THE AMINO ACID PRECURSORS ARE DERIVED FROM THE VACUOLE* (Received for publication, July 23,1992)

The cellular localization of the origin of a-aminoadipate used in penicillin biosynthesis and the first enzymic step in Penicillium chrysogenum involved, &(aaminoadipy1)-L-cysteinyl-D-valine synthetase (ACVS), has been studied. Subcellular fractions were obtained from protoplasts of a high penicillin-producing strain upon lysis by Triton X100, and vacuoles purified from them. They were identified by the aid of a-mannosidase as a marker enzyme, by the presence of polyphosphate, and their ability to sequester [“Cllysin, added to the protoplasts prior to subcellular fractionation. 15.6 and 26.5%, respectively, of 6-[“C]a-aminoadipate, and 8.5 and 10.3%, respectively, of [“Clvaline added accordingly were also found in the vacuole, and the higher proportion was found in vacuoles isolated from penicillin-producing mycelia. ACVS protein was detected in the membrane as well as the soluble fraction of the purified vacuoles. We propose therefore that ACVS is located either within or bound to the vacuolar membrane, and that the precursor amino acids for penicillin biosynthesis are withdrawn from the vacuolar amino acid pool.