ABSTRACT
The second and final step in the synthesis of anticoagulant Mitrin 3 was catalyzed by 6-O sulfotransferase (6-OST) and 3-O sulfotransferase (3-OST) (Scheme 1). There are three heparan sulfate 6-O sulfotransferase isoforms: 6-OST1, 6-OST2 (6-OST2a and 6-OST2b are two splice variants), and 6-OST3 [15]. It is known that all three isoforms sulfate completely desulfated N-sulfated (CDSNS)–heparin equally well [15]. However, N-sulfoheparosan was shown to be preferentially sulfated by these isoforms in the following order: 6-OST2>6-OST3>6OST1. Thus, in essence, we have utilized the 6-OST2a isoform to catalyze the 6-O sulfation of glucosamine units. The 6-O sulfation was coupled with 3-O sulfation, which is catalyzed by 3-OST1 sulfotransferase to generate Mitrin anticoagulant [16]. There are as many as six isoforms of heparan sulfate 3-O sulfotransferases: 3-OST1, 3-OST2, 3OST3 (3-OST3a and 3-OST3b are two variants), 3-OST4, 3-OST5, and 3-OST6 [17,18]. It was demonstrated earlier that 3-OST1 is primarily involved in generating anticoagulant heparan sulfate [19]. It was also shown that 3-OST1 generally acts on glucosamine units flanked by the reducing side of GlcA and the nonreducing side of IdoA2S to generate ATIII binding structures containing GlcAGlcNS3S and GlcA-GlcNS3S6S [19-21]. Because 6-O sulfation and 3-O sulfation are not coupled to generate anticoagulant structures in vivo, it was anticipated that
Scheme 1 Enzymatic synthesis of Mitrin anticoagulants.
