Biosynthesis of sulfated glycoprotein in the endometrium of rabbit uterus

The endometrial scrapings obtained from the uteri of estrogen-treated rabbits were incubated with N-acetyl-d[1-3H]glucosamine and [35S]sulfate, and then the incubation medium (M-Fr) was separated from the tissue. The tissue was subsequently homogenized exhaustively in 0.25m sucrose, and the insoluble residue (R-Fr) was separate. The supernatant at 8,500Xg for 10 min of the homogenate was subjected to subcelular fractionation by discontinuous sucrose gradient ultracentrifugation, and a thiamine pyrophosphatase-rich fraction (g-fr) was obtained. Complex carbohydrates were then separated from M-Fr, R-Fr, and G-Fr. The radioactivities incorporated into these complex carbohydrates suggested that sulfated glycoprotein synthesized in G-Fr was secreted into M-Fr. In order to confirm the above observation, labelled sulfated glycoprotein was isolated from the incubation medium. Subsequently, N-ACETYL-D[1-3H]glucosamine was incorporated into N-acetylglucosamine residues and [35S]sulfate into sulfates located most probably at the 6-position of N-acetylglucosamine residues of sulfated glycoprotein.