CRYSTAL STRUCTURE OF A MURINE MONO-CLONAL ANTI-ELAM IGG1 ANTIBODY 7A9 Fab FRAGMENT Adela Rodriguez-Romero,1 Orna Almog,1 Maria Tordova,1 Zafar Randawa,2 and Gary L. Gilliland.1 1Center for Advanced Research in Biotechnology of the University of Maryland Biotechnology Institute and National Insititute of Standards and Technology, 9600 Gudelsky Dr., Rockville, MD 20850. 2Otsuka America Pharmaceutical, Inc., 9900 Medical Center Dr., Rockville, MD 20850.
Neutrophils, involved in the early stages of many forms of acute inflammation, can cause disease by damaging normal host tissue, as in the Adult Respiratory Distress Syndrome (ARDS). In this process, cell surface receptors such as ELAM-1 (endothelial leukocyte adhesion molecule) react with a carbohydrate residue on the cell surface of the neutrophil, while the leukocyte integrins containing a CD18 antigen react with ICAM-1 (intercellular adhesion molecule-1). The 7A9 antibody effectively binds to ELAM-1 blocking neutrophil accumulation in the lungs [Mulligen et al., (1991) J. Clin. Invest. 88, 1396-1406]. The three-dimensional structure of the 7A9 Fab was undertaken to further our understanding of how this antibody binds to ELAM-1. Crystals of the protein (0.2 x 0.3 x0.5 mm) were grown in vapor diffusion experiments at room temperature in the presence of 20% PEG (8K), 20 mM ammonium sulfate, 100 mM cacodylate buffer at pH 6.5. The orthorhombic crystals diffract to 2.8 and have space group P212121. The unit cell parameters are a = 44.5 , b = 83.8 , and c = 132.5 . The structure was determined by the molecular replacing technique using the program XPLOR. The probe molecule, the Fab fragment of an antibody directed against the rhinovirus [Tormo et al., (1992) Protein Sci. 1, 1154-1161], was selected because of its sequence homology with the 7A9 antibody. The structure has been refined using XPLOR to a current R factor of 0.18. Details of the structure determination, a description of the structure, and a comparison of the structure with those of related Fabs will be presented.