MURINE STEROID SULFATASE (MSTS) - PURIFICATION, CHARACTERIZATION AND MEASUREMENT BY ELISA

The murine steroid sulfatase (mSTS) is a microsomal enzyme, important in steroid metabolism. In the mouse, the gene encoding mSTS is pseudoa utosomal and thus escapes X-inactivation. We have purified steroid sul fatase approximately 30-fold from mouse liver microsomes and its prope rties have been investigated. The major steps in the purification proc edure included solubilization with Triton X-100, gel filtration chroma tography, DEAE-Sephadex chromatography and HPLC gel filtration chromat ography. The purified sulfatase showed a relative molecular weight of 128 kDa on HPLC gel filtration, whereas the enzyme migrated as two ban ds of 60 and 68 kDa on sodium dodecyl sulfate-polyacrylamide gel elect rophoresis. The isoelectric point of steroid sulfatase was estimated t o be 6.2 by column chromatofocusing. Polyclonal antibodies to the puri fied protein were-prepared. An Enzyme Linked Immunosorbent Assay (ELIS A) was developed using purified monospecific anti-mSTS antibodies labe lled with peroxidase. The standard criteria of precision and reproduci bility were satisfied. The assay was applicable to routine determinati on of mSTS samples in research laboratories. Differences in mSTS liver concentrations were used to identify putative alleles for the mSTS ge ne (Sts). Results in ELISA confirmed the polymorphism previously demon strated for an enzymatic mSTS activity assay in two inbred mouse strai ns.