Wei-Jia Zhang

Structural insight into magnetochrome-mediated magnetite biomineralization

Magnetotactic bacteria align along the Earth’s magnetic field using an organelle called the magnetosome, a biomineralized magnetite (Fe(ii)Fe(iii)2O4) or greigite (Fe(ii)Fe(iii)2S4) crystal embedded in a lipid vesicle. Although the need for both iron(ii) and iron(iii) is clear, little is known about the biological mechanisms controlling their ratio. Here we present the structure of the magnetosome-associated protein MamP and find that it is built on a unique arrangement of a self-plugged PDZ domain fused to two magnetochrome domains, defining a new class of c-type cytochrome exclusively found in magnetotactic bacteria. Mutational analysis, enzyme kinetics, co-crystallization with iron(ii) and an in vitro MamP-assisted magnetite production assay establish MamP as an iron oxidase that contributes to the formation of iron(iii) ferrihydrite eventually required for magnetite crystal growth in vivo. These results demonstrate the molecular mechanisms of iron management taking place inside the magnetosome and highlight the role of magnetochrome in iron biomineralization.

Ferrous iron transport protein B gene (feoB1) plays an accessory role in magnetosome formation in Magnetospirillum gryphiswaldense strain MSR-1

To investigate the role of ferrous iron transport (Feo) systems in magnetosome formation, the gene for protein FeoB (feoB1), encoding 704 amino acids, was cloned from magnetotactic bacterium Magnetospirillum gryphiswaldense strain MSR-1. feoB1 constitutes a putative operon with feoA1, and the interval between the two genes is 36 base pairs. A feoB1-deficient mutant (DeltafeoB1) was constructed, and compared with wild-type in terms of iron uptake, iron content and functional complementation. Ferrous iron and ferric iron uptake in wild-type were respectively 1.8-fold and 1.3-fold higher than in the DeltafeoB1 mutant. Iron content (w/w) of DeltafeoB1 mutant was enhanced only slightly as extracellular iron concentration (either ferrous or ferric citrate) increased, whereas iron content of wild-type increased about 2-fold as extracellular iron concentration rose from 20 to 80 microM. Transmission electron microscopy revealed that DeltafeoB1 cells grown with either ferrous or ferric citrate produced fewer magnetosomes, with smaller diameter, compared to wild-type cells. Assay of feoAB1 promoter-lacZ transcriptional fusions indicated that the feoAB1 putative operon was downregulated when MSR-1 cells were grown under iron-rich condition. Magnetosome formation was reduced but not abolished in the feoB1 mutant, indicating that FeoB1 protein plays a significant role in this process. Other iron transport systems are presumed to be involved in iron uptake in MSR-1.

A novel genus of multicellular magnetotactic prokaryotes from the Yellow Sea

Multicellular magnetotactic prokaryotes (MMPs) are a group of magnetotactic microorganisms composed of 10-40 Gram-negative cells. Currently, all the identified MMPs show a spherical morphology and synthesize mainly iron sulfide magnetosomes. In this study, we report a novel genus of MMPs with peculiar ellipsoidal morphology and iron oxide magnetosomes, which were discovered in intertidal sediment of the Yellow Sea in China. Optical and fluorescence microscopy revealed that this organism was ~10 × 8 µm in size and composed of ~40 cells enveloped by an outer layer. Scanning electron microscopy showed that the cells were arranged in 4-6 interlaced circles. Bullet-shaped magnetite magnetosomes were organized in chains roughly parallel to the long axis of the ellipsoidal MMPs when analysed by transmission electron microscopy. These MMPs displayed special escape motility, i.e. swimming rapidly from the edge to the centre of the droplet and then slowly back to the edge. In addition, they exhibited negative phototaxis. Light microscopy observations showed that the ellipsoidal MMPs reproduced by division along the body long axis. Both analysis of 16S rRNA gene sequence and fluorescence in situ hybridization revealed the ellipsoidal MMPs as a new genus of the Deltaproteobacteria. In summary, this novel genus of MMPs exhibit unique morphology, peculiar division process and distinct phylogenetic affiliation compared with the other MMPs.

Comparative genomic analysis provides insights into the evolution and niche adaptation of marine Magnetospira sp. QH‐2 strain

Magnetotactic bacteria (MTB) are capable of synthesizing intracellular organelles, the magnetosomes, that are membrane-bounded magnetite or greigite crystals arranged in chains. Although MTB are widely spread in various ecosystems, few axenic cultures are available, and only freshwater Magnetospirillum spp. have been genetically analysed. Here, we present the complete genome sequence of a marine magnetotactic spirillum, Magnetospira sp. QH-2. The high number of repeats and transposable elements account for the differences in QH-2 genome structure compared with other relatives. Gene cluster synteny and gene correlation analyses indicate that the insertion of the magnetosome island in the QH-2 genome occurred after divergence between freshwater and marine magnetospirilla. The presence of a sodium-quinone reductase, sodium transporters and other functional genes are evidence of the adaptive evolution of Magnetospira sp. QH-2 to the marine ecosystem. Genes well conserved among freshwater magnetospirilla for nitrogen fixation and assimilatory nitrate respiration are absent from the QH-2 genome. Unlike freshwater Magnetospirillum spp., marine Magnetospira sp. QH-2 neither has TonB and TonB-dependent receptors nor does it grow on trace amounts of iron. Taken together, our results show a distinct, adaptive evolution of Magnetospira sp. QH-2 to marine sediments in comparison with its closely related freshwater counterparts.

Fur in Magnetospirillum gryphiswaldense Influences Magnetosomes Formation and Directly Regulates the Genes Involved in Iron and Oxygen Metabolism

Magnetospirillum gryphiswaldense strain MSR-1 has the unique capability of taking up large amounts of iron and synthesizing magnetosomes (intracellular magnetic particles composed of Fe3O4). The unusual high iron content of MSR-1 makes it a useful model for studying biological mechanisms of iron uptake and homeostasis. The ferric uptake regulator (Fur) protein plays a key role in maintaining iron homeostasis in many bacteria. We identified and characterized a fur-homologous gene (MGR_1314) in MSR-1. MGR_1314 was able to complement a fur mutant of E. coli in iron-responsive manner in vivo. We constructed a fur mutant strain of MSR-1. In comparison to wild-type MSR-1, the mutant strain had lower magnetosome formation, and was more sensitive to hydrogen peroxide and streptonigrin, indicating higher intracellular free iron content. Quantitative real-time RT-PCR and chromatin immunoprecipitation analyses indicated that Fur protein directly regulates expression of several key genes involved in iron transport and oxygen metabolism, in addition it also functions in magnetosome formation in M. gryphiswaldense.

Complex Spatial Organization and Flagellin Composition of Flagellar Propeller from Marine Magnetotactic Ovoid Strain MO-1

Marine magnetotactic ovoid bacterium MO-1 is capable of swimming along the geomagnetic field lines by means of its two sheathed flagellar bundles at a speed up to 300 μm/s. In this study, by using electron microscopy, we showed that, in each bundle, six individual flagella were organized in hexagon with a seventh in the middle. We identified 12 flagellin paralogs and 2 putative flagellins in the genome of MO-1. Among them, 13 were tandemly located on an ~ 17-kb segment while the 14th was on a separated locus. Using reverse transcription PCR and quantitative PCR, we found that all the 14 flagellin or putative flagellin genes were transcribed and that 2 of them were more abundantly expressed than others. A nLC (nanoliquid chromatography)-ESI (electrospray ionization)-MS/MS (mass spectrometry/mass spectrometry) mass spectrometry analysis identified all the 12 flagellin proteins in three glycosylated polypeptide bands resolved by one-dimensional denaturing polyacrylamide gel electrophoresis and 10 of them in 21 spots obtained by means of two-dimensional electrophoresis of flagellar extracts. Most spots contained more than one flagellin, and eight of the ten identified flagellins existed in multiple isoforms. Taken together, these results show unprecedented complexity in the spatial organization and flagellin composition of the flagellar propeller. Such architecture is observed only for ovoid-coccoid, bilophotrichously flagellated magnetotactic bacteria living in marine sediments, suggesting a species and environmental specificity.

Configuration of redox gradient determines magnetotactic polarity of the marine bacteria MO-1

Magnetotactic bacteria are capable of aligning and swimming along the geomagnetic field lines; such a behaviour is called magnetotaxis. Previous studies reported that bacteria in the northern hemisphere migrate preferentially towards the North Pole of the Earths magnetic field (north-seeking, NS), whereas those in the southern hemisphere swim towards the South Pole (south-seeking, SS). The orientated swimming is thought to guide bacteria migrating downward to the favourable microaerobic or anaerobic regions in stratified water column or sediments. Recent identification of SS populations in northern hemisphere challenged the model of the adaptive value of magnetotaxis. To seek explanation for the apparent discrepancy, we analysed magnetotaxis polarity of axenic cultures under simulated growth conditions in hypomagnetic, northern-hemisphere-like or southern-hemisphere-like magnetic fields. We found that NS and SS cells could obviously coexist in hypomagnetic field and even, when the oxidation-reduction gradient configuration is suitable, in the geomagnetic field. These results reveal the selectivity of the redox gradient configuration on magnetotactic polarity of the cells and reconcile the discrepancy of the early reports.

Complete Genome Sequence of Magnetospirillum gryphiswaldense MSR-1.

ABSTRACT We report the complete genomic sequence of Magnetospirillum gryphiswaldense MSR-1 (DSM 6361), a type strain of the genus Magnetospirillum belonging to the Alphaproteobacteria. Compared to the reported draft sequence, extensive rearrangements and differences were found, indicating high genomic flexibility and “domestication” by accelerated evolution of the strain upon repeated passaging.

Swimming behaviour and magnetotaxis function of the marine bacterium strain MO-1

Magnetotactic bacteria (MTB) have the unique capacity to align and swim along the geomagnetic field lines downward to the oxic-anoxic interface in chemically stratified water columns and sediments. They are most abundant within the first few centimetres of sediments below the water-sediment interface. It is unknown how MTB penetrate into the sediment layer and swim in the pocket water, while their movements are restricted by the alignment along the magnetic field lines. Here we characterized the swimming behaviour of the marine fast-swimming magnetotactic ovoid bacterium MO-1.We found that it rotates around and translates along its short body axis to the magnetic north (northward). MO-1 cells swim forward constantly for a minimum of 1770 μm without apparent stopping. When encountering obstacles, MO-1 cells squeeze through or swim southward to circumvent the obstacles. The distance of southward swimming is short and inversely proportional to the magnetic field strength. Using a magnetic shielding device, we provide direct evidence that magnetotaxis is beneficial to MO-1 growth and becomes essential at low cell density. Environmental implications of the fast-swimming magnetotactic behaviour of magnetococci are discussed.

Calcium ion-mediated assembly and function of glycosylated flagellar sheath of marine magnetotactic bacterium.

Flagella of some pathogens or marine microbes are sheathed by an apparent extension of the outer cell membrane. Although flagellar sheath has been reported for almost 60 years, little is known about its function and the mechanism of its assembly. Recently, we have observed a novel type of sheath that encloses a flagellar bundle, instead of a single flagellum, in a marine magnetotactic bacterium MO‐1. Here, we reported isolation and characterization of the sheath which can be described as a six‐start, right‐handed helical tubular structure with a diameter of about 100 nm, and a pitch of helix of about 260 nm. By proteomic, microscopic and immunolabelling analyses, we showed that the sheath of MO‐1 consists of glycoprotein with an apparent molecular mass > 350 kDa. This protein, named sheath‐associated protein (Sap), shows homology with bacterial adhesins and eukaryotic calcium‐dependent adherent proteins (cadherin). Most importantly, we showed that calcium ions mediate the assembly of the tubular‐shaped sheath and disintegration of the sheath was deleterious for smooth swimming of MO‐1 cells. The disintegrated sheath was efficiently reconstituted in vitro by adding calcium ions. Altogether, these results demonstrate a novel bacterial Ca2+‐dependent surface architecture, which is essential for bacterial swimming.