The cDNA of human cytosolic purine 5'-nucleotidase (EC 3.1.3.5), which is the supposed regulatory allosteric enzyme of purine nucleotide degradation, has been cloned from a human placenta cDNA library. The predicted open reading fram encodes a protein of 561 amino acids with a molecular mass of 64,966 Da. The deduced amino acid sequence exhibits 95% identity with the sequence of the B type subunit of chicken cytosolic purine 5'-nucleotidase. Northern blot analysis of human placenta poly(A) +RNA revealed a single band of 3.6 kilobases using the cloned cDNA.