DIRECT-DETECTION OF RADICALS IN INTACT SOYBEAN NODULES - PRESENCE OF NITRIC-OXIDE LEGHEMOGLOBIN COMPLEXES

Electron paramagnetic resonance spectroscopy has been employed to exam ine the nature of the metal ions and radicals present in intact root n odules of soybean plants grown in the absence of nitrate, The spectra obtained from nodules of different ages using this non-invasive techni que show dramatic differences, suggesting that there are both qualitat ive and quantitative changes in the metal ion and radical species pres ent. A major component of the spectra obtained from young nodules is a ssigned to a complex (Lb-NO) of nitric oxide (NO.) with the heme prote in leghemoglobin (Lb). This Lb-NO species, which has not been previous ly detected in intact root nodules of plants grown in the absence of n itrate, is thought to be formed by reaction of nitric oxide with iron( II) leghemoglobin. The nitric oxide may be generated from arginine via a nitric oxide synthase-like activity present in the nodules of the s oybean plants, in a manner analogous to that recently described for Lu pinus albus. This Lb-NO complex is present at lower concentrations in older nodules, and is almost completely absent from senescent nodules. Exposure of young and mature nodules to oxidant stress, in the form o f hydrogen peroxide, results in changes in the EPR spectra, with the l oss of the signals from the Lb-NO complex and appearance of absorption s similar to those from untreated senescent nodules. These results sug gest that there are characteristic changes in both the metal ion compl exes and radicals present in intact root nodules of different ages, an d support the theory that nitric oxide and other radicals play a signi ficant role in determining the nitrogen fixing activity of root nodule s; the modulatory activity of NO. may involve regulation of gene activ ity. (C) 1998 Elsevier Science Inc.