Sulfatase 1, also known as SULF1, is an enzyme which in humans is encoded by the SULF1 gene. Heparan sulfate proteoglycans (HSPGs) act as co-receptors for numerous heparin-binding growth factors and cytokines and are involved in cell signaling. Heparan sulfate 6-O-endo-sulfatases, such as SULF1, selectively remove 6-O-sulfate groups from heparan sulfate. This activity modulates the effects of heparan sulfate by altering binding sites for signaling molecules.
Function Heparan sulfate proteoglycans (HSPGs) are widely expressed throughout most tissues of nearly all multicellular species. The function of HSPGs extends beyond providing an extracellular matrix (ECM) structure and scaffold for cells. They are integral regulators of essential cell signaling pathways affecting cell growth, proliferation, differentiation, and migration. Although the core protein is important, the large heparan sulfate (HS) chains extending from the core are responsible for most receptor signaling. HS chains are heterogeneous structures that differ in specific and conditional cell contexts. Of particular importance is the HS sulfation pattern, which was once thought to be static after HS biosynthesis in the Golgi. However, this paradigm changed after the discovery of two extracellular 6-O-S glucosamine arylsulfatases, Sulf1 and Sulf2. These two enzymes allow rapid extracellular modification of sulfate content in HSPGs, impacting signaling involving Shh, Wnt, BMP, FGF, VEGF, HB-EGF, GDNF, and HGF. In addition, Sulfs may exercise another level of regulation over HS composition by down or upregulating HS biosynthetic enzymes present in the Golgi through the very same signaling pathways they modify.
Discovery 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. Validation of the predicted N-linked glycosylation sites on QSulf1 were performed using tunicamycin and QSulf1 variants missing the N-terminal (catalytic) domain or HD, which contain predicted N-linked glycosylation sites. The N- and C-terminal showed unbranched N-linked glycosylation, but was absent in the hydrophilic domain even though it contains two putative sites. In addition, O-linked or sialylated glycosylation were not present in QSulf1. Importantly, proper glycosylation is necessary to localize to the cell surface, possibly to bind HS moieties, and was required for enzymatic activity.
Structure and mechanism Sulf1 and Sulf2 are new members of a superfamily of arylsulfatases, being closely related to arylsulfatase A, B (ARSA; ARSB) and glucosamine 6-sulfatase (G6S). The x-ray crystal structure of neither Sulf1 or Sulf2 has been attempted, but ARSA active site crystal structure was deciphered. In ARSA, the conserved cysteine, which is posttranslationally modified to a C alpha formylglycine (FG) is critical for catalytic activity. In the first step, one of the two oxygens of the aldehyde hydrate attacks the sulfur of the sulfate ester. This leads to a transesterification of the sulfate group onto the aldehyde hydrate. Simultaneously the substrate alcohol is released. In the second step, sulfate is eliminated from the enzyme-sulfate intermediate by an intramolecular rearrangement. The “intramolecular hydrolysis” allows the aldehyde group to be regenerated. The active site of ARSA contains nine conserved residues that were found to be critical for catalytic activity. Some residues, such as Lys123 and Lys302, bind the substrate while others either participate in catalysis directly, such as His125 and Asp281, or indirectly. In addition a magnesium ion is needed to coordinate the oxygen that attacks the sulfur in the first step of sulfate cleavage. The crystal structure and residue mutations need to be performed in Sulf1 and Sulf2 to determine if any differences exist from lysosomal sulfatases.
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