Crystallization and functional analysis of a soluble deglycosylated form of the human costimulatory molecule B7-1

The interactions of B7-1 with CD28 and CTLA-4 modulate the course of human immune responses, making B7-1 an important target for developing structure- based therapeutics. B7-1 is, however, one of the most heavily glycosylated proteins found at the leukocyte cell surface, complicating the structural a nalysis of this molecule. Methods for the production, crystallization and s elenomethionine labelling of a soluble deglycosylated form of this molecule are described. The protein readily forms both tetragonal plate and bipyram idal crystals belonging to space groups I4(1)22, with unit-cell parameters a = b = 56.9, c = 298.7 Angstrom, and P4(1)22 (or P4(3)22), with unit-cell parameters a = b = 89.0, c = 261.9 Angstrom, respectively. The I4(1)22 and primitive crystal forms diffract to 2.7 and 3.5 Angstrom, respectively. Sur face plasmon resonance-based assays indicate that the ligand-binding proper ties of sB7-1 are unaffected by deglycosylation. Since none of the methods relied on any special structural properties of sB7-1, it is proposed that t his novel combination of procedures could in principle be adapted to the sy stematic analysis of many other glycoproteins of structural or functional i nterest.