| 1999 |
HS3ST1 (3-OST-1) encodes heparan sulfate D-glucosaminyl 3-O-sulfotransferase-1, a type II integral membrane protein with a self-contained C-terminal sulfotransferase domain (~260 residues) and a divergent N-terminal region. Expression in COS-7 cells confirmed 3-O-sulfation of specific glucosaminyl residues within heparan sulfate and conversion of non-anticoagulant to anticoagulant heparan sulfate (HSact). 3-OST-1 showed 300-fold greater HSact conversion activity than 3-OST-2 and 3-OST-3A isoforms. |
cDNA expression in COS-7 cells, radiolabeled PAPS sulfotransferase assay, disaccharide analysis, Northern blot |
The Journal of biological chemistry |
High |
9988767 9988768
|
| 2001 |
The sequence-specific properties of 3-OST-1 (preferential generation of antithrombin-binding sites) reside entirely within the sulphotransferase domain and are not influenced by the divergent N-terminal region. Domain-swap chimeras (N-terminal of 3-OST-3A fused to sulphotransferase domain of 3-OST-1, and vice versa) demonstrated that the sulphotransferase domain alone determines isoform-specific substrate selectivity. |
cDNA domain-swap chimera expression in COS-7 cells, antithrombin-binding site generation assay, HSV-1 entry assay in CHO transfectants |
The Biochemical journal |
High |
11563988
|
| 2000 |
3-OST-1 enzymatically modifies heparan sulfate immobilized on a biochip by introducing a 3-O-sulfo group, converting low-affinity antithrombin III (ATIII) binding to high-affinity binding, as detected by surface plasmon resonance. This demonstrates the specific structural modification required for ATIII recognition. |
Surface plasmon resonance (SPR) on streptavidin biochip with biotinylated heparan sulfate, enzymatic modification with 3-OST-1 and PAPS |
Biochemical and biophysical research communications |
Medium |
11006120
|
| 2002 |
Recombinant human 3-OST-1 expressed in E. coli is a soluble, active enzyme with Km values in the low micromolar range (KmHS = 4.3 µM; KmPAPS = 38.6 µM) and Vmax of ~18–21 pmol sulfate/min/pmol enzyme, comparable to baculovirus-expressed enzyme. This established the kinetic parameters of the catalytic mechanism. |
Expression in E. coli, purification, radiochemical in vitro sulfotransferase kinetic assay with [35S]PAPS |
Biochemical and biophysical research communications |
Medium |
11811991
|
| 2002 |
Structure-based homology modeling of 3-OST-1 identified key structural motifs and specific amino acids predicted to be important for enzymatic function and substrate recognition, providing a structural framework for the 3-O-sulfotransferase catalytic mechanism. |
Computational homology modeling using crystallographic data of related enzymes, structural analysis tools |
Biochemical and biophysical research communications |
Low |
11811992
|
| 2001 |
6-O-sulfotransferase-1 (6-OST-1) is a limiting enzyme in the anticoagulant heparan sulfate (HSact) biosynthetic pathway downstream of 3-OST-1. In CHO cells expressing 3-OST-1, a mutant deficient in 6-O-sulfation failed to produce HSact; transfection with 6-OST-1 rescued HSact production to ~50%. Both 3-O- and 6-O-sulfation can each be the terminal step in HSact biosynthesis. |
Retroviral transduction of 3-OST-1 into CHO cells, chemical mutagenesis screen, 6-OST-1 transfection rescue, in vitro modification with purified 6-OST-1, capillary HPLC-MS disaccharide analysis |
The Journal of biological chemistry |
High |
11551899
|
| 2003 |
Hs3st1 knockout mice are devoid of 3-OST-1 enzyme activity in plasma and tissues, showing dramatic reductions in tissue HSact. However, hemostasis is normal (normal fibrin accumulation, normal carotid artery occlusion times, normal thrombin-antithrombin complexes). Instead, Hs3st1−/− mice exhibit genetic background-specific lethality and intrauterine growth retardation, indicating that bulk HSact is dispensable for normal hemostasis and that 3-OST-1-derived structures serve alternative biological roles. |
Hs3st1 knockout mouse generation, enzyme activity assay in plasma and tissue extracts, ferric chloride carotid artery injury assay, tissue fibrin accumulation under normoxic and hypoxic conditions, thrombin-antithrombin complex measurement |
The Journal of clinical investigation |
High |
12671048 12975616
|
| 2012 |
Crystal structure of the ternary complex of 3-OST-1 with 3'-phosphoadenosine 5'-phosphate (PAP) and a heptasaccharide substrate was determined. Comparison to 3-OST-3 structures revealed distinct substrate-binding modes and saccharide conformations for each isoform. Site-directed mutagenesis identified Arg268 in 3-OST-1 (and Lys259, Thr256, Trp283 in 3-OST-3) as key residues for substrate binding and isoform specificity. |
X-ray crystallography (ternary complex), site-directed mutagenesis, structural comparison to 3-OST-3 crystal structures |
Proceedings of the National Academy of Sciences of the United States of America |
High |
22431632
|
| 2013 |
Golgi-targeted HS3ST1 localizes to the Golgi and produces a single type of antithrombin (AT)-binding site in CHO cells with high anti-factor Xa activity (137 ± 36 units/mg). Untargeted HS3ST1 is broadly distributed throughout CHO cells and produces no detectable AT-binding sites. Stable HS3ST1 overexpression also upregulates 2-O-, 6-O-, and N-sulfo group-containing disaccharides, revealing a concerted interplay between HS biosynthetic enzymes. |
Stable CHO cell transfection with Golgi-targeted vs. untargeted HS3ST1, anti-factor Xa anticoagulant activity assay, immunofluorescence localization, disaccharide composition analysis by AMAC-LCMS |
The Journal of biological chemistry |
High |
24247246
|
| 2017 |
3-O-sulfation of heparan sulfate catalyzed by Hs3st1 enhances binding of neuropilin-1 to heparan sulfate. 3-O-sulfated heparan sulfate dodecamers inhibited neuropilin-1-dependent semaphorin-3a-induced growth cone collapse of neurons and enhanced inhibition of endothelial cell sprouting. The effect was specific to Hs3st2 (not Hs3st1) for growth cone collapse in neurons from isoform-specific knockout mice. |
Affinity chromatography with 3-O-sulfated HS columns, SPR binding assay, thermal denaturation protection assay, growth cone collapse assay with Hs3st1−/− and Hs3st2−/− neuron cultures, endothelial cell sprouting assay |
ACS chemical biology |
High |
26731579
|
| 2017 |
In Hs3st1−/− mice, antithrombin (AT) treatment during LPS-induced septic shock induced pro-inflammatory rather than anti-inflammatory effects: increased LPS-lethality, increased leukocyte firm adhesion to endothelium, and vasoconstriction of coronary arterioles (opposite of effects in wild-type mice). In humans, the rs16881446G allele in HS3ST1 was associated with reduced HS3ST1 expression in primary endothelial cells and with increased severity of coronary artery disease. This establishes that the primary function of HS3ST1-derived HSAT+ is to mediate AT's anti-inflammatory activity. |
Hs3st1−/− mouse LPS challenge model, intravital microscopy for leukocyte adhesion, ex vivo coronary arteriole dilation assay, human candidate-gene association study in >2000 patients, eQTL analysis in primary endothelial cells |
Matrix biology : journal of the International Society for Matrix Biology |
High |
28126521
|
| 2019 |
3-O-sulfation of heparan sulfate (catalyzed by HS3ST1/3-OST-1) significantly enhances tau binding to heparan sulfate. In Hs3st1−/− cells, reduced 3-O-sulfation diminished both cell-surface binding and internalization of tau. NMR titrations mapped 3-O-S binding sites on tau to the microtubule binding repeat 2 (R2) and proline-rich region 2 (PRR2) domains. |
HS microarray, SPR binding assay with structurally defined oligosaccharides, Hs3st1−/− cell-based tau binding and uptake assay, NMR titration |
Angewandte Chemie (International ed. in English) |
High |
31692167
|
| 2020 |
ZNF263, a C2H2 zinc finger transcription factor, represses HS3ST1 transcription. CRISPR-mediated knockout or siRNA knockdown of ZNF263 dramatically increased HS3ST1 expression, leading to enhanced 3-O-sulfation, increased antithrombin binding, increased Factor Xa inhibition, and increased neuropilin-1 binding. ZNF263 is expressed at distinctively low levels in mast cells (heparin-producing cells). |
CRISPR KO and siRNA knockdown of ZNF263 in mammalian cell lines and primary human cells, qRT-PCR, antithrombin binding flow cytometry, Factor Xa inhibition assay, transcriptomics analysis of mast cell expression |
Proceedings of the National Academy of Sciences of the United States of America |
High |
32277030
|
| 2022 |
3-O-sulfation of heparan sulfate, as generated by HS3ST1, contributes to cellular internalization of tau aggregates. In HS3ST1−/− cells (confirmed by LC-MS/MS), uptake of tau aggregates was reduced. Aggregated tau shows higher affinity for 3-O-sulfated HS than non-3-O-sulfated HS, and competition with antithrombin III (which binds 3-O-sulfated HS) reduced tau uptake. |
HS3ST1 KO cell line, LC-MS/MS with 13C-labeled HS calibrants to confirm 3-O-sulfation loss, tau aggregate uptake assay, antithrombin III competition assay |
BMC molecular and cell biology |
High |
36564747
|
| 2023 |
A specific 3-O-sulfated heparan sulfate disaccharide structure made by 3-OST-1 (encoded by HS3ST1) is increased sevenfold in Alzheimer's disease brains versus controls. This structure was identified by LC-MS/MS and confirmed using recombinant sulfotransferases and Hs3st1 knockout mouse HS. A synthetic 14-mer carrying this 3-O-sulfated domain showed stronger inhibition of tau internalization than a 14-mer lacking it. |
LC-MS/MS analysis of brain HS, recombinant enzyme modification assay, Hs3st1−/− mouse HS analysis, synthetic tetradecasaccharide inhibition assay for tau internalization |
Science advances |
High |
37235665
|
| 2023 |
ApoE (all isoforms: ApoE2, ApoE3, ApoE4) recognizes and binds 3-O-sulfated heparan sulfate. In HS3ST1 knockout cells, cell-surface binding and uptake of ApoE were reduced. NMR titration localized ApoE/3-O-S binding to the vicinity of the canonical HS binding motif in ApoE. |
Glycan microarray, SPR binding assay, NMR titration, HS3ST1 KO cell-based ApoE binding and uptake assay |
Angewandte Chemie (International ed. in English) |
High |
37014788
|
| 2012 |
HS3ST1 (Hs3st1) is involved in the insulin secretion pathway in pancreatic β-cells. siRNA-mediated silencing of Hs3st1 reduced glucose-induced insulin secretion (GIIS) in MIN6T3 cells, acting upstream of membrane depolarization. |
siRNA knockdown of Hs3st1 in MIN6T3 cells, glucose-induced insulin secretion assay, sodium chlorate inhibition of sulfation, qRT-PCR |
Journal of diabetes investigation |
Medium |
24843591
|
| 2019 |
HS3ST1 overexpression in renal epithelial cells (HKC8-HS3ST1) leads to prolonged STAT3 phosphorylation in response to HB-EGF (heparin-binding EGF-like growth factor), compared to transient STAT3 phosphorylation in control cells. Pro-fibrotic factors TGFβ1 and TGFβ2/IL1β significantly downregulate HS3ST1 expression in renal epithelial cells and fibroblasts. |
Stable overexpression of HS3ST1 in HKC8 cells, HB-EGF stimulation with STAT3 phosphorylation time-course, qRT-PCR for HS3ST1 in TGFβ-treated cells |
Biochimica et biophysica acta. General subjects |
Medium |
30794825
|
| 2023 |
In castration-resistant prostate cancer (CRPC) cells (C4-2) under hormone depletion, HS3ST1 produces 3-O-sulfated heparan sulfate that facilitates EGF binding to the cell surface and activation of EGFR-ERK1/2 signaling, enabling hormone-independent growth. HS3ST1 knockdown suppressed hormone-independent cell growth, EGF binding, and EGFR-ERK1/2 activation. |
HS3ST1 siRNA knockdown in C4-2 cells, EGF binding assay, EGFR-ERK1/2 phosphorylation assay, cell proliferation assay under hormone-depleted conditions, in vivo xenograft in castrated mice with gefitinib treatment |
Scientific reports |
Medium |
37463954
|
| 2025 |
HS3ST1-generated HSAT (antithrombin-binding heparan sulfate) is expressed in basement membranes of epithelial cells, not only in endothelium. Inactivation of HS3ST1 in pancreatic ductal adenocarcinoma (PDAC) cells eliminated HSAT expression, induced an inflammatory phenotype, suppressed apoptosis markers, and increased metastasis in a mouse PDAC model. HSAT-positive cells bind antithrombin, which inhibits thrombin generation by tissue factor/factor VIIa. |
HS3ST1 inactivation in PDAC cells, mouse experimental metastasis model, antithrombin binding assay, tissue factor/factor VIIa thrombin generation assay, immunostaining of human tissues |
The Journal of clinical investigation |
High |
40924474
|
| 2020 |
HRPII from Plasmodium falciparum competes with antithrombin for binding to 3-O-sulfated heparan sulfate on endothelial cells, disrupting AT's anti-inflammatory/barrier-protective signaling. siRNA knockdown of HS3ST1 (3-OST-1) in endothelial cells downregulated HRPII's pro-inflammatory effects, supporting the mechanism that HRPII's action depends on competition at the 3-O-sulfated HS/AT interface. |
siRNA knockdown of HS3ST1 in endothelial cells, endothelial barrier permeability assay, Src-dependent VE-cadherin phosphorylation assay, competition experiments with AT |
Journal of thrombosis and haemostasis : JTH |
Medium |
31858717
|
| 2026 |
An E86Q active-site mutant of 3-OST-1 acts as a dominant-negative inhibitor: it retains substrate and donor binding but abolishes catalytic activity, reducing wild-type 3-OST-1 activity by >80% in vitro and significantly decreasing 3-O-sulfated HS products in cells without affecting total HS abundance. An LC-MS/MS assay was developed that can quantify endogenous 3-OST-1 activity at nanogram sensitivity. |
Site-directed mutagenesis (E86Q), in vitro LC-MS/MS activity assay with 13C-labeled internal standards, cellular overexpression of dominant-negative mutant, heparinase digestion and AMAC derivatization |
Glycobiology |
High |
41914643
|
| 2025 |
Rational design of 3-OST-1 mutants W72R and H144R, individually and as double mutant W72R/H144R, increased enzyme activity 1.7-, 1.1-, and 2.2-fold respectively over wild-type. Enhanced activity resulted from increased substrate affinity, as shown by enzyme kinetics and molecular dynamics simulations. The mutant-modified heparin showed superior anticoagulant properties. |
Rational mutagenesis, enzyme kinetic assays (Km, Vmax), molecular dynamics simulations, anti-factor Xa anticoagulant activity assay |
Journal of agricultural and food chemistry |
Medium |
40267027
|