ABSTRACT
The GD2 ganglioside is a cell-surface component that appears on the surface of metastatic melanoma cells and is a marker for the progression of the disease. The ME36.1 monoclonal antibody binds to the GD2 ganglioside and has shown potential as a therapeutic antibody. ME36.1 is a possible alternative therapy to radiation, which is often ineffective in late-stage melanoma. The crystal structure of the Fab fragment of ME36.1 has been determined using molecular replacement and refined to an R factor of 20.4% at 2.8 A resolution. The model has good geometry with root-mean-square deviations of 0.008 A from ideal bond lengths and 1.7 degrees from ideal bond angles. The crystal structure of the ME36.1 Fab shows that its complementarity determining region forms a groove-shaped binding site rather than the pocket-type observed in other sugar binding Fabs. Molecular modeling has placed a four-residue sugar, representative of GD2, in the antigen binding site. The GD2 sugar moiety is stabilized by a network of hydrogen bonds that define the specificity of ME36.1 toward its antigen.
Subject(s)
Antibodies, Monoclonal/chemistry , Gangliosides/immunology , Immunoglobulin Fab Fragments/chemistry , Melanoma/chemistry , Antibodies, Monoclonal/immunology , Binding Sites, Antibody , Carbohydrate Conformation , Carbohydrate Sequence , Crystallization , Crystallography, X-Ray , Gangliosides/chemistry , Hydrogen Bonding , Immunoglobulin Fab Fragments/immunology , Immunoglobulin Variable Region , Melanoma/immunology , Melanoma/therapy , Models, Molecular , Molecular Sequence Data , Molecular Structure , Protein Conformation , SoftwareABSTRACT
An Fab fragment from a monoclonal antibody (ME36.1) to the melanoma-associated GD2 ganglioside has been purified and crystallized in space group P2(1) with unit-cell dimensions a = 37.6, b = 94.1, c = 67.4 A, beta = 101.0 degrees. The crystals, which grow to a size of up to 0.6 x 0.5 x 0.3 mm, diffract to 2.5 A and native data have been collected to 2.8 A resolution. The crystal density is 1.22 g ml(-1) indicating one molecule of 48 kDa per asymmetric unit and a solvent content of 51%. A soluble form of the carbohydrate was obtained from the scarce GD2 glycolipid by enzymatic digestion with ceramide-glycanase. Small co-crystals of the Fab-GD2 complex have been obtained. As ME36.1 has been used in immunotherapy to treat malignant melanoma, knowledge of its interactions with the ganglioside could increase the efficacy of these treatments.
