title: The ferredoxin-binding site of ferredoxin: Nitrite oxidoreductase. Differential chemical modification of the free enzyme and its complex with ferredoxin.
abstract: Spinach (Spinacea oleracea) leaf ferredoxin (Fd)-dependent nitrite reductase was treated with either the arginine-modifying reagent phenyl-glyoxal or the lysine-modifying reagent pyridoxal-5'-phosphate under conditions where only the Fd-binding affinity of the enzyme was affected and where complex formation between Fd and the enzyme prevented the inhibition by either reagent. Modification with [14C]phenylglyoxal allowed the identification of two nitrite reductase arginines, R375 and R556, that are protected by Fd against labeling. Modification of nitrite reductase with pyridoxal-5'-phosphate, followed by reduction with NaBH4, allowed the identification of a lysine, K436, that is protected by Fd against labeling. Positive charges are present at these positions in all of the Fd-dependent nitrite reductase for which sequences are available, suggesting that these amino acids are directly involved in electrostatic binding of Fd to the enzyme.
authors.affiliation: Department of Chemistry and Biochemistry, Texas Tech University, Lubbock 79409-1061, USA.
authors.forename: M M
authors.initials: MM
authors.lastname: Dose
authors.forename: M
authors.initials: M
authors.lastname: Hirasawa
authors.forename: S
authors.initials: S
authors.lastname: Kleis-SanFrancisco
authors.forename: E L
authors.initials: EL
authors.lastname: Lew
authors.forename: D B
authors.initials: DB
authors.lastname: Knaff
mesh_terms: D000595:Amino Acid Sequence; D001120:Arginine; D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D050900:Ferredoxin-Nitrite Reductase; D005288:Ferredoxins / Q000378:metabolism*; D008969:Molecular Sequence Data; D009572:Nitrite Reductases / Q000737:chemistry* / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D010658:Phenylglyoxal / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D018724:Spinacia oleracea / Q000201:enzymology*
chemical_list: D005288:Ferredoxins; D010446:Peptide Fragments; D001120:Arginine; D009572:Nitrite Reductases; D050900:Ferredoxin-Nitrite Reductase; D010658:Phenylglyoxal