CHARACTERISTICS OF MODIFIED LEGHEMOGLOBINS ISOLATED FROM SOYBEAN (GLYCINE-MAX MERR) ROOT-NODULES

Hemoprotein derivatives of an abundant soybean (Glycine max Merr.) roo t nodule leghemoglobin, Lba, were studied for their modified spectral characteristics and physical properties. Three modified hemoprotein de rivatives of Lba (Lbam(1), Lbam(2), and Lbam(3)) were purified by prep arative isoelectric focusing. The ferric forms of these pigments were green and exhibited anomalous spectra in the visible region as compare d to the Lba(3+) forms. These modified pigments showed a hypochromic s hift of 10 nm for the charge transfer absorption maximum; however, dif ferences were not apparent in the Soret region. Upon binding with nico tinate, the alpha and beta bands were shifted significantly into the r ed region as compared to the Lba(3+) nicotinate complex. The three Lba m fractions were reduced by dithionite or by NADH in the presence of r iboflavin. Lbam(2+) also bound nicotinate and displayed absorption spe ctra indistinguishable from those of Lba(2+) nicotinate. In contrast t o Lba(2+), Lbam(2+) displayed aberrant spectra when bound with either O-2 Or CO. These complexes exhibited a prominent charge transfer band at approximately 620 nm and failed to exhibit spectra characteristic o f Lba(2+)O(2) and Lba(2+)CO. The protein moiety of these modified pigm ents was intact because their tyrosine/tryptophan ratios and their ami no acid compositions were identical with those of Lba, nor were differ ences observed in the peptide profiles resulting from trypsin digests of purified Lba and Lbams. Automated Edman degradation of selected pea ks 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 compa red 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.