Flavin-mediated reduction of ferric leghemoglobin from soybean nodules |
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Authors: | Manuel Becana Marvin L Salin Lin Ji Robert V Klucas |
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Institution: | (1) Department of Biochemistry, University of Nebraska, 68583 Lincoln, NE, USA;(2) Department of Biochemistry and Molecular Biology, Mississippi State University, 39762 Starkville, MS, USA;(3) Present address: Laboratorio de Fijación de Nitrógeno, E.E. de Aula Dei, Apdo. 202, 50080 Zaragoza, Spain |
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Abstract: | The reduction of ferric leghemoglobin (Lb3+) from soybean (Glycine max (L.) Merr.) nodules by riboflavin, FMN and FAD in the presence of NAD(P)H was studied in vitro. The system NAD(P)H + flavin reduced Lb3+ to oxyferrous (Lb2+ · O2) or deoxyferrous (Lb2+) leghemoglobin in aerobic or anaerobic conditions, respectively. In the absence of O2 the reaction was faster and more effective (i.e. less NAD(P)H oxidized per mole Lb3+ reduced) than in the presence of O2; this phenomenon was probably because O2 competes with Lb3+ for reductant, thus generating activated O2 species. The flavin-mediated reduction of Lb3+ did not entail production of superoxide or peroxide, indicating that NAD(P)H-reduced flavins were able to reduce Lb3+ directly. The NAD(P)H + flavin system also reduced the complexes Lb3+ · nicotinate and Lb3+ · acetate to Lb2+ · O2, Lb2+ or Lb2+ · nicotinate, depending on the concentrations of ligands and of O2. In the presence of 200 M nitrite most Lb remained as Lb3+ in aerobic conditions but the nitrosyl complex (Lb2+ · NO) was generated in anaerobic conditions. The above-mentioned characteristics of the NAD(P)H + flavin system, coupled with its effectiveness in reducing Lb3+ at physiological levels of NAD(P)H and flavins in soybean nodules, indicate that this mechanism may be especially important for reducing Lb3+ in vivo.Abbreviations and Terminology FLbR
ferric leghemoglobin reductase
- Hb2+ /Hb3+
hemoglobin containing Fe2+ /Fe2+
- Lb2+ /Lb3+
leghemoglobin containing Fe2+ /Fe3+
- Lb3+ · nicotinate/acetate
Lb in which nicotinate or acetate are complexed to Lb3+
- Lb2+ · O2/CO/NO/nicotinate
Lb in which O2, CO, NO or nicotinate are complexed to Lb2+
- Rfl
riboflavin
- SOD
superoxide dismutase (EC 1.15.1.1)
Published as Paper No. 9237, Journal Series, Nebraska Agricultural Research DivisionWe thank M.B. Crusellas for his skillful drawings. M. Becana thanks the Spanish Ministry of Education and Science/Fulbright Commission for financial support. |
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Keywords: | Flavin Glycine (nitrogen fixation) Leghemoglobin Nitrogen fixation Oxygen (activated) Pyridine nucleotides Symbiosis (legume-Rhizobium) |
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