title: Structural and chemical properties of a flavodoxin from Anabaena PCC 7119.
abstract: Structural and chemical properties of a flavodoxin from Anabaena PCC 7119 are described. The first 36 residues of the amino-terminal amino acid sequence have been determined and show extensive homology with flavodoxins isolated from other sources. Anabaena flavodoxin exhibits a net negative change (-3) in the helix-1 segment as found with other cyanobacterial flavodoxins Synechococcus 6301 (Anacystis nidulans) and Nostoc MAC, but in contrast to the net positive charge found in this region in the case of flavodoxins isolated from nitrogen-fixing bacteria (Azotobacter and Klebsiella). The FMN cofactor can be reversibly resolved from the apoprotein by trichloroacetic acid treatment. Apoflavodoxin, thus prepared, binds FMN with a Kd value of 0.1 nM and binds riboflavin with a decreased affinity (Kd = 5 microM) at pH 7.2. The apoprotein is stable in dilute solutions at pH values around 7 but readily denatures at pH 8 as judged from loss in flavin-binding ability and by ultraviolet circular dichroism spectroscopy. Oxidation-reduction potential studies at pH values of 7 and 8 show OX/SQ couples of -195 mV and -255 mV, respectively, and show SQ/HQ couples of -390 mV and -418 mV, respectively. From these data, the binding constant for the FMN semiquinone is calculated to be approx. 5-fold tighter and the binding of the FMN hydroquinone is approx. 10(5)-fold weaker than that of the oxidized FMN to the apoprotein. Anabaena flavodoxin functions as an effective mediator of electron transfer from ferredoxin-NADP(+)-reductase to cytochrome c with a turnover number [4.5-5) x 10(3) min-1); a values similar to that determined for Anabaena ferredoxin. The flavodoxin binds tightly to the reductase with Kd values of 6.4 and 8.5 microM at pH values of 7.0 and 8.0, respectively.
authors.affiliation: Departamento de Bioquimica y Biologia Molecular y Celular, Facultad de Ciencias, Universidad de Zaragoza, Spain.
authors.forename: M F
authors.initials: MF
authors.lastname: Fillat
authors.forename: D E
authors.initials: DE
authors.lastname: Edmondson
authors.forename: C
authors.initials: C
authors.lastname: Gomez-Moreno
mesh_terms: D000595:Amino Acid Sequence; D002942:Circular Dichroism; D000458:Cyanobacteria / Q000378:metabolism*; D005486:Flavin Mononucleotide / Q000378:metabolism; D005418:Flavodoxin / Q000235:genetics / Q000378:metabolism*; D005420:Flavoproteins / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D011485:Protein Binding; D011487:Protein Conformation; D012689:Sequence Homology, Nucleic Acid; D013050:Spectrometry, Fluorescence
chemical_list: D005418:Flavodoxin; D005420:Flavoproteins; D005486:Flavin Mononucleotide