MamA as a Model Protein for Structure-Based Insight into the Evolutionary Origins of Magnetotactic Bacteria

International audienceMamA is a highly conserved protein found in magnetotactic bacteria (MTB), a diverse group of prokaryotes capable of navigating according to magnetic fields - an ability known as magnetotaxis. Questions surround the acquisition of this magnetic navigation ability; namely, whether it arose through horizontal or vertical gene transfer. Though its exact function is unknown, MamA surrounds the magnetosome, the magnetic organelle embedding a biomineralised nanoparticle and responsible for magnetotaxis. Several structures for MamA from a variety of species have been determined and show a high degree of structural similarity. By determining the structure of MamA from Desulfovibrio magneticus RS-1 using X-ray crystallography, we have opened up the structure-sequence landscape. As such, this allows us to perform structural-and phylogenetic-based analyses using a variety of previously determined MamA from a diverse range of MTB species across various phylogenetic groups. We found that MamA has remained remarkably constant throughout evolution with minimal change between different taxa despite sequence variations. These findings, coupled with the generation of phylogenetic trees using both amino acid sequences and 16S rRNA, indicate that magnetotaxis likely did not spread via horizontal gene transfer and instead has a significantly earlier, primordial origin

Cation Diffusion Facilitators Transport Initiation and Regulation Is Mediated by Cation Induced Conformational Changes of the Cytoplasmic Domain

Cation diffusion facilitators (CDF) are part of a highly conserved protein family that maintains cellular divalent cation homeostasis in all domains of life. CDF's were shown to be involved in several human diseases, such as Type-II diabetes and neurodegenerative diseases. In this work, we employed a multi-disciplinary approach to study the activation mechanism of the CDF protein family. For this we used MamM, one of the main ion transporters of magnetosomes - bacterial organelles that enable magnetotactic bacteria to orientate along geomagnetic fields. Our results reveal that the cytosolic domain of MamM forms a stable dimer that undergoes distinct conformational changes upon divalent cation binding. MamM conformational change is associated with three metal binding sites that were identified and characterized. Altogether, our results provide a novel auto-regulation mode of action model in which the cytosolic domain's conformational changes upon ligand binding allows the priming of the CDF into its transport mode

The dual role of MamB in magnetosome membrane assembly and magnetite biomineralization

Magnetospirillum gryphiswaldense MSR‐1 synthesizes membrane‐enclosed magnetite (Fe$_3$O$_4$) nanoparticles, magnetosomes, for magnetotaxis. Formation of these organelles involves a complex process comprising key steps which are governed by specific magnetosome‐associated proteins. MamB, a cation diffusion facilitator (CDF) family member has been implicated in magnetosome‐directed iron transport. However, deletion mutagenesis studies revealed that MamB is essential for the formation of magnetosome membrane vesicles, but its precise role remains elusive. In this study, we employed a multi‐disciplinary approach to define the role of MamB during magnetosome formation. Using site‐directed mutagenesis complemented by structural analyses, fluorescence microscopy and cryo‐electron tomography, we show that MamB is most likely an active magnetosome‐directed transporter serving two distinct, yet essential functions. First, MamB initiates magnetosome vesicle formation in a transport‐independent process, probably by serving as a landmark protein. Second, MamB transport activity is required for magnetite nucleation. Furthermore, by determining the crystal structure of the MamB cytosolic C‐terminal domain, we also provide mechanistic insight into transport regulation. Additionally, we present evidence that magnetosome vesicle growth and chain formation are independent of magnetite nucleation and magnetic interactions respectively. Together, our data provide novel insight into the role of the key bifunctional magnetosome protein MamB, and the early steps of magnetosome formation

Purification of the M. magneticum Strain AMB-1 Magnetosome Associated Protein MamAΔ41

Magnetotactic bacteria comprise a diverse group of aquatic microorganisms that are able to orientate themselves along geomagnetic fields. This behavior is believed to aid their search for suitable environments (1). This capability is conferred by the magnetosome, a subcellular organelle that consists of a linear-chain assembly of lipid vesicles each able to biomineralize and enclose a ~50-nm crystal of magnetite or greigite. A principle component of the magnetosome that was shown to be required for the formation of functional vesicles is MamA. MamA is a highly abundant magnetosome-associated protein which is one of the most characterized magnetosome-associated proteins in vivo(2-6). This article focuses on the purification of MamA, which despite being studied in vivo, no clear functional or structural details have been identified for it. Bioinformatics analysis suggested that MamA is a tetra-tricopeptide repeat (TPR) containing protein. TPR is a structural motif found as such or forming part of a bigger fold in a wide range of proteins, it serves as a template for protein-protein interactions and mediates multi-protein complexes (7). TPRs are involved in many crucial tasks in eukaryotic cell organelle processes and many bacterial pathways (8-14). In order to understand MamA, a unique TPR containing protein, highly purified protein is required as a first step. In this article, we present the purification protocol for a stable MamA deletion mutant (MamAΔ41) from M. magneticum AMB-1

From conservation to structure, studies of magnetosome associated cation diffusion facilitators (CDF) proteins in Proteobacteria

International audienceMagnetotactic bacteria (MTB) are prokaryotes that sense the geomagnetic field lines to geo-locate and navigate in aquatic sediments. They are polyphyletically distributed in several bacterial divisions but are mainly represented in the Proteobacteria. In this phylum, magne-totactic Deltaproteobacteria represent the most ancestral class of MTB. Like all MTB, they synthesize membrane-enclosed magnetic nanoparticles, called magnetosomes, for magnetic sensing. Magnetosome biogenesis is a complex process involving a specific set of genes that are conserved across MTB. Two of the most conserved genes are mamB and mamM, that encode for the magnetosome-associated proteins and are homologous to the cation diffusion facilitator (CDF) protein family. In magnetotactic Alphaproteobacteria MTB species, MamB and MamM proteins have been well characterized and play a central role in iron-transport required for biomineralization. However, their structural conservation and their role in more ancestral groups of MTB like the Deltaproteobacteria have not been established. Here we studied magnetite cluster MamB and MamM cytosolic C-terminal domain (CTD) structures from a phylogenetically distant magnetotactic Deltaproteobacteria species represented by BW-1 strain, which has the unique ability to biomineralize magnetite and greigite. We characterized them in solution, analyzed their crystal structures and compared them to those characterized in Alphaproteobacteria MTB species. We showed that despite the high phylogenetic distance, MamB BW-1 and MamM BW-1 CTDs share high structural similarity with known CDF-CTDs and will probably share a common function with the Alphapro-teobacteria MamB and MamM

Correction: MamA as a Model Protein for Structure-Based Insight into the Evolutionary Origins of Magnetotactic Bacteria

<p>Correction: MamA as a Model Protein for Structure-Based Insight into the Evolutionary Origins of Magnetotactic Bacteria</p

ArsTM crystal packing, asymmetric unit composition and overall structure.

<p><b><i>(A)</i></b> ArsTM crystal packing and asymmetric unit composition. The molecules are shown in three rotation-related views. <b><i>(B)</i></b> Overlay of all six ArsTM monomers reveals the high degree of structural similarity. The representative ArsTM monomer contains five sequential TPR motifs. The molecule is shown in two views, related by a 90° rotation. <b><i>(C)</i></b> An overlay of representative monomers from ArsTM (green), MamAΔ41_Mbav (PDB ID: 3VTX, orange) and <i>Magnetospirillum</i> species MamAΔ41_AMB-1 (PDB ID: 3AS5 chain A and B in light pink and brown, respectively) related by a 180° rotation. A high structural similarity of MamAΔ41 between the species can be observed, apart from the helical conformation of the identical His-tag linker sequence remaining after thrombin proteolysis (H11: ELALVPR) seen in the 3AS5 chain B and 3VTX. In addition, a light flexibility is observed at the NTD of the monomers. All images were produced by PyMOL.</p