Identification of arginyl residues involved in the binding of ferredoxin-NADP+ reductase from Anabaena sp. PCC 7119 to its substrates.
Ferredoxin-NADP+ reductase from the cyanobacterium Anabaena sp. PCC 7119 was chemically modified by the alpha-dicarbonyl reagent phenylglyoxal. The studies of the inactivation by this compound, which is specific for arginyl residues, of both the diaphorase and NADPH-cytochrome c reductase activities, characteristic of the enzyme, are indicative of the involvement of at least one group of this kind in the binding site of NADP+ and a second one implicated in the interaction with ferredoxin. After specific cleavage of a FNR sample incubated with [7-14C]phenylglyoxal, two major labeled peptides were identified. The peptide which exhibited the higher degree of modification corresponded to residues 208-242. It contained four arginine residues but only two of them were the target of the modification: Arg224 and Arg233. Protection studies with protein substrates and sequence comparison with other reductases allow us to propose that these residues in Anabaena sp. PCC 7119 FNR must be involved in the interaction with the pyridine nucleotide. The second peptide corresponds to residues 75-103 and although it contains three arginine residues, Arg77 is the only one that exhibits the modification. This residue seems to be a key one in the interaction of this reductase with ferredoxin.
Medina
M
M
Departamento de Bioquímica y Biología Molecular y Celular, Facultad de Ciencias, Universidad de Zaragoza, Spain.
Mendez
E
E
Gomez-Moreno
C
C
D000595:Amino Acid Sequence; D017033:Anabaena / Q000201:enzymology*; D001120:Arginine / Q000737:chemistry; D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D005287:Ferredoxin-NADP Reductase / Q000737:chemistry / Q000378:metabolism*; D005288:Ferredoxins / Q000378:metabolism; D008969:Molecular Sequence Data; D009249:NADP / Q000378:metabolism; D017434:Protein Structure, Tertiary; D013057:Spectrum Analysis; D013329:Structure-Activity Relationship
D005288:Ferredoxins; D009249:NADP; D001120:Arginine; D005287:Ferredoxin-NADP Reductase