PERF 15 is the major protein of the rat perforatorium found between the inner acrosomal and the outer face of the nuclear envelope of the sperm head. Screening of a testicular lambda-gt 11 phage cDNA library with an affinity-purified polyclonal antibody to this polypeptide allowed the isolation of several positive clones. Sequencing of a positive cDNA clone revealed a 396-nucleotide open reading frame encoding a protein of 132 amino acids (15,060 Da). The amino terminal sequence of the isolated PERF 15 protein was identical to the deduced amino acid sequence of the cDNA clone. Riboprobes encoding the PERF 15 cDNA recognized a complementary 1.0-kb mRNA only in the testis. Developmental Northern blots and in situ hybridization revealed that this mRNA is first transcribed in pachytene spermatocytes and reaches a peak of expression in round spermatids. PERF 15 belongs to a family of fatty acid binding proteins, showing a 61% sequence similarity to myelin P2, a 58% sequence similarity to adipocyte lipid binding protein, and a 47% similarity to heart fatty acid binding protein. It contains a stretch of amino acids, including the motif Ile-Ser-Phe-Lys, present in myelin P2 protein that has been implicated in the induction of experimental autoimmune neuritis. This homology taken together with its subacrosomal location indicates that the PERF 15 protein may interact with the membrane of the acrosome and play a major role in the structural arrangement and stability of this organelle.