Before the cloning and characterization of Sulf1 and Sulf2, HS composition was thought to be unchanging after localization to the cell surface. However, this changed when the quail orthologue of Sulf1, QSulf1, was identified in a screen for Sonic hedgehog (Shh) response genes activated during somite formation in quail embryos. Sequence alignment analysis indicates QSsulf1 is homologous with lysosomal N-acetyl glucosamine sulfatases (G6-sulfatases) that catalyze the hydrolysis of 6-O sulfates from N-acetyl glucosamines of heparan sulfate during the degradation of HSPGs. In contrast to lysosomal active sulfatases, QSulf1 localizes exclusively to the cell surface by interacting hydrophilically with a non-heparan sulfate outer membrane component, and is enzymatically active at a neutral pH. By mutating the catalytically active cysteines to alanine, thereby blocking N-formylglycine formation, they found QSulf1 was responsible for Wingless (Wnt) release from HS chains to activate the Frizzled receptor; this was the first evidence that an extracellular sulf was capable of modifying HS and therefore cell signaling. The overall structure of QSulf is followed closely by its orthologues and paralogues, including human and mouse. The human and murine orthologues of QSulf1, HSulf1 and MSulf1, respectively, were cloned and characterized after the discovery of QSulf1. In addition, a paralogue, Sulf2, sharing 63-65% identity (both mouse and human) with Sulf1 also was discovered through BLAST sequence analysis. The HSulf1 gene (GenBank accession number AY101175) has an open reading frame of 2616 bp, encoding a protein of 871 amino acid (aa), and HSulf2 (GenBank accession number AY101176) has an open reading frame of 2613 bp, encoding a protein of 870 aa. The HSulf1 and 2 genes localize to 8q13.2-13.3 and 20q13.12, respectively. They contain putative Asn-linked glycosylation sites, and furin cleavage sites responsible for proteolytic processing in the Golgi. The function or substrate specificity these cleavage sites impart has yet to be determined.
