Purification, crystallization, X-ray diffraction analysis and phasing of a Fab fragment of monoclonal neuroantibody αD11 against nerve growth factor

• Published in: Acta crystallographica. Section D, Biological crystallography. Vol. 60; no. 7; pp. 1323 - 1327
• Main Authors: Covaceuszach, Sonia, Cassetta, Alberto, Cattaneo, Antonino, Lamba, Doriano
• Format: Journal Article
• Language: English
• Published: 5 Abbey Square, Chester, Cheshire CH1 2HU, England Munksgaard International Publishers 01.07.2004
• Subjects: nerve growth factor; neuroantibodies; αD11
• ISSN: 1399-0047; 0907-4449; 1399-0047
• DOI: 10.1107/S0907444904011485

The rat monoclonal neuroantibody αD11 is a potent antagonist that prevents the binding of nerve growth factor (NGF) to its tyrosine kinase A receptor (TrkA) in a variety of systems, most notably in two in vivo systems linked to crucial pathological states, such as Alzheimer's disease and HIV infection. To provide further insights into the mechanism of action of this potentially therapeutic monoclonal antibody, structural studies of the antigen‐binding fragment (Fab) of αD11 were performed. αD11 IgG2a immunoglobulin was obtained from hybridomas by in vitro tissue culture. The αD11 Fab crystallizes in two crystal forms. Form I belongs to space group P1, with unit‐cell parameters a = 42.7, b = 50.6, c = 102.7 Å, α = 82.0, β = 89.1, γ = 86.0°. With two molecules in the asymmetric unit, VM is 2.3 Å3 Da−1 and the solvent content is 46%. A complete data set has been collected at 2.7 Å resolution on beamline XRD‐1 (ELETTRA, Trieste, Italy). Form II belongs to space group C2, with unit‐cell parameters a = 114.8, b = 69.4, c = 64.10 Å, β = 117.0°. With one molecule in the asymmetric unit, VM is 2.4 Å3 Da−1 and the solvent content is 48%. A complete data set has been collected at 1.7 Å resolution on beamline ID14‐1 (ESRF, Grenoble, France). Phasing was successfully performed by Patterson search techniques and refinement of the structures is currently under way. Crystal forms I and II display a close‐packing pattern.