2-O sulfatase compositions and related methods
The invention relates to 2-O sulfatase and uses thereof. In particular, the invention relates to recombinantly produced 2-O sulfatase, functional variants and nucleic acid molecules that encode these molecules. The invention also provides methods of using 2-O sulfatase for a variety of purposes, including degrading and analyzing glycosaminoglycans (GAGs) present in a sample. For instance, 2-O sulfatase may be used for determining the purity, identity, composition and sequence of glycosaminoglycans present in a sample. The invention also relates to methods of inhibiting angiogenesis and cellular proliferation as well as methods for treating cancer, neurodegenerative disease, atherosclerosis and microbial infection using 2-O sulfatase and/or GAG fragments produced by degradation with 2-O sulfatase.
1. An isolated 2-O sulfatase produced by expressing an isolated nucleic acid molecule selected from the group consisting of:
(a) nucleic acid molecules which hybridize to a nucleic acid molecule having a nucleotide sequence selected from the group consisting of nucleotide sequences set forth as SEQ ID NOs: 1 and 3, wherein the hybridization conditions are 1) hybridization at 65° C. in hybridization buffer that consists of 3.5×SSC, 0.02% Ficoll, 0.02% polyvinyl pyrrolidone, 0.02% Bovine Serum Albumin, 2.5 mM NaH 2 PO 4 , pH7, 0.5% SDS, 2 mM EDTA, —wherein SSC is 0.15M sodium chloride/0.015M sodium citrate, pH7; SDS is sodium dodecyl sulphate; and EDTA is ethylenediaminetetracetic acid and 2) washing in 2×SSC at room temperature and then in 0.1–0.5×SSC/0.1×SDS at 68° C.,
(b) nucleic acid molecules that differ from the nucleic acid molecules of (a) in codon sequence due to degeneracy of the genetic code, and
(c) full complements of (a) or (b).
2. An isolated polypeptide comprising:
a 2-O sulfatase having an amino acid sequence selected from the group consisting of SEQ ID NOs: 2 and 4.
3. An isolated 2-O sulfatase having an amino acid sequence selected from the group consisting of SEQ ID NOs: 2 and 4.
4. A composition comprising the 2-O sulfatase of claim 1 or 3 and a pharmaceutically acceptable carrier.
5. The 2-O sulfatase of claim 1 , wherein the nucleic acid molecule is expressed recombinantly.
6. The 2-O sulfatase of claim 5 , wherein the 2-O sulfatase is recombinantly expressed in E. coli.
7. The isolated polypeptide of claim 2 , wherein the 2-O sulfatase has the amino acid sequence as set forth in SEQ ID NO: 4.
8. The isolated 2-O sulfatase of claim 3 , wherein the isolated 2-O sulfatase is synthetic.
9. A composition comprising: the isolated polypeptide of claim 2 and a pharmaceutically acceptable carrier.