Biosynthesis of the carbohydrate antigenic determinants, Globo H, blood group H, and Lewis b: A role for prostate cancer cell α1,2-L-fucosyltransferase

Prostate carcinoma LNCaP cells were unique among several human cancer cell lines which include two other prostate cancer cell lines, PC-3 and DU-145, in expressing α1,2-L-fucosyltransferase (FT) as an exclusive FT activity. Affinity gel-GDP and Sephacryl S100 HR columns were used for a partial purification of this enzyme from 3.9 × 10⁹ LNCaP cells (∼200-fold; 40% yield). The K_m value (2.7 mM) for the LacNAc type 2 acceptor was quite similar to the one reported for the cloned blood group H gene-specified α1,2-FT [Chandrasekaran et al. (1996) Biochemistry 35, 8914-8924]. N-Ethylmaleimide was a potent inhibitor (K_i 12.5 μM). The enzyme showed four-fold acceptor preference for the LacNAc type 2 unit in comparison to the T-hapten in mucin core 2 structure. Its main features were similar to those of the cloned enzyme: (1) C-6 sulfation of terminal Gal in the LacNAc unit increased the acceptor efficiency, whereas C-6 sialylation abolished acceptor ability; (2) C-6 sulfation of GlcNAc in LacNAc type 2 decreased by 80% the acceptor ability, whereas LacNAc type 1 was unaffected; (3) Lewis x did not serve as an acceptor; (4) the C-4 hydroxyl rather than the C-6 hydroxyl group of the GlcNAc moiety in LacNAc type1 was essential for activity; and (5) the acrylamide copolymer of Galβ1,3GlcNAcβ-O-Al was the best acceptor among the acrylamide copolymers. Additionally, highly significant biological features of α1,2FT were identified in the present study. The synthesis of Globo H and Lewis b determinants became evident from the fact that Galβ1,3GalNAcβ1,3Galα-O-Me and Galβ1,3(Fucα1,4)Glc-NAcβ1,3Galβ-O-Me served as high-affinity acceptors for this enzyme. Further, D-Fucβ1,3Gal-NAcβ1,3Galα-O-Me was a very efficient acceptor, indicating that the C-6 hydroxyl group of the terminal Gal moiety in Globo H is not essential for the enzyme activity. Thus, the present study was able to demonstrate three different catalytic roles of LNCaP α1,2-FT, namely, the expressions of blood group H, Lewis b from Lewis a, and Globo H.