Mutation of the Secys residue 266 in human type 2 selenodeiodinase alters Se-75 incorporation without affecting its biochemical properties

Type 2 deiodinase (D2) is a low K-m iodothyronine deiodinase that metaboliz es thyroxine (T-4) to the active metabolite T-3. We have recently shown tha t the cDNA for the human D2 coding region contains two in-frame selenocyste ine (TGA) codons. The 3' TGA is seven codons 5' to a universal stop codon, TAA. The human D2 enzyme, transiently expressed in HEK-293 cells, can be in vivo labeled with Se-75 as a doublet of approximately 31 kDa. This doublet is consistent with the possibility that the carboxy-terminal TGA codon can either encode selenocysteine or function as a stop codon. To test this hyp othesis we mutagenized the second selenocysteine codon to a cysteine (TGC) or to an unambigous stop codon (TAA). While the selenium incorporation patt ern is different between the wild-type and mutant proteins, the deiodinatio n properties of the enzyme are not affected by mutating the 3'TGA codon. Th us, we conclude that neither this residue nor the remaining seven carboxy-t erminal amino acids are critical for the deiodination process. (C) Societe francaise de biochimie et biologie moleculaire / Elsevier, Paris.