The rheological properties and end-use qualities of many foods are mainly determined by the types and levels of grain storage proteins (GSPs) in wheat. GSP levels are influenced by various factors, including tillage management, irrigation, and fertiliser application. However, the effects of irrigation and nitrogen on GSPs remain unclear. To address this knowledge gap, a stationary split–split block design experiment was carried out in low- and high-fertility (LF and HF) soil, with the main plots subjected to irrigation treatments (W0, no irrigation; W1, irrigation only during the jointing stage; W2, irrigation twice during both jointing and flowering stages), subplots subjected to nitrogen application treatments (N0, no nitrogen application; N180, 180 kg/ha; N240, 240 kg/ha; N300, 300 kg/ha), and cultivars tested in sub–sub plots (FDC5, the strong-gluten cultivar Fengdecun 5; BN207, the medium-gluten cultivar Bainong 207). The results showed that GSP levels and processing qualities were significantly influenced by nitrogen application (p < 0.01), N240 was the optimal nitrogen rate, and the influence of irrigation was dependent on soil fertility. Optimal GSP levels were obtained under W2 treatment at LF conditions, and the content was increased by 17% and 16% for FDC5 and BN207 compared with W0 under N240 treatment, respectively. While the optimal GSP levels were obtained under W1 treatments at HF conditions, and the content was increased by 3% and 21% for FDC5 and BN207 compared with W0 under N240 treatment, respectively. Irrigation and nitrogen application increased the glutenin content by increasing Bx7 and Dy10 levels in FDC5, and by increasing the accumulation of Ax1 and Dx5 in BN207. Gliadins were mainly increased by enhancing α/β-gliadin levels. Correlation analysis indicated that a higher soil nitrate (NO3-N) content increased nitrogen remobilisation in leaves. Path analysis showed that Dy10, Dx5, and γ-gliadin largely determined wet glutenin content (WGC), dough stability time (DST), dough water absorption rate (DWR), and sedimentation value (SV). Therefore, appropriate irrigation and nitrogen application can improve nitrogen remobilisation, GSP levels, and processing qualities, thereby improving wheat quality and production.