Characteristics of Modified Leghemoglobins lsolated from Soybean (Glycine max Merr.) Root Nodules

Authors

Hyung-Kyun Jun, University of Nebraska - Lincoln
Gautam Sarath, University of Nebraska–LincolnFollow
Jose F. Moran, Consejo Superior de lnvestigaciones Científicas
Manuel Becana, Consejo Superior de lnvestigaciones Científicas
Robert V. Klucas, University of Nebraska - Lincoln
Fred W. Wagner, University of Nebraska - LincolnFollow

ORCID IDs

Gautam Sarath

Date of this Version

1994

Citation

Plant Physiol. (1994) 104: 1231-1236

Comments

Copyright American Society of Plant Biologists. Used by permission.

Abstract

Hemoprotein derivatives of an abundant soybean (Glycine max Merr.) root nodule leghemoglobin, Lba, were studied for their modified spectral characteristics and physical properties. Three modified hemoprotein derivatives of Lba (Lbam₁, Lbam₂, and Lbam₃) were purified by preparative isoelectric focusing. The ferric forms of these pigments were green and exhibited anomalous spedra in the visible region as compared to the Lba³⁺ forms. These modified pigments showed a hypochromic shift of 10 nm for the charge transfer absorption maximum; however, differences were not apparent in the Sdret region. Upon binding with nicotinate, the α and β bands were shifted significantly into the red region as compared to the Lba³⁺ nicotinate complex. The three Lbam fractions were reduced by dithionite or by NADH in the presence of riboflavin. Lbam²⁺ also bound nicotinate and displayed absorption spectra indistinguishable from those of Lbam²⁺ nicotinate. In contrast to Lha²⁺, Lbam²⁺ displayed aberrant spectra when bound with either O₂ or CO. These complexes exhibited a prominent charge transfer band at approximately 620 nm and failed to exhibit spectra characteristic of Lba²⁺O₂ and Lba²⁺CO. The protein moiety of these modified pigments was intact because their tyrosineltryptophan ratios and their amino acid compositions were identical with those of Lba, nor were differences observed in the peptide profiles resulting from trypsin digests of purified Lba and Lbams. Automated Edman degradation of seleded peaks further confirmed the intactness of the protein backbone including the absence of deamination. Pyridine hemochromogen for heme from Lbams could be formed, and the spectra displayed distinct differences compared to those of Lba. A new peak at 580 nm and a loss of a peak at 480 nm were observed for all three Lbams.