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Human plasma glutathione peroxidase (GSHPx) has been shown to be a selenium-containing enzyme immunologically distinct from cellular GSHPx. Oligonucleotide probes, based on the partial amino acid sequence of plasma GSHPx, were synthesized and used to screen a human placenta cDNA library. Nucleotide sequence analysis of the obtained clones revealed that GSHPx consisted of a 678-base pair open reading frame coding for a 226-amino acid polypeptide with a Mr of 25,389. About 50% of the deduced amino acid sequence was confirmed by partial amino acid sequencing of the peptides in a lysine endopeptidase-digest of the purified enzyme. The amino acid sequence exhibited only 44% homology with that of human cellular GSHPx. Northern blot analysis revealed a single transcript of 2.2 kilobases in the poly(A)+ RNA fractions of human placenta and HepG2 (a human hepatic cell line), but not that of human liver and endothelial cells.

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Mixtures of 17-base long oligonucleotides possibly encoding a hexapeptide of Drosophila melanogaster glucose-6-phosphate dehydrogenase were synthesized and used as probes for screening a genomic library of D. melanogaster constructed in Charon 4A vectors. A total of about 60,000 plaques were initially screened, and after two successive plaque purification three clones carrying the identical 13-kb EcoRI fragment were isolated.That these clones contain the G6PD coding sequence was demonstrated by in situ hybridization of the cloned DNA fragments to salivary gland polytene chromosomes and in vitro translation of the hybrid-selected mRNA. As suggested by Torczynski et al. (1984), this method using short synthetic probes appears versatile for isolating low-copy genes from genomic libraries.

The cDNA coding rabbit glutathione peroxidase was isolated from liver cDNA library in lambda gtll by cross hybridization with the rat glutathione peroxidase cDNA which was cloned in this laboratory and reported elsewhere(l). The cDNA consisted of 600 bp of the coding region and the nucleotide sequence revealed that TGA, which is to be the stop codon in general, encoded seleno-cysteine(SeC) residue as was proved to be so with glutathione peroxidases of mouse (2), man(3) and rat(l). The amino acid sequence deduced from cDNA possesses 84, 84, 87 and 85% homology with rat, mouse, human, and bovine(4) enzymes, respectively.