PURIFICATION AND CHARACTERIZATION OF A UTERINE RETINOL-BINDING PROTEIN IN THE BITCH

A major canine endometrial secreted protein (cP6, 23000-M(t)) was purified by ion-exchange and gel-filtration chromatography and characterized by two-dimensional gel electrophoresis. Anti[human retinol-binding protein (hRBP)] serum identified cP6 on immunoblot analysis and immunoprecipitated cP6 from culture medium. This major protein was also shown to bind [H-3]retinol. N-terminal and internal amino acid sequences were determined and compared with previously identified protein, RNA, or DNA sequences. N-terminal analysis revealed that cP6 had high identity and similarity to serum retinol-binding proteins (RBPs), while internal sequence analysis showed a strong similarity to rat androgen-dependent epididymal protein and beta-lactoglobulins. Amino acid analysis, however, showed significant differences between these proteins and cP6 in both total amino acid content and certain selected amino acids. Immunohistochemical analysis showed staining for REP only in the uterine luminal epithelium. These studies suggest that bitch endometrium secretes a family of proteins (cP6), some of which bind [H-3]retinol, are immunologically related to the REP family, and have N-terminal and internal sequences with a high similarity to REP, beta-lactoglobulins and other members of the lipocalin family. This family of proteins may be important in early development for supplying retinol or derivatives to the developing embryo.