The objective of this study was to investigate the effects of tree cover and season on soil N dynamics and microbial biomass in a semi-arid southern African savanna in Zimbabwe. We used a randomized complete block design with five blocks of 100 × 100 m, demarcated in a 10-ha pocket of Colophospermum mopane-dominated woodland protected from grazing and fire. In each block, we randomly selected three mopane trees with large canopies (8.3 m crown diameter) and another three with small canopies (2.7 m crown diameter). We determined soil organic carbon and nutrient concentrations, litterfall N and C inputs, microbial biomass and N transformations beneath large and small mopane trees as well as in the intercanopy areas. Soil organic carbon, microbial biomass, N, P and K were more than twice those beneath large trees than in the intercanopy areas. Rainy-season net mineral N accumulation rate in the surface soil (0–10 cm) ranged from 3.71 μg g−1 mo−1 in the intercanopy areas to 8.80 μg g−1 mo−1 beneath large trees; correspondingly, net nitrate accumulation rate ranged from 1.33 to 3.60 μg g−1 mo−1. Dry-season net mineral N and net nitrate accumulation rates were similar across sampling sites and did not exceed 2 and 0.4 μg g−1 mo−1, respectively. Litterfall N inputs were positively and significantly correlated with soil N availability, microbial biomass N and N transformations. At all sampling sites, microbial biomass and mineral N pools in the dry season were maximum when soil moisture (∼5%) and N transformations were minimum. In contrast, when soil moisture (9–13%) and N transformations were maximum in the rainy season, microbial biomass and mineral N pools were minimum. It is concluded that the improved soil conditions beneath isolated trees in semi-arid savannas may enhance herbaceous biomass yield especially of canopy shade-tolerant species.