What are the effects of no-take marine reserves on trophic relationships of coral reef fish? Previous studies often have lacked detailed dietary information on major predators, and have often been confounded by differences in habitat complexity between reserve and fished sites. This study investigates the effects of marine reserve protection on predator-prey interactions of coral reef fish on the inshore islands of the Great Barrier Reef (GBR). The abundance of species of prey fish of Plectropomus leopardus (Serranidae), a piscivore and the major target of the hook and line fisheries on the GBR, were estimated in protected and fished zones. These prey species were identified from previous detailed studies of the diet of P. leopardus. Fish populations and habitat characteristics were surveyed by underwater visual census. Previous studies had determined that the biomass of P. leopardus was 3–4 times higher in protected than fished zones in the Whitsunday and Palm Islands, central GBR, after 14 years of protection. Eight of the nine prey species had a higher density within fished zones than protected zones, six significantly so. The density of all prey fish was twice that in the fished than the protected zone (p < 0.001). There were no significant differences in availability of different sized refuge holes, structural complexity or live coral cover between zones. Thus, important attributes of habitat complexity did not confound the comparisons between reserve and fished zones. Finally, a significant negative correlation (r = 0.46) between coral trout biomass and summed prey fish biomass suggested that predation may be an important structuring process in this system. The results have implications for the conservation of fishery targets and their prey. The study highlights the potential ecosystem implications of the use of no-take marine reserves as conservation and fisheries management tools.