PDBsum entry 4nqt

Immune system

PDB id: 4nqt

Contents

Protein chain

208 a.a.

Waters ×106

PDB id:

4nqt

Name:

Immune system

Title:

Anti-parallel fc-hole(t366s/l368a/y407v) homodimer

Structure:

Ig gamma-1 chain c region. Chain: a. Fragment: unp residues 118-330. Synonym: hole fc t366s/l368a/y407v. Engineered: yes. Mutation: yes

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: ighg1. Expressed in: escherichia coli. Expression_system_taxid: 562

Resolution:

2.10Å R-factor: 0.198 R-free: 0.253

Authors:

C.Eigenbrot,M.Ultsch

Key ref:

J.M.Elliott et al. (2014). Antiparallel conformation of knob and hole aglycosylated half-antibody homodimers is mediated by a CH2-CH3 hydrophobic interaction. J Mol Biol, 426, 1947-1957. PubMed id: 24576605 DOI: 10.1016/j.jmb.2014.02.015

Date:

25-Nov-13 Release date: 12-Mar-14

Protein chain

P01857  (IGHG1_HUMAN) -  Immunoglobulin heavy constant gamma 1 from Homo sapiens

Seq: 399 a.a.

Struc: 208 a.a.*

* PDB and UniProt seqs differ at 5 residue positions (black crosses)

DOI no: 10.1016/j.jmb.2014.02.015

J Mol Biol 426:1947-1957 (2014)

PubMed id: 24576605

Antiparallel conformation of knob and hole aglycosylated half-antibody homodimers is mediated by a CH2-CH3 hydrophobic interaction.

J.M.Elliott, M.Ultsch, J.Lee, R.Tong, K.Takeda, C.Spiess, C.Eigenbrot, J.M.Scheer.

ABSTRACT

Bispecific antibody and antibody-like molecules are of wide interest as potential therapeutics that can recognize two distinct targets. Among the variety of ways such molecules have been engineered is by creating "knob" and "hole" heterodimerization sites in the CH3 domains of two antibody heavy chains. The molecules produced in this manner maintain their biological activities while differing very little from the native human IgG sequence. To better understand the knob-into-hole interface, the molecular mechanism of heterodimerization, and to engineer Fc domains that could improve the assembly and purity of heterodimeric reaction products, we sought crystal structures of aglycosylated heterodimeric and homodimeric "knob" and "hole" Fc fragments derived from bacterial expression. The structure of the knob-into-hole Fc was determined at 2.64Å. Except for the sites of mutation, the structure is very similar to that of the native human IgG1 Fc, consistent with a heterodimer interaction kinetic KD of <1nM. Homodimers of the "knob" and "hole" mutants were also obtained, and their X-ray structures were determined at resolutions 2.5Å and 2.1Å, respectively. Both kinds of homodimers adopt a head-to-tail quaternary structure and thus do not contain direct knob/knob or hole/hole CH3 interactions. The head-to-tail arrangement was disfavored by adding site-directed mutations at F241 and F243 in the CH2 domains, leading to increases in both rate and efficiency of bispecific (heterodimer) assembly.