NONENZYMATIC COVALENT MODIFICATION OF HUMAN HEMOGLOBIN BY PYRIDOXAL-5-PHOSPHATE UNDER THE ACTION OF VISIBLE-LIGHT

Reaction of pyridoxal-5-phosphate (PLP) with apo-, oxy-, and deoxyhemo globin under the action of visible light at 4-20 degrees C at neutral pH gives rise to stable adducts, which withstand dialysis and Sephadex gel filtration. The yield of the adducts is increased with increasing illumination intensity and exposure time. Apohemoglobin was the most effective in stable adduct formation. With methemoglobin, this process was slowed down by visible light and addition of an electron acceptor , methylene blue. The adducts are strongly fluorescent, and the fluore scence parameters are identical to those for the stable PLP adducts fo rmed upon Schiff-base reduction with NaBH4. It is suggested that the s table adducts are formed owing to reduction of the Schiff-base aldimin e bond by light-ejected electrons. Electrophoregrams of the stable add ucts of hemoglobins and PLP formed under the action of visible light o r NaBH4 exhibit the same number of new bands with identical mobilities but different intensities. This is due to different electron-acceptin g properties of the Schiff bases formed by PLP and the alpha-NH2 group of Val-1 and the epsilon-NH2 groups of LYs residues of the alpha and beta chains of Hb.