THE EFFECT OF LYSINE-MODIFYING AND ARGININE-MODIFYING REAGENTS ON SPINACH FERREDOXIN - NITRITE OXIDOREDUCTASE

Treatment of ferredoxin-dependent nitrite reductase, isolated from spinach leaves, with either the lysine-modifying reagent N-acetyl succinimide or the arginine-modifying reagent phenylglyoxal, caused substantial loss of enzymatic activity when reduced ferredoxin, the physiological donor, served as the source of electrons for nitrite reduction. In contrast, treatment with either modifier had little effect on the ability of the enzyme to catalyze nitrite reduction when a non-physiological electron donor, reduced methyl viologen, was supplied. Ferredoxin, but not nitrite, protected the enzyme against loss of ferredoxin-dependent activity caused by both modifiers. Modification caused only minor changes in the absorbance and circular dichroism spectra of nitrite reductase, suggesting that neither N-acetyl succinimide nor phenylglyoxal treatment causes a major conformational change in the enzyme. Modification of nitrite reductase with either N-acetyl succinimide or phenylglyoxal caused the enzyme to lose the ability to form a tight, electrostatically-stabilized complex with ferredoxin. These results have been interpreted in terms of the likely presence of both lysine- and arginine-residues at the ferredoxin-binding domain of spinach nitrite reductase.