To define domains involved in IGF binding 60 N-terminal amino acid residues of IGFBP-1 were deleted. This deletion resulted in loss of IGF binding suggesting that the N-terminus may enclose an IGF binding domain. However, most point mutations introduced in this region did not affect IGF binding. In contrast to Cys-34, only substitution of Cys-38 for a tyrosine residue abolished IGF binding. With the determination that all 18 cysteine residues are involved in disulphide bond formation our data suggest that, although not all cysteines contribute to the same extent, the ligand binding site may be spatially organized.