The neurosecretory bag cells of the mollusk, Aplysia, produce a peptide egg-laying hormone, ELH, via a multistep proteolytic processing sequence analogous to those which have been demonstrated for secretory peptides in other systems. The major members of this processing sequence were analyzed by sequential sodium dodecyl sulfate-polyacrylamide gel electrophoresis and isoelectric focusing of bag cell proteins synthesized in the presence of labeled precursors and to elucidate the precursor-product relationships between these proteins in pulse-chase experiments. Major members [8] of the processing sequence were identified. The ultimate precursor is a 29,000-dalton, pI = 7.7, protein which gives rise to a pI = 7.2 protein with an apparent MW of 6000 and heterogeneous species of MW 16,000-20,000. The latter protein or proteins are processed to apparent end products of 13,000-14,500 daltons: the pI = 7.2 species yields precursors to the final secretory products. These include a pI = 7.5 peptide which is cleaved to ELH (MW 4385, pI > 9) and a MW 45,000, pI = 4.1 species which yields the other secretory product, AP (MW 4500, pI = 4.9). A single precursor probably is processed to yield 3 products, 2 of which are secreted; each product is generated via at least 1 intermediate form.