Figure 2. Immunoblots showing the presence of nitrated proteins in bean and soybean nodules.

(a) Bean plants were grown in the absence of NO₃⁻ or under common conditions with 0.25 mM NH₄NO₃, and soybean plants were grown in the field. Nodule extracts were analyzed on 12.5% SDS-gels. Immunoblots (up) and Coomassie-stained gels (down) are shown. Protein loaded was 50 µg (bean) or 20 µg (soybean) per lane.

(b) Extracts from similar nodule samples as above were loaded on preparative native gels, then Lb bands were eluted and proteins were resolved on 15% SDS-gels and blotted. Protein loaded was 20 µg per lane.

Y, young nodules; M, mature nodules; S, senescent nodules; 4N, young nodules after 4 days of NO₃⁻ treatment. Molecular mass (kDa) markers are indicated on the left. Nitrated protein bands and nitrated Lb bands are marked with arrowheads and arrows, respectively. Immunoblots are representative of six (a) or three (b) blots obtained with different nodule samples, except that in a few cases nitration intensity was similar in Y and M nodules of bean plants grown on 0.25 mM NH₄NO₃.