Nitrogen (N) is a key nutrient influencing wheat growth, grain yield, and quality. A long-term field experiment was conducted using cultivar Zhengmai 1860 to clarify the effects of N topdressing on grain protein composition, starch accumulation, and yield. Treatments included a basal N application of 150 kg ha−1 (N1) combined with four topdressing rates at jointing: 37.5, 75, 112.5, and 150 kg ha−1 (N1 + 37.5, N1 + 75, N1 + 112.5, N1 + 150). Nitrogen topdressing significantly affected the physiological and biochemical characteristics of grains at different spike positions. Amylopectin, globulin, soluble starch (SS), and soluble starch synthase (SSS) accumulated most under 75–112.5 kg ha−1, with N1 + 75 showing the strongest response in basal and middle spike grains. Amylose and granule-bound starch synthase (GBSS) peaked at the middle spike under N1 + 112.5. Protein component (gliadin, glutelin, albumin), amino acids, glutamate synthase (GOGAT), and glutamine synthetase (GS) increased progressively with higher N rates, with maximum accumulation at N1 + 150. Nitrogen topdressing also enhanced spike number (5.05–37.13%), grains per spike (3.86–16.22%), and 1000-grain weight (2.72–5.79%), with the highest yield (9451.7 kg ha−1) at N1 + 112.5. These results highlight the critical role of optimized N management in improving grain composition and yield in wheat.