Peroxisomes are ubiquitous, oxidative subcellular organelles with important functions in cellular lipid metabolism and redox homeostasis. Loss of peroxisomal functions causes severe disorders with developmental and neurological abnormalities. Zebrafish are emerging as an attractive vertebrate model to study peroxisomal disorders as well as cellular lipid metabolism. Here, we combined bioinformatics analyses with molecular cell biology and reveal the first comprehensive inventory of Danio rerio peroxisomal proteins, which we systematically compared with those of human peroxisomes. Through bioinformatics analysis of all PTS1-carrying proteins, we demonstrate that D. rerio lacks two well-known mammalian peroxisomal proteins (BAAT and ZADH2/PTGR3), but possesses a putative peroxisomal malate synthase (Mlsl) and verified differences in the presence of purine degrading enzymes. Furthermore, we revealed novel candidate peroxisomal proteins in D. rerio, whose function and localisation is discussed. Our findings confirm the suitability of zebrafish as a vertebrate model for peroxisome research and open possibilities for the study of novel peroxisomal candidate proteins in zebrafish and humans.