We have screened a cDNA library (20,000 clones) made from rat kidney poly(A)+ RNA, using an oligonucleotide probe that was a mixture of 14-base DNA oligomers containing all 32 possible sequences coding for residues 32-36 of the gamma-glutamyl transpeptidase (EC 2.3.2.2.) heavy chain. We isolated and sequenced two cDNAs corresponding to the mRNA coding for the entire length of the enzyme precursor. The nucleotide sequence that we obtained (2072 bases) reveals an open reading frame of 1707 nucleotides coding for the common precursor of both enzyme subunits. The amino acid sequence begins with the 21 residues located at the NH2-terminal hydrophobic region of the heavy subunit. We show that this sequence, which is not processed, is the only possible signal peptide in the sequence. Five potential N-glycosylation sites are present in the gamma-glutamyl transpeptidase sequence. Using one of the two cDNA clones as probe, a 2.2-kilobase sequence was detected by blot analysis in rat kidney and human fetal liver RNA.