title: Effects of mutations and glycosylations on STS activity: a site-directed mutagenesis study.
abstract: Steroid sulphatase (STS) catalyses the formation of active steroids from inactive steroid sulphates. High levels of intra-tumoural STS mRNA are associated with a poor prognosis in post-menopausal patients with oestrogen receptor positive breast cancer. In this study, analysis of the mutated STS protein showed that N- and C-terminal truncated STS constructs are inactive. Histidine 136, located inside the active site, is crucial for STS activity whereas proline 212, which allows the protein turn into the membrane, is not. Mutations in glycosylation sites asparagine 47 and 259 decreased STS activity while asparagine 333 and 459 mutations did not affect it. However, immunoblot studies revealed that all four N-linked sites are glycosylated to some extent. In addition, a polyclonal antibody raised in rabbits against human STS was developed and characterised. These data increase our knowledge of the STS enzyme structure and may help design new STS inhibitors.
authors.affiliation: Endocrinology and Metabolic Medicine and Sterix Ltd., Faculty of Medicine, Imperial College London, St. Mary's Hospital, London, W2 1NY, UK. c.stengel@imperial.ac.uk
authors.forename: Chloe
authors.initials: C
authors.lastname: Stengel
authors.forename: Simon P
authors.initials: SP
authors.lastname: Newman
authors.forename: Joanna M
authors.initials: JM
authors.lastname: Day
authors.forename: Helena J
authors.initials: HJ
authors.lastname: Tutill
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Reed
authors.forename: Atul
authors.initials: A
authors.lastname: Purohit
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D006031:Glycosylation; D006801:Humans; D007106:Immune Sera; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D050505:Mutant Proteins / Q000378:metabolism; D017354:Point Mutation / Q000235:genetics*; D043266:Steryl-Sulfatase / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*
chemical_list: D007106:Immune Sera; D050505:Mutant Proteins; D043266:Steryl-Sulfatase