Tree pruning is a management tool in agroforestry systems for reducing shade, enhancing nutrient cycling or providing fodder. However, little information is available on the effect of pruning management on plant growth, non-structural carbohydrate (NSC) levels in the roots and N2 fixation of Sesbania sesban. A glasshouse experiment was conducted to assess the effect of pruning frequency on biomass production, NSC levels and N2 fixation of Sesbania sesban. Treatments included pruning at (i) 3 months only, (ii) 3 and 6 months, and (iii) 3, 6 and 9 months, with each pruning removing shoot biomass above 50% of the initial height. Increased pruning frequency decreased above- and belowground dry matter (DM), and root NSC levels. The decrease in aboveground DM correlated with reduced levels of starch, sugar and total non-structural carbohydrates. Pruning more frequently significantly decreased nodulation, the percentage N derived from the atmosphere and N2 fixed. It could be concluded that more frequent prunings decreases biomass and NSC levels which could reduce the regeneration capacity of trees since they rely on NSCs to regrow. The suppressive effects of increased pruning frequency on N2 fixation could decrease the desirable benefits of legume trees on soil N fertility improvement.