We have cloned a gene (pepF) encoding a serine carboxypeptidase, proteinase F (PEPF), from Aspergillus niger. The sequences were identified in a phage lambda genomic DNA library using a synthetic probe based on the N-terminal sequence of PEPF. Nucleotide sequence data from pepF genomic and cDNA clones reveals that it is composed of four exons of 199, 283, 227 and 881 bp, interrupted by three introns of 53, 69 and 59 bp. The sequence of pepF codes for a polypeptide of 530 amino acids (aa), of which the first 52 aa are not present in the mature PEPF. This region may represent a prepro sequence that is removed by proteolytic cleavage as a monobasic cleavage site (Lys52). Northern blot analysis of total cellular RNA extracted from A. niger cells indicates that pepF is transcribed as a single 1.8-kb mRNA, which is regulated by nitrogen and carbon repression, specific induction and the pH of the culture medium.