A human cDNA encoding 41-kDa phosphoribosylpyrophosphate (PRPP) synthetase (PRS)-associated protein (PAP41) was cloned from two expressed sequence tag (EST) clones having the nucleotide similarity of 61.5 and 70.0% to human PAP39 cDNA. The predicted open reading frame of 1107 base pairs (bp) has the nucleotide identity of 91.8% to rat PAP41 and encodes a protein of 369 amino acids with a calculated molecular weight (MW) of 40,925. The deduced amino acid sequence exhibits the 98.9% identity to rat PAP41 and 72.2, 50.6, and 50.0% identity with human PAP39, PRS I, and PRS II, respectively, but lacks the PRPP binding site. Southern blot analysis suggested that the PAP41 gene exists as a single copy in the human genome. The single PAP41 mRNA of about 2.1 kb was shown to be present in five human cell lines by Northern blot analysis.