Immunological comparisons of higher plant nitrate reductases

Abstract

Monospecific antiserum raised against barley nitrate reductase was used to investigate the immunological relationship of barley nitrate reductase to nitrate reductases from nine higher plant species (hexaploid, tetraploid, and diploid wheat, rye, oat, maize, tobacco, soybean and pea). Rocket immunoelectrophoresis and antiserum titration were used to estimate antigenicity. The nitrate reductase from the monocotyledonous species appeared to be antigenically similar to the barley nitrate reductase. The wheat nitrate reductase could not be distinguished from the barley nitrate reductase by rocket immunoelectrophoresis or antiserum titration. The rye nitrate reductase was slightly different, while the oat and maize nitrate reductases were easily differentiated from the barley nitrate reductase. Nitrate reductases of the dicotyledonous species were different from the barley nitrate reductase as indicated by their inability to form rockets with the barley nitrate reductase antiserum. Nitrate reductases from all species were inactivated by the barley nitrate reductase antiserum, but required different amounts of the antiserum. The nitrate reductase from the monocotyledonous species required 1–1.5-fold more antiserum for 50% inactivation than required for 50% inactivation of equivalent amount of barley nitrate reductase. In contrast, the nitrate reductase from tobacco, soybean, and pea required 2-, 7-, and 10-fold more antiserum than barley for 50% inactivation, respectively. Thus, structural differences had occurred among these higher plant nitrate reductases but the active site appears to be highly conserved.