Common conformational changes in flavodoxins induced by FMN and anion binding: The structure of Helicobacter pylori apoflavodoxin

Abstract

Flavodoxins, noncovalent complexes between apoflavodoxins and flavin mononucleotide (FMN), are useful models to investigate the mechanism of protein/flavin recognition. In this respect, the only available crystal structure of an apoflavodoxin (that from Anabaena) showed a closed isoalloxazine pocket and the presence of a bound phosphate ion, which posed many questions on the recognition mechanism and on the potential physiological role exerted by phosphate ions. To address these issues we report here the X-ray structure of the apoflavodoxin from the pathogen Helicobacter pylori. The protein naturally lacks one of the conserved aromatic residues that close the isoalloxazine pocket in Anabaena, and the structure has been determined in a medium lacking phosphate. In spite of these significant differences, the isoallozaxine pocket in H. pylori apoflavodoxin appears also closed and a chloride ion is bound at a native-like FMN phosphate site. It seems thus that it is a general characteristic of apoflavodoxins to display closed, non-native, isoalloxazine binding sites together with native-like, rather promiscuous, phosphate binding sites that can bear other available small anions present in solution. In this respect, both binding energy hot spots of the apoflavodoxin/FMN complex are initially unavailable to FMN binding and the specific spot for FMN recognition may depend on the dynamics of the two candidate regions. Molecular dynamics simulations show that the isoalloxazine binding loops are intrinsically flexible at physiological temperatures, thus facilitating the intercalation of the cofactor, and that their mobility is modulated by the anion bound at the phosphate site.

The Flavodoxin from Helicobacter pylori: Structural Determinants of Thermostability and FMN Cofactor Binding

“…The x-ray structure of the H. pylori protein reveals (18) that a tryptophan residue typically involved in cofactor binding in many flavodoxins appears replaced by an alanine, which creates a distinct pocket near the si-face of the redox cofactor (20). The structure of the apo form (PDB code: 2bmv) has also been recently determined (21) and it is similar to that of the holoprotein, except at the FMN binding site.…”

FtsH cleavage of non-native conformations of proteins

“…In principle, this different reactivity could be related to structural differences between the two forms, but a comparison of the X-ray structures of the apo and holo flavodoxins from Anabaena PCC 7119 (Genzor et al, 1996a;Rao et al, 1992) and from Helicobacter pylori (Freigang et al, 2002;Martinez-Julvez et al, 2007) reveals that the association of the FMN cofactor hardly induces structural changes in the apoprotein. Nevertheless, the bound cofactor makes the holo forms of these flavodoxins much more stable that their corresponding apo forms Cremades et al, 2008).…”

Folding of proteins with a flavodoxin‐like architecture

“…Again, FMN only binds to native apoflavodoxin, which strongly stabilizes the protein , thereby enabling it to be functional between pH 2 and 10 . Apoflavodoxin and flavodoxin are structurally similar, but in apoprotein, the loops binding the isoalloxazine are flexible . This flavodoxin strongly resembles A. vinelandii and Anabaena flavodoxins, but its FMN‐binding site is more accessible .…”

“…Apoflavodoxin and flavodoxin are structurally similar, but in apoprotein, the loops binding the isoalloxazine are flexible . This flavodoxin strongly resembles A. vinelandii and Anabaena flavodoxins, but its FMN‐binding site is more accessible . As a result, inhibitors can bind specifically to H. pylori flavodoxin and thereby disrupt its electron transfer function .…”