Reggie/Flottilin proteins are upregulated during neuronal regeneration and play crucial roles in various cell signaling processes. We have analyzed the genomes of prokaryotic organisms and found a large number of reggie-like genes. Remarkably, these reggie-like genes are very often accompanied by a gene of another transmembrane protein belonging to the diverse NfeD family, both of them organized in the same operon structure and thus co-expressed and co-regulated. This is a clear indication for a functional relationship of both genes and potentially even for a physical interaction of their respective proteins.
In B. subtilis, the reggie-like operon is regulated by the σ^W-factor leading to enhanced expression under conditions of cellular stress, like osmotic shock or presence of cell envelope-compromising antibiotics. It appears that reggie-like proteins (RLPs) and their associated partner proteins (RAPs) altogether act in response to these stress conditions.

Fig.1 (right): ¹H-¹⁵N-HSQC of the soluble C-terminus of YuaF from B. subtilis. Expression as MBP fusion yields soluble and well-behaved protein showing NMR spectra of excellent quality. The large dispersion in the HN dimension is characteristic of a protein rich β-sheet structure.

As a first step on the way to elucidate the functional network of these proteins, we have determined 3D structures in solution of the cytosolic portion of three different RAPs. Despite very low homology at the level of primary structure, all RAPs share a structural core composed of a five-stranded β-barrel, well known from the diverse class of OB-fold proteins that are usually involved in oligonucleotide and oligosaccharide binding. However, considering the pattern of conserved amino acids RAPs appear to be more likely involved in protein-protein interactions. These interactions may be responsible for co-localizing RAPs and RLPs in lipid microdomains as determined by fluorescence microscopy (Collaboration with Prof. P. Graumann, University of Freiburg).

Fig.2 (left): Solution structure of the cytoplasmic domain of YuaF from B. subtilis. Apparently, all SPFH-associated NfeD proteins adopt the OB-fold, a five-stranded β-barrel that is well known from oligosaccharide and oligonucleotide binding proteins. The N-terminal helix of YuaF presumably forms a spacer between the β-barrel core and the plasma membrane to which full-length YuaF is anchored by three transmembrane helices.

Fig.3: Hypothetical model of SPFH and NfeD co-evolution. In eukaryotes the SPFH protein superfamily shows a tremendous diversity. The low degree of sequence conservation made it impossible to establish a reliable phylogenetic tree of the eukaryotic SPFH proteins alone. However, when taking into account additionally the sequences of prokaryotic SPFH proteins, their operon structures, and the sequences of NfeD proteins that turn out to possess a conserved 3D structure, phylogenetic relationships can be proposed that explain the low sequence conservation of eukaryotic SPFH proteins by independent transitions from the prokaryotic to the eukaryotic world.

1. Walker, C.A.; Hinderhofer, M.; Witte, D.J.; Boos, W. and Möller*, H. M. Solution structure of the soluble domain of the NfeD protein YuaF from Bacillus subtilis. J. Biomol. NMR. 42, 69-76 (2008).


2. Hinderhofer, M.; Walker, C.A.; Wolf, J.; Friemel, A.; Boos, W. and Möller*, H. M. Resonance Assignment of the Soluble Domain of the NfeD Protein YqiJ from Eschericia coli. BMRB entry 15664 (2008).


3. Walker, C.A.; Hinderhofer, M.; Witte, D.J.; Boos, W. and Möller*, H. M. Solution structure of the soluble domain of the NfeD protein YuaF from Bacillus subtilis. RCSB PDB entry 2K14 (2008).


4. Walker, C.A.; Hinderhofer, M.; Witte, D.J.; Boos, W. and Möller*, H. M. Solution structure of the soluble domain of the NfeD protein YuaF from Bacillus subtilis. BMRB entry 15475 (2008).


5. Hinderhofer, M.; Walker, C. A.; Friemel, A.; Stuermer, C. A. O.; Möller^§, H. M. and Reuter^§*, A. Evolution of prokaryotic SPFH proteins. BMC Evolutionary Biology 9(10) (2009). (^§ equal contribution)