Effect of different doses of nitrogen (N) (90, 120, 150 and 180 kg Nha–1) on the activities of aminotransferases and alkaline inorganic pyrophosphatase (AIP) in relation to the accumulation of proteins, amino acids and sugars in roots and internodes at 15 and 40 days post anthesis (DPA) stages was studied in six wheat genotypes namely HD 2967, GLU 1101, PBW 343, BW 9022, PH-132-4840 and PBW 550. Supra-optimal N doses (150 kg Nha–1 and 180 kg Nha–1) accentuated glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT) and alkaline inorganic pyrophosphatase activities in correspondence with an increase in amino acid, protein and sugar content in both roots and internodes in all the six genotypes. Activities of analyzed enzymes were significantly high at 15 days post anthesis (DPA) stage and thereafter declined at maturity (40 DPA) in parallel with decrease in amino acid contents. Maximum activity of GOT, GPT and AIP was observed in HD 2967 and GLU 1101 genotypes along with higher build up of proteins and amino acids which resulted in higher grain yield. Activity of GPT was comparatively high over GOT, indicating its major role towards protein synthesis. Grain filling processes in terms of proteins and amino acids were positively correlated with GOT and GPT activities while sugars were correlated to AIP. Thus, nitrogen acquisition and assimilation resulted in favoured utilization of N in form of amino acid and proteins accumulation while sugar content was also stimulated. Due to immense activities of aminotransferases and higher contents of amino acids and proteins in GLU 1101 and HD 2967 genotypes at optimal dose and higher dose of N, these genotypes hold future potential for developing new cultivars with better grain quality characteristics.