Biochar application can alter soil properties and enhance soil quality. However, the effects of the regulatory mechanisms underlying nitrogen fertilizer application on soil physicochemical properties and microbial characteristics remain unclear. We conducted a three-year maize field experiment to investigate the effects of four biochar application rates (0, 8, 16, and 24 t/ha) and three nitrogen fertilizer rates (0, 150, and 300 kg/ha) on soil physicochemical properties, enzyme activities, microbial community structure, and maize yield. In the 0–20 cm soil layer, the combined application of biochar and nitrogen fertilizer significantly increased soil porosity (SP) and soil water content (SWC), reduced soil bulk density (SBD), and increased soil organic carbon (SOC) and total nitrogen (TN) levels. Additionally, β-D-cellobiohydrolase and leucine aminopeptidase activities were enhanced, along with nitrogen acquisition potential. The relative abundance of Proteobacteria was highest at a biochar application rate of 16 t/ha. The combined 16 t/ha biochar and 150 kg/ha nitrogen treatment exhibited the highest maize yield. Maize yield was positively correlated with SP, SWC, SOC, and TN. Consequently, the combined application of 16 t/ha biochar and 150 kg/ha nitrogen improved soil physicochemical properties, enhanced soil enzyme activities, regulated key soil microbial taxa, and increased maize yield.