Crystal structure of a chimeric Fab′ fragment of an antibody binding tumour cells

Brady, R.L.; Edwards, David J.; Hubbard, Roderick E.; Jiang, Jiahao; Lange, Gudrun; Roberts, Sally; Todd, R.; Adair, John R.; Emtage, J.S.; King, David J.; Low, D.C.

doi:10.1016/0022-2836(92)90695-g

Cited by 45 publications

(24 citation statements)

References 24 publications

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“…The structure was determined by molecular replacement with the program Phaser 45 . The search model of the Fab domain was derived from the published human IgG4 Fab crystal structure (PDB ID: 1BBJ) 46 , and that of the MG1 domain from the published human C5 crystal structure (PDB ID: 3CU7) 47 . A model was built with the Coot program 48 and refined with the program Refmac5 49 .…”

Section: Methodsmentioning

confidence: 99%

Long lasting neutralization of C5 by SKY59, a novel recycling antibody, is a potential therapy for complement-mediated diseases

Fukuzawa

Sampei

Haraya

et al. 2017

Sci Rep

112

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Dysregulation of the complement system is linked to the pathogenesis of a variety of hematological disorders. Eculizumab, an anti-complement C5 monoclonal antibody, is the current standard of care for paroxysmal nocturnal hemoglobinuria (PNH) and atypical hemolytic uremic syndrome (aHUS). However, because of high levels of C5 in plasma, eculizumab has to be administered biweekly by intravenous infusion. By applying recycling technology through pH-dependent binding to C5, we generated a novel humanized antibody against C5, SKY59, which has long-lasting neutralization of C5. In cynomolgus monkeys, SKY59 suppressed C5 function and complement activity for a significantly longer duration compared to a conventional antibody. Furthermore, epitope mapping by X-ray crystal structure analysis showed that a histidine cluster located on C5 is crucial for the pH-dependent interaction with SKY59. This indicates that the recycling effect of SKY59 is driven by a novel mechanism of interaction with its antigen and is distinct from other known pH-dependent antibodies. Finally, SKY59 showed neutralizing effect on C5 variant p.Arg885His, while eculizumab does not inhibit complement activity in patients carrying this mutation. Collectively, these results suggest that SKY59 is a promising new anti-C5 agent for patients with PNH and other complement-mediated disorders.

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Section: Methodsmentioning

confidence: 99%

Long lasting neutralization of C5 by SKY59, a novel recycling antibody, is a potential therapy for complement-mediated diseases

Fukuzawa

Sampei

Haraya

et al. 2017

Sci Rep

112

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“…2 a ELLIPS2 output for IgG antibody domain B72.3c Fab’ (see Carrasco et al 2001). b The axial ratios of ( a/b, b/c ) =1.60, 1.42) used for this example were obtained from the crystal structure of Brady et al (1992) using the algorithm of Taylor et al (1983), which fits a surface ellipsoid to crystallographic atomic co-ordinate data from the protein data bank (PDB)…”

Section: Evaluation Of Hydrodynamic Properties From the Axial Dimensimentioning

confidence: 99%

Hydrodynamic modelling of protein conformation in solution: ELLIPS and HYDRO

Torre

Harding

2013

Biophys Rev

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The last three decades has seen some important advances in our ability to represent the conformation of proteins in solution on the basis of hydrodynamic measurements. Advances in theoretical modeling capabilities have been matched by commensurate advances in the precision of hydrodynamic measurements. We consider the advances in whole-body (simple ellipsoid-based) modeling—still useful for providing an overall idea of molecular shape, particularly for those systems where only a limited amount of data is available—and outline the ELLIPS suite of algorithms which facilitates the use of this approach. We then focus on bead modeling strategies, particularly the surface or shell–bead approaches and the HYDRO suite of algorithms. We demonstrate how these are providing great insights into complex issues such as the conformation of immunoglobulins and other multi-domain complexes.

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“…These have elbow angles ranging from 1378 to 1458, an 88 variation. In the Fab for B72.3, the elbow angle was 1378, 8 the lower end of the range for HuCC49DCH2.…”

Section: Discussionmentioning

confidence: 94%

“…8 It is, therefore, available for comparison. No structures of Fab fragments from CC49 have been reported, but Tamura et al 23 have used homology rules to identify its six canonical hypervariable loops.…”

Section: Discussionmentioning

confidence: 99%

“…B72.3 is noteworthy because of the structural similarity to CC49 found here, and the fact that, because of its earlier discovery, 7 it has been the subject of more extensive studies and characterization, including an X-ray structure analysis of its antigen binding fragment. 8 The engineered antibody studied here, HuCC49DCH2 form B, is a chimeric antibody in which the complementarity-determining regions (CDR) of the Fabs are of mouse origin, while the remainder of the antibody is derived from a human antibody sequence. The C H 2 domain is entirely missing, and the C H 3 domain is linked directly to the Fabs through polypeptides, which are 20 amino acid residues in length.…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

The Structure of an Antitumor CH2-domain-deleted Humanized Antibody

Larson

Day

Glaser

et al. 2005

Journal of Molecular Biology

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C H 2-domain-deleted CC49 (HuCC49DCH2), a recombinant humanized antibody that recognizes the TAG-72 antigen expressed on a variety of human carcinomas, is secreted from cultured cells as a mixture of two homodimeric isoforms. Isoform A contains two covalent interchain disulfide bonds at heavy chain positions 239 and 242, while isoform B fails to develop any interchain disulfide bonds but has 239-242 intrachain disulfide bonds instead. Form A is currently in preclinical development as a therapeutic agent for treating colorectal carcinoma, though form B shows equal efficacy.HuCC49DCH2 form B can be crystallized from sodium formate only in the presence of detergents. X-ray diffraction data were collected on a single cryo-cooled crystal grown with Triton X-100 and the structure was solved by molecular replacement. The model has refined to RZ0.246 (R free Z0.297) for 2.8 Å data. The antibodies pack in the crystal around crystallographic 2-fold axes as tetramers with approximate 222 symmetry. Atomic force microscopy studies show that this tetrameric structure is the crystal building block and also exists free in the mother liquor. The tetramer is composed of two rings, back-to-back, with a thickness of w83 Å . Each ring is composed of two antibodies with the complementarity-determining regions (CDR) of the two Fabs of one antibody interacting with the CDR regions of the second antibody in a head-to-head fashion. These rings are approximately 167 Å long and 112 Å wide. The C H 3 domain is inverted with respect to the Fabs when compared to the usual orientation found in conventional antibodies. The polypeptides joining the C H 3 domains to the Fab portions of the antibody are not seen and are almost certainly disordered. The antigen combining site of HuCC49DCH2 is very similar, but not identical, in topology and charge distribution to that of antibody B72.3, which binds a similar epitope on TAG-72. The combining site consists of a deep cleft, heavily lined with aromatic amino acid side-chains but bounded by numerous charged groups.

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Crystal structure of a chimeric Fab′ fragment of an antibody binding tumour cells

Cited by 45 publications

References 24 publications

Long lasting neutralization of C5 by SKY59, a novel recycling antibody, is a potential therapy for complement-mediated diseases

Long lasting neutralization of C5 by SKY59, a novel recycling antibody, is a potential therapy for complement-mediated diseases

Hydrodynamic modelling of protein conformation in solution: ELLIPS and HYDRO

The Structure of an Antitumor CH2-domain-deleted Humanized Antibody

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