The primary structure of the insulin-like growth factor-binding protein (IGFBP) produced by the bovine kidney cell line, MDBK, has been deduced from the cDNA clone. The MDBK binding protein precursor consists of a hydrophobic pre-peptide of at least 26 amino acids and a mature protein of 284 amino acids. The predicted protein sequence shares extensive sequence similarity with both the rat (82%) and human (89%) IGFBP-2s, so that the MDBK binding protein is clearly the bovine counterpart of IGFBP-2. The protein has limited similarity with classes 1 (31%) and 3 (31%) human IGFBPs, except that all 18 cysteine residues are conserved. Other features deduced from the bovine IGFBP-2 cDNA include: an abundance of leucine in the pre-peptide, an Arg-Gly-Asp sequence, absence of N-linked glycosylation sites, and an imperfect polyadenylation signal as well as an ATTTA motif in the 3′ non-coding DNA. Western blotting indicated that this binding protein is widely distributed in bovine fluids as well as in media conditioned by bovine cell lines. Proteins immunologically related to bovine IGFBP-2 were detected not only in sheep, but also in chickens, indicating that this IGFBP is not exclusively mammalian.