Long-chain flavodoxin FldB from Escherichia coli.

Abstract

Flavodoxins are a family of small electron transferases widely distributed in prokaryotes. They utilize a noncovalently bound flavin mononucleotide (FMN) molecule as the redox center, and are able to switch between three different redox states, namely the oxidized (ox) state, the one-electron reduced semiquinone (sq) state, and the twoelectron reduced hydroquinone (hq) state (Knight and Hardy 1967). Since flavodoxins display two redox potentials and can transfer either one or two electrons at a time, they are functionally versatile. Flavodoxins have been identified to play important roles in various biological processes, including photosynthesis, methionine synthesis, biotin synthesis, and the activation of important enzymes such as pyruvate-formate lyase and ribonucleotide reductase (Sancho 2006). In eukaryotes, flavodoxins are integrated into multi-domain proteins and carry out similar redox functions. Flavodoxins can be further classified into the long-chain and short-chain subfamilies, which are distinguished based on the presence or absence of a 20-amino acid extra segment. The flavodoxin-like domains in eukaryotic multidomain proteins are more closely related to the short-chain subfamily. Previous investigations on the extra sequence suggested it is not directly involved in cofactor binding but may play a role in protein partner recognition and interactions, and it was proposed that the short-chain may derive from the long-chain group during evolution (LopezLlano et al. 2004a, b). However, the underlying molecular mechanism is yet unclear and the structural determinants for the distinct biological functions of different flavodoxins remain elusive. The Escherichia coli genome harbors several genes encoding flavodoxin proteins. Among them, the fldA gene encodes the well-characterized flavodoxin 1 protein which belongs to the long-chain subfamily and is essential for bacteria survival (Gaudu and Weiss 2000). The fldB gene encodes another long-chain flavodoxin which shares over 40 % sequence identity with FldA yet is functionally distinct. Insertion mutation of the fldB gene is not lethal to E. coli, and over expression of FldB could not substitute FldA (Gaudu and Weiss 2000). Evidence suggested that the fldB gene is a member of the superoxide response soxRS regulon, and its expression level was induced by paraquat (methyl viologen) (Gaudu and Weiss 2000). However, the exact function of FldB protein remains unclear. We have previously carried out structural and dynamic studies of E. coli flavodoxins including the short-chain MioC and YqcA proteins as well as the long-chain FldA by solution NMR spectroscopy (Hu et al. 2006; Ye et al. 2014). The FldA protein exhibit global conformational exchanges without the presence of FMN cofactor, rendering one third of its backbone amide signals missing in the NMR spectra, and we were unable to determine its solution structures in the apo-form. In contrast, the FldB protein Q. Ye W. Fu Y. Hu (&) C. Jin (&) Beijing Nuclear Magnetic Resonance Center, Peking University, Beijing 100871, China e-mail: yunfei@pku.edu.cn