system for studying immunoglobulin folding and assembly is the non-covalent homodimer formed by the C-terminal domains (CH3) of the heavy chains of IgG. Here, we describe the structure of recombinant CH3 dimer as determined by X-ray crystallography and an analysis of the folding pathway of this protein.Under conditions where prolyl isomerization does not contribute to the folding kinetics, formation of the beta-sandwich structure is the rate-limiting step. beta-Sheet formation of CH3 is a slow process, even compared to other antibody domains, while the subsequent association of the folded monomers is fast. After long-time denaturation, the majority of the unfolded CH3 molecules reaches the native state in two serial reactions, involving the re-isomerization of the Pro35-peptide bond to the cis configuration. The species with the wrong isomer accumulate as a monomeric intermediate. Importantly, the isomerization to the correct cis configuration is the prerequisite for dimerization of the CH3 domain. In contrast, in the Fab fragment of the same antibody, prolyl isomerization occurs after dimerization demonstrating that within one protein, comprised of highly homologous domains, both the kinetics of beta-sandwich formation and the stage at which prolyl isomerization occurs during the folding process can be completely different.

Selected figure(s)

Figure 1.
Figure 1. Diagram and GRASP model of the C[H]3 domain. (a) Overall dimeric model of C[H]3. The tryptophan and cis-proline residues are in ball-and-stick representation. The disulfide bridge connecting the sheets of each monomer is highlighted in yellow. This Figure was generated in SETOR [Evans 1993]. (b) Interface of the C[H]3 dimer. The residues interacting within the dimeric interface are indicated in stick representation. This Figure was generated in SETOR [Evans 1993]. (c) A representation of a C[H]3 monomer with the dimerization interface in white. (d) A surface representation of the dimeric interface of a C[H]3 monomer showing positive (in blue), negative (in red) and neutral (in white) charged surfaces. This model was made using GRASP [Nicholls and Honig 1991].

Figure 2.
Figure 2. GdmCl-dependent unfolding and refolding transitions of C[H]3. Samples were incubated for six days at 20°C in 0.1 M Tris-HCl (pH 8.0) and varying GdmCl concentrations. For the fluorescence measurements the excitation wavelength was set to 280 nm. Spectra were recorded from 290 to 400 nm and analyzed at 355 nm. The C[H]3 dimer concentration was 0.2 µM (0m), 1.6 µM ( open ) and 4.1 µM ( up triangle, open ), respectively. For the CD experiment, the ellipticity was recorded at 213 nm ( triangle, filled ) at a protein concentration of 4.1 µM. All measurements were carried out at 20°C.

The above figures are reprinted by permission from Elsevier: J Mol Biol (1999, 293, 67-79) copyright 1999.

Figures were selected by an automated process.