4NQS

Knob-into-hole IgG Fc

Summary for 4NQS

• Entry DOI: 10.2210/pdb4nqs/pdb
• Related: 4NQT 4NQU
• Descriptor: Ig gamma-1 chain C region, miniZ, ... (4 entities in total)
• Functional Keywords: immunoglobulin, immune system
• Biological source: Homo sapiens (human); More
• Cellular location: Secreted: P01857 P01857
• Total number of polymer chains: 8
• Total formula weight: 113173.82

Authors

Eigenbrot, C.,Ultsch, M. (deposition date: 2013-11-25, release date: 2014-03-12, Last modification date: 2024-11-06)

Primary citation

Elliott, J.M.,Ultsch, M.,Lee, J.,Tong, R.,Takeda, K.,Spiess, C.,Eigenbrot, C.,Scheer, J.M.
Antiparallel Conformation of Knob and Hole Aglycosylated Half-Antibody Homodimers Is Mediated by a CH2-CH3 Hydrophobic Interaction.
J.Mol.Biol., 426:1947-1957, 2014

PubMed 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 2.64 Å. 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 K(D) of <1 nM. Homodimers of the "knob" and "hole" mutants were also obtained, and their X-ray structures were determined at resolutions 2.5 Å and 2.1 Å, 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.

PubMed: 24576605
DOI: 10.1016/j.jmb.2014.02.015

Experimental method

X-RAY DIFFRACTION (2.64 Å)