Structure of a [2Fe–2S] ferredoxin from Rhodobacter capsulatus likely involved in Fe–S cluster biogenesis and conformational changes observed upon reduction

Abstract

FdVI from Rhodobacter capsulatus is structurally related to a group of [2Fe-2S] ferredoxins involved in iron-sulfur cluster biosynthesis. Comparative genomics suggested that FdVI and orthologs found in alpha-Proteobacteria are involved in this process. Here, the crystal structure of FdVI has been determined for both the oxidized and the reduced protein. The [2Fe-2S] cluster lies 6 A below the protein surface in a hydrophobic pocket without access to the solvent. This particular cluster environment might explain why the FdVI midpoint redox potential (-306 mV at pH 8.0) did not show temperature or ionic strength dependence. Besides the four cysteines that bind the cluster, FdVI features an extra cysteine which is located close to the S1 atom of the cluster and is oriented in a position such that its thiol group points towards the solvent. Upon reduction, the general fold of the polypeptide chain was almost unchanged. The [2Fe-2S] cluster underwent a conformational change from a planar to a distorted lozenge. In the vicinity of the cluster, the side chain of Met24 was rotated by 180 degrees , bringing its S atom within hydrogen-bonding distance of the S2 atom of the cluster. The reduced molecule also featured a higher content of bound water molecules, and more extensive hydrogen-bonding networks compared with the oxidized molecule. The unique conformational changes observed in FdVI upon reduction are discussed in the light of structural studies performed on related ferredoxins.

Molecular Characterization of a Class I P450 Electron Transfer System from Novosphingobium aromaticivorans DSM12444

“…Cysteine residues are also found at the equivalent position to Cys 43 of Arx in Fe-S cluster biogenic ferredoxins, e.g. FDVI from Rhodobacter capsulatus (56) and Fdx from E. coli (57), and in the P450-associated ferredoxin terpredoxin (58). The corresponding residue in both Pdx and Pux is alanine while it is a leucine in Adx (24,29,59).…”

Functional Characterization of Fdx1: Evidence for an Evolutionary Relationship between P450-Type and ISC-Type Ferredoxins

“…This additional cysteine residue between the first and the second cluster-binding cysteine residues was proposed to be characteristic for ISC-type ferredoxins. 17 It was suggested that this additional cysteine residue might be involved in the binding and reduction of S 0 and/or Fe 3+ . 32 However, there were also ISC-type ferredoxins reported that are lacking this special cysteine residue (e.g., Etp1 fd from S. pombe, 13 FdxB from Pseudomonas putida 33 or human Fdx2 8 ).…”

“…Another interaction distinguishing vertebrate-type and plant-type ferredoxins is the hydrogen bond between the cluster-binding C44 and T46 (in Fdx1), which is also found only in the former class of ferredoxins. 17 The obligate presence of this OH-S bond explains the requirement of a threonine or a serine residue at the respective position. Apart from T46, three more residues are highly but not absolutely conserved.…”

“…cellulosum So ce56 is a soil-dwelling Gramnegative bacterium, whose genome sequence was published recently. 15 Its genome contains eight fdx genes 16 including one gene (sce7334) displaying the ISC-type motif C-X (3) -C-X-C-X (2) -C-X (∼ 35-38) -C, 17 and being part of the putative ISC operon in S. cellulosum So ce56, suggesting that Fdx1 might play a role in iron-sulfur cluster assembly. 16 In this work, we present a detailed study of Fdx1 from the myxobacterium S. cellulosum So ce56 and prove its functions as an ISC-associated ferredoxin.…”

Structural and thermodynamic characterization of the adrenodoxin-like domain of the electron-transfer protein Etp1 from Schizosaccharomyces pombe

“…So far, no crystal structure of an eukaryotic adrenodoxin-type ferredoxin involved in heme A synthesis and assembly of iron-sulfur clusters has been described, whereas three structures of bacterial proteins with corresponding function are already solved [13][14][15][16]. The corresponding E. coli ferredoxin gene was shown to be crucial for the efficient biosynthesis of Fe-S clusters in vivo [17,18], and the encoded protein was proven to support in vitro assembly of Fe-S clusters [19].…”