A family of flavoproteins in the domains Archaea and Bacteria

Abstract

A family of flavoproteins, called A-type flavoproteins, is described. It consists of 14 protein sequences of 385Ϫ597 amino acids in length, 7 from methanogens (domain: Archaea), 5 from phototrophic prokaryotes, one from Escherichia coli, and a partial sequence from the sulfate reducer Desulfovibrio gigas (domain : Bacteria). No similar sequence could be found in the domain Eucarya. All sequences show significant similarity over a 385Ϫ400 amino acid portion overlapping a recognizable flavodoxin signature starting at positions 245Ϫ285 of the common core sequence. Cofactor analysis and, to some extent, analysis of the primary structure of six A-type flavoproteins, three of which are structurally characterized here, support the existence of four sub-families: Here we have aligned 14 sequences and FAD, which enables them to catalyze a variety of oneelectron and two-electron transfer reactions [1]. A number of and characterized the R. capsulatus protein and 2 additional members of this group of proteins as simple or complex flavoflavoproteins are known that contain additional redox centers like a disulfide bond, a heme group, a metal, or iron-sulfur cen-proteins. As a result, we propose to group them into a heterogeneous family, the A-type flavoproteins. ters [2]. Flavoproteins known to bind several cofactors, like cytochromes P-450, have distinct binding sites often located on separate protein domains [3Ϫ5].EXPERIMENTAL PROCEDURES We have recently purified an FMN-containing flavoprotein from cells of Methanobacterium thermoautotrophicum, flavoMaterials, Chemicals and enzymes. Restriction enzymes, protein A, which, like flavodoxins, is up-regulated under iron T4 DNA ligase, and Taq polymerase were purchased from and limitation and possesses a perfectly conserved flavodoxin signa-used as recommended by Fermentas. Oligonucleotide primers ture [6]. However, it differs from typical flavodoxins in being were synthesized at Microsynth. Q Sepharose Fast Flow, Sephmuch larger (a dimer of a 404-amino-acid polypeptide) and in acryl S-200 and S-300 HR were from Pharmacia, and Fractogel having a relatively high redox potential of Ϫ230 mV for the EMD-TMAE(650)-S from Merck (Darmstadt, Germany). Other quinone/hydroquinone couple [7]; so far, its function remains chemicals were obtained from Fluka (Buchs, Switzerland), and hypothetical [6Ϫ7]. At the time of its discovery, there was only were from the highest grade available. one flavoprotein clearly homologous to M. thermoautotroSequence analysis. Sequence analyses were carried out with phicum flavoprotein A, namely the product of an open reading the University of Wisconsin's Genetics Computer Group packframe in an operon involved in nitrogen fixation in Rhodobacter age, versions 8 and 9 and at the National Center for Biotechnolcapsulatus [8Ϫ9]. Since then, the completion of several se-ogy Information via the BLAST network service. quencing projects has permitted the discovery of ten predicted Amplification, cloning, and expression of E. coli orf479, polypeptide sequences unambiguou...

Functional Analysis of the Small Component of the 4-Hydroxyphenylacetate 3-Monooxygenase of Escherichia coli W: a Prototype of a New Flavin:NAD(P)H Reductase Subfamily

“…4) (42). It was suggested that these flavoproteins, which bind FAD and FMN at the same time in equimolecular amounts, might have evolved by the fusion of two flavin-binding domains located in the N-and C-terminus regions of the protein, showing different activities and functions (42).…”

“…4) (42). It was suggested that these flavoproteins, which bind FAD and FMN at the same time in equimolecular amounts, might have evolved by the fusion of two flavin-binding domains located in the N-and C-terminus regions of the protein, showing different activities and functions (42). The existence of a HpaC-like FMN binding domain in A-type flavoproteins suggests that a fusion between the reductase and oxygenase components of the enzymes of the TC-FDM family might already exist in nature or, at least, it would be feasible to design in vitro such monocomponent monooxygenase by protein engineering (unpublished data).…”

Operon flv4-flv2 Provides Cyanobacterial Photosystem II with Flexibility of Electron Transfer

“…Flavodiiron proteins (FDPs), originally known as A-type flavoproteins (Flv) (Wasserfallen et al, 1998), function in detoxification of O 2 and/or NO in many strict and facultative anaerobes (Vicente et al, 2008a). FDPs form a complex group of enzymes, and many of them have been thoroughly characterized.…”

Flavodiiron Protein Flv2/Flv4-Related Photoprotective Mechanism Dissipates Excitation Pressure of PSII in Cooperation with Phycobilisomes in Cyanobacteria

“…Flavodiiron proteins (FDPs), originally called A-type flavoproteins or Flvs (Wasserfallen et al, 1998), were recently demonstrated to have an important role in photoprotection of the photosynthetic machinery (Zhang et al, 2009Allahverdiyeva et al, 2011Allahverdiyeva et al, , 2013Ermakova et al, 2013). FDPs in general are most widespread among strict and facultative anaerobic bacteria.…”