Evidence for the presence of thioltransferase in the lens.

Thioltransferase (TTase) activity was identified and partially purified from the ocular tissue for the first time. The enzyme activity depended on the presence of reduced glutathione (GSH), glutathione reductase (GR) and NADPH to reduce the disulfide bond in a synthetic substrate, hydroxyl ethyl disulfide (HEDS). Maximum activity was obtained in a pH 7.4 phosphate buffer at 30 degrees C. This enzyme distinguishes from other reducing enzymes such as thioredoxin that do not require GSH and GR for their catalytic activity. It also differs from the 52 kDa enzyme, protein disulfide isomerase by its smaller molecular size and its stability against heat treatment. TTase activity was higher in the epithelial layer but distributed evenly in the rest of the lens also, TTase showed similar activity in the lenses obtained from rats, pigs, bovine, guinea pigs, chick embryos and humans. The molecular weight of this enzyme was estimated to be 11.5 kDa on a SDS-PAGE system. Western blot analysis showed the protein reacted positively to the antibody raised by the purified pig liver TTase. Similarly the antibody raised by the partially purified lens enzyme reacted positively with the purified pig lever TTase. The presence of TTase in the lens was confirmed further with the slot blot analysis where it demonstrated a 32P-labeled cDNA from pig liver TTase hybridizing with the RNA in the pig lens or rabbit lens epithelium cells. Based on the above information it was concluded that the lens TTase is comparable to TTase from other tissues in its functional and structural properties. It is hypothesized that the lens TTase has a significant physiological role in sulfhydryl homeostasis in the lens by protecting the SH groups of the proteins from S-thiolation. It is speculated that, lens TTase may be primary antioxidant in the lens along with GSH and GR by protecting the vulnerable lens proteins against oxidative damage.