Fab MOR03268 Triggers Absorption Shift of a Diagnostic Dye via Packaging in a Solvent-shielded Fab Dimer Interface

Hillig, R.C.; Urlinger, Stefanie; Fanghänel, Jörg; Brocks, Bodo; Haenel, Cornelia; Stark, Yvonne; Sülzle, Detlev; Svergun, Dmitri I.; Baesler, Siegfried; Malawski, G.; Moosmayer, Dieter; Menrad, Andreas; Schirner, Michael; Licha, Kai

doi:10.1016/j.jmb.2007.12.071

Cited by 9 publications

(6 citation statements)

References 27 publications

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“…Dimerization of Fab molecules in crystals is observed in cases where the antigen itself has internal twofold symmetry, as expected (20,21), but histone peptides described here do not form dimers or have internal symmetry. Antigen clasping is also distinct from the binding mode of the bispecific, chelating recombinant antibody (CRAb) because a CRAb is constructed by linking two different scFv molecules (22) and the two scFv units bind to nonoverlapping epitopes of a protein.…”

Section: Discussionsupporting

confidence: 73%

Antigen clasping by two antigen-binding sites of an exceptionally specific antibody for histone methylation

Hattori

Lai

Dementieva

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Antibodies have a well-established modular architecture wherein the antigen-binding site residing in the antigen-binding fragment (Fab or Fv) is an autonomous and complete unit for antigen recognition. Here, we describe antibodies departing from this paradigm. We developed recombinant antibodies to trimethylated lysine residues on histone H3, important epigenetic marks and challenging targets for molecular recognition. Quantitative characterization demonstrated their exquisite specificity and high affinity, and they performed well in common epigenetics applications. Surprisingly, crystal structures and biophysical analyses revealed that two antigen-binding sites of these antibodies form a head-to-head dimer and cooperatively recognize the antigen in the dimer interface. This "antigen clasping" produced an expansive interface where trimethylated Lys bound to an unusually extensive aromatic cage in one Fab and the histone N terminus to a pocket in the other, thereby rationalizing the high specificity. A long-neck antibody format with a long linker between the antigen-binding module and the Fc region facilitated antigen clasping and achieved both high specificity and high potency. Antigen clasping substantially expands the paradigm of antibody-antigen recognition and suggests a strategy for developing extremely specific antibodies.antibody engineering | epigenetics | antibody validation | protein-protein interaction | data reproducibility

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Section: Discussionsupporting

confidence: 73%

Antigen clasping by two antigen-binding sites of an exceptionally specific antibody for histone methylation

Hattori

Lai

Dementieva

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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“…Data were indexed and scaled to 2.8 Å resolution using HKL2000 [19]. The structure was solved by the molecular replacement method using Balbes [17] automatically using 2JB5 [20] for light chain, 3KDM [21] for heavy chain, 1B5L [22] for IFNα1b as search models. The initial phases were improved with OASIS [23].…”

Section: Structure Determination and Refinementmentioning

confidence: 99%

Structural insights into a human anti-IFN antibody exerting therapeutic potential for systemic lupus erythematosus

Ouyang

Gong

Li³

et al. 2012

J Mol Med

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Increasing evidences suggest that the type I interferon α (IFN α) plays a critical role in the etiopathogenesis of systemic lupus erythematosus (SLE), which makes it a promising therapeutic target for the treatment of the disease. By screening a large size non-immune human antibody library, we have developed a human single-chain antibody (ScFv) AIFN α 1bScFv01 and corresponding whole antibody AIFN α 1bIgG01 to human interferon α 1b (IFN α 1b) with high specificity and high affinity. The IgG antibody could down-regulate the expression of ISG15 and IFIT-1 induced by either recombinant IFN α 1b or naïve IFN α from SLE patients' sera, and reduced total serum IgG and IgM antibodies level in a pristane-primed lupus-like mouse model. The crystal structure of AIFN α 1bScFv01-IFN α 1b complex solved to 2.8 Å resolution revealed that both Pro26-Gln40 region in loop AB and Glu147-Arg150 region in helix E of IFN α 1b contribute to binding with AIFN α 1bScFv01. Four residues of above two regions (Leu30, Asp32, Asp35 and Arg150) are critical for the formation of antigen-antibody complexes. AIFN α 1bScFv01 shares partial epitopes of IFN α 1b with its receptor IFNAR2 but with much higher binding affinity to IFN α 1b than IFNAR2. Thus, AIFN α 1bIgG01 exhibits its neutralizing activity through competition with IFNAR2 to bind with IFN α and prevents the activation of IFN α-mediated signaling pathway. Our results highlight the potential use of the human antibody for modulating the activity of IFN α in SLE.

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“…Such a 2:1 binding stoichiometry in a Fab/epitope complex was described only recently by Hillig et al . for an antifluorescent dye antibody 31…”

Section: Discussionmentioning

confidence: 99%

The structure of the anti‐c‐myc antibody 9E10 Fab fragment/epitope peptide complex reveals a novel binding mode dominated by the heavy chain hypervariable loops

Krauß¹,

Wessner²,

Welfle³

et al. 2008

Proteins

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The X-ray structure of the Fab fragment from the anti-c-myc antibody 9E10 was determined both as complex with its epitope peptide and for the free Fab. In the complex, two Fab molecules adopt an unusual head to head orientation with the epitope peptide arranged between them. In contrast, the free Fab forms a dimer with different orientation. In the Fab/peptide complex the peptide is bound to one of the two Fabs at the "back" of its extended CDR H3, in a cleft with CDR H1, thus forming a short, three-stranded antiparallel beta-sheet. The N- and C-terminal parts of the peptide are also in contact with the neighboring Fab fragment. Comparison between the CDR H3s of the two Fab molecules in complex with the peptide and those from the free Fab reveals high flexibility of this loop. This structural feature is in line with thermodynamic data from isothermic titration calorimetry.

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Fab MOR03268 Triggers Absorption Shift of a Diagnostic Dye via Packaging in a Solvent-shielded Fab Dimer Interface

Cited by 9 publications

References 27 publications

Antigen clasping by two antigen-binding sites of an exceptionally specific antibody for histone methylation

Antigen clasping by two antigen-binding sites of an exceptionally specific antibody for histone methylation

Structural insights into a human anti-IFN antibody exerting therapeutic potential for systemic lupus erythematosus

The structure of the anti‐c‐myc antibody 9E10 Fab fragment/epitope peptide complex reveals a novel binding mode dominated by the heavy chain hypervariable loops

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