NADPH with Cytosol Stimulates Deiodination by Detergent-Solubilized Hepatic Microsomes: Evidence for NADPH-Dependent Cytosolic Non-Glutathione Reductase System
Based upon reconstitution experiments in which very low 5’-deiodinase (5’-DI) activity of isolated rat liver microsomes was restored to various degrees by the addition of cytosol, the existence of an endogenous cytosolicstimulating cofactor has been previously demonstrated (1). From similar reconstitution experiments using starved rats, it has been proposed that NADPH and/or GSH were cofactors for 5’-DI, the augmenting action of NADPH, being attributed to the generation of GSH through glutathione reductase (2). However, supporting evidence indicating that glutathione, as well as NADPH are essential endogenous cofactors in mediating 5’-DI stimulation has been controversial (3,4). We previously observed an important role of NADPH in stimulating microsomal 5’-DI in the presence of cytosol (5). Furthermore, using a microsome preparation, we have recently demonstrated a new non-glutathione NADPH-dependent cytosolic reductase system, which operates in the presence of intermediate (fraction B) and high molecular weight (MW) components (fraction A), without very low M.W. components including glutathione (GSH) (5). On the other hand, our laboratory has recently achieved solubilization of 5’-DI by detergents (6) and partial purification of the enzyme by DEAE-Sephacel column chromatography. Accordingly, it was the purpose of the present investigation to examine the effect of NADH, NADPH, and GSH on the stimulation of 5’-DI in a reconstitution assay system utilizing a detergent solubilized 5’-DI preparation with cytosol or fractionated cytosolic components (i.e., fraction A and B).
KeywordsGlutathione Reductase Mount Sinai Hospital Cytosolic Component Iodothyronine Deiodinase Reconstitution Experiment
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