The crystal structure of Shethna protein II (FeSII) from Azotobacter vinelandii suggests a domain swap

Abstract

The Azotobacter vinelandii FeSII protein forms an oxygen-resistant complex with the nitrogenase MoFe and Fe proteins. FeSII is an adrenodoxin-type ferredoxin that forms a dimer in solution. Previously, the crystal structure was solved [Schlesier et al. (2016), J. Am. Chem. Soc. 138, 239-247] with five copies in the asymmetric unit. One copy is a normal adrenodoxin domain that forms a dimer with its crystallographic symmetry mate. The other four copies are in an `open' conformation with a loop flipped out exposing the 2Fe-2S cluster. The open and closed conformations were interpreted as oxidized and reduced, respectively, and the large conformational change in the open configuration allowed binding to nitrogenase. Here, the structure of FeSII was independently solved in the same crystal form. The positioning of the atoms in the unit cell is similar to the earlier report. However, the interpretation of the structure is different. The `open' conformation is interpreted as the product of a crystallization-induced domain swap. The 2Fe-2S cluster is not exposed to solvent, but in the crystal its interacting helix is replaced by the same helix residues from a crystal symmetry mate. The domain swap is complicated, as it is unusual in being in the middle of the protein rather than at a terminus, and it creates arrangements of molecules that can be interpreted in multiple ways. It is also cautioned that crystal structures should be interpreted in terms of the contents of the entire crystal rather than of one asymmetric unit.

Keywords: Azotobacter vinelandii; FeSII protein; Shethna protein II; domain swapping; nitrogen fixation; oxygen protection; structural biology.

Figure 1

The contents of two unit cells of the FeSII protein, showing the five unique chains A, B, C, D, E and A′, B′, C′, D′, E′ for the unit cell next to chain E.

Figure 2

The region around the 2Fe–2S cluster for closed and open chains. In chain E (salmon) in the crystal, the lid helix from chain E covers the iron–sulfur cluster. In chain C (magenta) the lid helix is flipped out in the open conformation, but the cluster is still capped by the lid helix from chain D. Weighted 2F_o − F_c density was calculated from the 1.65 Å resolution data set and contoured at 0.9σ; selected residues are labelled.

Figure 3

Cartoon and schematic view of the dimers of FeSII in the P2₁2₁2 crystal form. (a) Closed dimer (chains E and E′). (b) Open dimer (chains A and B). (c) Domain-swap dimer (chains A and B′). (d) Open and domain-swap dimers (A, B and B′). The chains are shown in blue and purple, with the lid helix (residues 68–79) in orange, except for the top lid helix when interpreted as part of the domain-swap dimer, when it is shown in red.