Nitrogen-fixing bacteria are essential for providing nitrogen (N) to soybean (Glycine max (L.) Merrill) plants, often resulting in a reduced or eliminated requirement for N-fertilizer. The application of these microorganisms can enhance N fixation and maximize soybean yield. Soybean is attaining significant attention in southeastern Anatolia as a second crop; however, the interactive effects of N-fixing bacteria and N doses on its yield are still unknown. This study investigated the effects of N application and seed inoculation with Bradyrhizobium japonicum bacteria on yield and quality attributes of soybean grown as a second crop during 2021 and 2022. Seed inoculation (bacteria-inoculated or non-inoculated) were the main plots, whereas N doses [0 (N₀), 40 (N₁), 80 (N₂), 120 (N₃), 160 (N₄), 200 (N₅), 240 (N₆), and 280 (N₇) kg ha^-1 were sub plots. Bacteria inoculation and N doses significantly (p≤0.01) altered the number of pods per plant, plant height, thousand seed weight, grain yield, protein ratio, and oil ratio. The highest (3645 kg ha^-1) and the lowest (3072 kg ha^-1) grain yield was recorded for bacteria-inoculated and non-inoculated seeds, respectively. Similarly, N₇ (3628 kg ha^-1) and N₀ (2791 kg ha^-1) produced the highest and the lowest grain yield, respectively. Regression analysis revealed that 144 kg ha^-1 pure N should be applied for the cultivation of the second crop soybean under the climatic conditions of Şanlıurfa, Türkiye, and seeds should be inoculated with B. japonicum for better yield and economic returns.