The medium compositions such as carbon and nitrogen sources, moisture content and inorganic salt affected the microbial protein (MP) production. Imbalance of carbon-nitrogen ratio in apple pomace (AP) limited the microbial utilization. Hence, those conditions must be optimized to achieve maximum MP. In this work, AP was pretreated by extrusion technology to obtain extruded apple pomace (EAP). Subsequently, the medium compositions were optimized using Plackett-Burman design (PBD) and Box-Behnken design (BBD). PBD determined four significant factors (bran, glucose, packing quantity (PQ), water to material ratio (W/M)) out of the eight variables. The BBD results showed that optimal true protein content (10.42%), effective viable count (1.94×109 CFU/g) and crude protein content (18.73%) were achieved at bran 16.22%, glucose 8.09%, PQ 9.88 g and W/M 1.56. Compared with AP, the true protein and crude protein content of optimal fermented EAP (FEAP) were increased by 152% and 216%, respectively. According to fluorescence microscopy, the cellulose of AP was little effected by extrusion technology while was mostly degraded by mixed strains (Aspergillus niger, Candida utilis, Geotrichum candidum and Lactic acid bacteria). Combination of extrusion and fermentation, the medium compositions were optimized to promote the bioconversion of AP into MP feed.