PDBsum entry 1t04

Immune system

PDB id: 1t04

Contents

Protein chains

214 a.a. *

219 a.a. *

Waters ×47

* Residue conservation analysis

PDB id:

1t04

Name:

Immune system

Title:

Three dimensional structure of a humanized anti-ifn-gamma fab in c2 space group

Structure:

Huzaf antibody light chain. Chain: a, c. Synonym: fab. Engineered: yes. Other_details: the source is engineered human constant and framework regions with cdrs from mouse.. Huzaf antibody heavy chain. Chain: b, d. Engineered: yes.

Source:

Homo sapiens. Human. Organism_taxid: 9606. Organism_taxid: 9606

Biol. unit:

Dimer (from PQS)

Resolution:

3.00Å R-factor: 0.217 R-free: 0.293

Authors:

P.C.Bourne,S.S.Terzyan,G.Cloud,N.F.Landolfi,M.Vasquez,A.B.Edmundson

Key ref:

P.C.Bourne et al. (2004). Three-dimensional structures of a humanized anti-IFN-gamma Fab (HuZAF) in two crystal forms. Acta Crystallogr D Biol Crystallogr, 60, 1761-1769. PubMed id: 15388922 DOI: 10.1107/S0907444904018670

Date:

07-Apr-04 Release date: 05-Oct-04

Protein chains

Q6GMW1  (Q6GMW1_HUMAN) -

Protein chains

No UniProt id for this chain

Struc: 219 a.a.

DOI no: 10.1107/S0907444904018670

Acta Crystallogr D Biol Crystallogr 60:1761-1769 (2004)

PubMed id: 15388922

Three-dimensional structures of a humanized anti-IFN-gamma Fab (HuZAF) in two crystal forms.

P.C.Bourne, S.S.Terzyan, G.Cloud, N.F.Landolfi, M.Vásquez, A.B.Edmundson.

ABSTRACT

Three-dimensional structures were determined for two crystal forms (orthorhombic P2(1)2(1)2(1) and monoclinic C2) of the Fab from the humanized version of a murine monoclonal antibody (AF2) that possesses binding and potent neutralizing activity against human interferon gamma (IFN-gamma). This humanized antibody (HuZAF; USAN name fontolizumab) is currently in phase II clinical trials for the treatment of Crohn's disease. HuZAF exhibits binding and IFN-gamma neutralizing capacities that closely approximate those of the original antibody. It is shown that HuZAF, whose VH domain was designed using a best-sequence-fit approach, is closer structurally to its mouse precursor than is a version whose VH was constructed using a human sequence with lower homology to the original mouse sequence. This work thus offers direct structural evidence in support of the best-sequence-fit approach and adds to previous results of biological and biochemical evaluations of distinctly engineered antibodies that also favored the use of a best-sequence-fit strategy. A second crystal type appeared during attempts to crystallize the Fab-IFN-gamma complex. The antibody-antigen complex that existed in solution dissociated in the crystallization mixture. A conformationally altered but unliganded HuZAF protein crystallized in a different space group (C2), with two Fab molecules in the asymmetric unit. In this crystal lattice, no space was available for accommodating the IFN-gamma antigen. Thus, there are currently three slightly different structures of the HuZAF Fab.

Selected figure(s)

Figure 2.
Figure 2 Stereo illustration of the three-dimensional structure of the HuZAF Fab, prepared with the program MOLSCRIPT (Kraulis, 1991[174] [Kraulis, P. J. (1991). J. Appl. Cryst. 24, 946-950.]-[175][bluearr.gif] ). Secondary elements are superimposed on backbone tracings, which are colored yellow in the L chain and steel blue in the H chain. [176][beta] -Strands are represented by directional arrows: cyan for the four-stranded [177][beta] -pleated sheets and magenta for the five-stranded (VL) and three-stranded layers (CL) of the L chain and red and white for the corresponding [178][beta] -pleated sheets in the H chain. Helices are identified by red and yellow spirals. CDRs are marked 1, 2 and 3.

Figure 3.
Figure 3 (a) Polypeptide tracings of the VH and VL domains of the HuAF2 Fab (PDB code [187]1b4j ) in red, superimposed on those of the murine domains in the mouse-human chimeric ChiAF2 Fab (PDB code [188]1b2w ) in white. CDRs in the HuAF2 have amino-acid sequences excerpted from those in the original murine AF2 antibody. (b) Backbone tracings of HuZAF V domains (cyan) overlaid on those of ChiAF2 (Maloney et al., 1997[189] [Maloney, D. G., Grillo-Lopez, A. J., White, C. A., Bodkin, D., Schilder, R. J., Neidhart, J. A., Janakiraman, N., Foon, K. A., Liles, T. M., Dallaire, B. K., Wey, K., Royston, I., Davis, T. & Levy, R. (1997). Blood, 90, 2188-2195.]-[190][bluearr.gif] ). In HuZAF, the substitution of two acceptor framework residues with mouse residues, Val11 by leucine and Arg38 by lysine, significantly improves the structural mimicry relative to ChiAF2.

The above figures are reprinted by permission from the IUCr: Acta Crystallogr D Biol Crystallogr (2004, 60, 1761-1769) copyright 2004.

Figures were selected by an automated process.