Structure of Escherichia coli Flavodiiron Nitric Oxide Reductase

The multidomain flavodiiron protein from Clostridium difficile 630 is an NADH:oxygen oxidoreductase

“…By reverse phase HPLC the flavin types were identified (data not shown): while the whole protein contains both FAD and FMN, in a close 1:1 ratio, the C-terminal domain contains only FAD. This result is in concordance with what is known for FDPs, that contain FMN in the flavodoxin core domain (e.g., 1 , 30 , 31 ), while NADH:rubredoxin oxidoreductases have been reported to possess FAD 23 – 25 .…”

“…The spectroscopic (EPR and UV-Visible) features of C. difficile FDP_F are characteristic of the types of centers present in FDPs, a major difference being the apparent absence of a stable semiquinone form of the FMN, which was not detected either by dithionite or NADH successive reduction of the enzyme. Interestingly, the amino acid sequence of FDP_F (as well as of all FDPs from this Class), shows that the arginine that establishes a hydrogen bond, through its main chain amino group, to the N5 of the isoalloxazine moiety of the FMN, and which has been considered responsible for the formation of the red semiquinone in the FDPs that contain this residue (e.g., 30 ), is absent in FDP_F (Fig. S1 ).…”

Krypton-derivatization highlights O2-channeling in a four-electron reducing oxidase

“…One main difference between the reduced and oxidized states is the loop 24 to 28 pointing from a b-hairpin in the b-lactamase-like domain which is unique in FDPs and covers the active site. 20 This loop is observed in the closed position in both tFprA structures as observed in the active reduced state of mFprA ( Fig. S4, ESI †).…”

“…Thus, a hydrophobic channel would make more sense to capture O 2 molecules with high specificity and efficiency, as it was predicted and calculated by Romão and co-workers. 20 To characterize experimentally this possible gas channel, we produced Kr-derivatives of oxidized tFprA by flash cooling crystals under gas pressure and collected datasets below and above the Kr K-edge (SI Method and Table S1, ESI †). Anomalous signal coming unambiguously from Kr atoms have been detected in an anomalous Fourier map at similar positions in each of the eight monomers constituting the asymmetric unit (Fig.…”

Functional and protective hole hopping in metalloenzymes

“…One of these chains is conserved among the ve enzymes, extending from the diiron site to the surface near the FMN cofactor. 126,129 The FDP enzyme from Methanothermobacter marburgensis has an extensive network of interconnected Trp and Tyr residues (PDB ID 2OHH, Fig. 9).…”