The soil nitrogen (N) availability and urease activity (UA) in a humid ecosystem with variable rainfall distribution and poor soil fertility are not well understood. A complete appreciation of N cycling in the soil–water–plant continuum is needed to better manage N and water in regions that will be strongly affected by climate change. A sorghum (Sorghum bicolor L.) study located in Florence, South Carolina, USA, was conducted using a variable-rate pivot system. We hypothesized that supplemental irrigation (SI) and N would enhance UA and N uptake while minimizing the concentration of N in porewater (TINW). The aim of the study was to assess the impact of SI (0, 50, and 100%) and N fertilization (0, 85, and 170 kg N ha−1) on: UA; total N (TNS); total inorganic N (TINS); TINW; and N uptake of sorghum. Results support our research hypothesis. The greatest UA was from 0% SI and 170 kg ha−1 (18.7 µg N g−1 ha−1). Porewater N (mg L−1), when averaged across SI and N showed a significantly lower concentration at lower soil depth (9.9 ± 0.7) than the upper depth (26.1 ± 2.4). The 100% SI had the greatest biomass N uptake (NUPB) of 67.9 ± 31.1 kg ha−1 and grain N uptake (NUG) of 52.7 ± 20.5 kg ha−1. The greatest NUPB (70.9 ± 30.3 kg ha−1) and NUG (55.3 ± 16.5 kg ha−1) was from the application of 170 kg N ha−1. Overall, results showed that proper use of water and N enhanced soil N dynamics, and improved biomass productivity and N uptake of sorghum.