We report the complete nucleotide sequence of two genes encoding major peroxisomal polypeptides (PXPs) of Candida tropicalis. One, POX4, encodes PXP-4, which is the most abundant polypeptide in cells grown on oleic acid, and the other, POX5, is the gene for PXP-5. Each of the two polypeptides was found to be the subunit of a distinct long-chain acyl-coenzyme A oxidase: acyl-CoA oxidase II (PXP-4) or acyl-CoA oxidase I (PXP-5). Both the genes had no intron and gave a single open reading frame. The NH2-terminal sequences, except the initiator methionine, and the calculated molecular weights of the deduced polypeptides were consistent with those of the respective PXPs. Well-conserved sequences of 12 and 16 hydrophobic amino acids were present in the middle of the polypeptide, instead of at the NH2 terminus, and may be internal signal sequences for the peroxisomal location of PXPs. Although the two polypeptides were significantly homologous throughout their sequences, the local homologies in two regions out offive were markedly diverged from the average (63%); the homology in the second region was 93%, whereas that in the fourth one was only 24%. The implications of this finding are discussed in respect to the multiplicity of peroxisomal enzymes and the presence of multifunctional proteins in peroxisomes.