The occurrence of three isoforms of heparan sulfate 6-O-sulfotransferase having different specificities for hexuronic acid adjacent to the targeted N-sulfoglucosamine

We previously cloned heparan sulfate 6-O-sulfotransferase (HS6ST) (Habuchi, H., Kobayashi, M., and Kimata, K. (1998) J. Biol. Chem. 273, 9208-9213). I n this study, we report the cloning and characterization of three mouse iso forms of HS6ST, a mouse homologue to the original human HS6ST (HS6ST-1) and two novel HS6STs (HS6ST-2 and HS6ST-3). The cDNAs have been obtained from mouse brain cDNA library by cross-hybridization with human HS6ST cDNA The t hree cDNAs contained single open reading frames that predicted type II tran smembrane proteins composed of 401, 506, and 470 amino acid residues, respe ctively. Amino acid sequence of HS6ST-1 was 51 and 57% identical to those o f HS6ST-2 and HS6ST-3, respectively. HS6ST-2 and HS6ST-3 had the 50% identi ty. Overexpression of each isoform in COS-7 cells resulted in about 10-fold increase of HS6ST activity, The three isoforms purified with anti-FLAG ant ibody affinity column transferred sulfate to heparan sulfate and heparin bu t not to other glycosaminoglycans. Each isoform showed different specificit y toward the isomeric hexuronic acid adjacent to the targeted N-sulfoglucos amine; HS6ST-1 appeared to prefer the iduronosyl N-sulfoglucosamine while H S6ST-2 had a different preference, depending upon the substrate concentrati ons, and HS6ST-3 acted on either substrate. Northern analysis showed that t he expression of each message in various tissues was characteristic to the respective isoform. HS6ST-1 was expressed strongly in liver, and HS6ST-2 wa s expressed mainly in brain and spleen. In contrast, HS6ST-3 was expressed rather ubiquitously. These results suggest that the expression of these iso forms may be regulated in tissue specific manners and that each isoform may be involved in the synthesis of heparan sulfates with tissue-specific stru ctures and functions.