Predicted nutrient assimilation pathways of carbon, nitrogen, phosphorous and sulfur in “<i>Ferrovum</i>” strain JA12.

<p>Inorganic phosphate is predicted to be taken up by the phosphate specific transport system (PST) and made available to the metabolism or stored as polyphosphate (P_n). A sulfate permease (SulP) is predicted to be responsible for the uptake of sulfate which then is activated by a sulfate adenylyltransferase to adenosine-phosphosulfate (APS) and subsequently reduced to sulfide. Carbon dioxide appears to be reduced to 3-phosphoglycerate in the Calvin-Benson-Bassham cycle indicated by its key enzyme RuBisCO. Bicarbonate may be fixed in the carboxysome using a carbonic anhydrase (CA) and a RuBisCO. “<i>Ferrovum</i>” strain JA12 is predicted to either take up ammonium directly by the ammonium transporter (AmtB) or reduce nitrate or nitrite to ammonium. No names for the nitrate and nitrite transporters and reductases are indicated due to contradicting nomenclature in the databases. Apparently, urea is taken up <i>via</i> an ABC transporter (Urt) and hydrolysed to ammonia and bicarbonate by the urease (UreABC). The subsequent spontaneous protonation of ammonia to ammonium at circum neutral pH reduces the proton concentration within the cytoplasm. The energy for all metabolic processes seems to be derived from the oxidation of ferrous iron.</p

Predicted electron transfer from ferrous iron to the terminal electron acceptors in “<i>Ferrovum</i>” strain JA12.

<p>Ferrous iron is probably oxidised at the cell surface by a high molecular mass cytochrome (Cyc2-like). The relevance of the identified high potential iron-sulfur protein (HiPIP) in this process remains unknown. The electrons appear to be further transferred <i>via</i> soluble <i>c</i>-type cytochromes in the periplasm either uphill driven by the proton motive force to the <i>bc</i>₁ complex and NADH dehydrogenase to produce reduction equivalents (NAD(P)H), or downhill <i>via</i> the <i>cbb</i>₃-type cytochrome <i>c</i> oxidase or alternatively <i>via</i> one of the quinol oxidases (cytochrome <i>bd</i> complex, cytochrome <i>bo</i>₃ ubiquinol oxidase) to the terminal acceptor oxygen. The reduction of oxygen to water at the terminal oxidases neutralises protons entering the cell during ATP synthesis <i>via</i> the ATP synthase (F₀F₁). The terminal oxidases <i>bo</i>₃ and <i>cbb</i>₃ pump protons into the periplasm driven by the downhill electron transfer to oxygen. Since the identity of the quinol derivates produced by “<i>Ferrovum</i>” strain JA12 was not investigated QH₂ represents the reduced form and Q the oxidised form of the quinol derivative, respectively. The genome also encodes other potential oxidoreductases which could channel electrons into the quinol pool (e.g. sulfide quinone oxidoreductase (SQR), electron transfer flavoprotein oxidoreductase (ETF), succinate dehydrogenase (SDH) of the citrate cycle).</p

Artificial circular plot of the “<i>Ferrovum</i>” strain JA12 genome.

<p>The three contigs of “<i>Ferrovum</i>” strain JA12 were concatenated to form an artificial circular genome in the order FERRO_contig000001, FERRO_contig000002 and FERRO_contig000003 (broken lines). The origin of replication is not indicated. G+C content and GC skew of the genome of “<i>Ferrovum</i>” strain JA12 are shown on ring 1 and 2 from the inside, respectively. The Blastn-based whole genome comparison of “<i>Ferrovum</i>” strain JA12 and “<i>F</i>. <i>myxofaciens</i>” P3G was conducted using BRIG [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0146832#pone.0146832.ref047" target="_blank">47</a>] with the genome sequence of “<i>Ferrovum</i>” strain JA12 set as reference (ring 3, orange). Blastn matches of “<i>F</i>. <i>myxofaciens</i>” P3G to the reference are shown on ring 4 (blue) with the colour intensity indicating the sequence identity of the match.</p

Urease-encoding gene clusters in “<i>Ferrovum</i>” strain JA12 and representative bacteria belonging to other taxa.

<p>The structure of the urease gene cluster in the genome of “<i>Ferrovum</i>” strain JA12 (A) was compared to urease encoding gene clusters of other urease active bacteria (B). With the exception of <i>Nitrosospira multiformis</i> ATCC 25196, which appears to use a urea-specific permease, urea is taken up <i>via</i> an ABC transport system (green), consisting of a periplasmic urea binding protein (<i>urtA</i>), two permeases (<i>urtBC</i>) and two ATP-hydrolysing enzymes (<i>urtDE</i>). The genes predicted to encode the three urease subunits (<i>ureABC</i>, orange), the Ni²⁺ transporter (<i>ureJ</i>, dark blue) and the urease accessory proteins (<i>ureH</i>, <i>ureEFG</i>, light blue) are often co-localised with the urea transporter. The accessory proteins encoded by <i>ureH</i> and <i>ureD</i> are thought to fulfil the same function during the formation of the active urease. Genes encoding other proteins involved in nitrogen metabolism are shown in grey. The location of potential promotors (red) were predicted using FGENESB.</p

Predicted stress management strategies in “<i>Ferrovum</i>” strain JA12.

<p>“<i>Ferrovum</i>” strain JA12 appears to employ strategies acting at different levels to maintain the intracellular pH homeostasis (A, green). Uncontrolled proton influx could be inhibited by incorporating cyclopropane fatty acyl phospholipids (Cfa) in the membrane or by building up an inside positive membrane potential by the increased uptake of K⁺-ions (Kef, FkuB). Apparently, the complexes of the respiratory chain (<i>bc</i>₁, <i>bo</i>₃, <i>cbb</i>₃) contribute to the active discharge of protons. Furthermore, “<i>Ferrovum</i>” strain JA12 is predicted to buffer protons by the decarboxylation of arginine (Adi), phosphatidylserine (Psd) or aspartate (PanD) and by the hydrolysis of urea <i>via</i> the urease (Ure). Damaged proteins appear to be restored by a number of chaperones. “<i>Ferrovum</i>” strain JA12 could cope with the high concentrations of metal and metalloid ions (B, light blue) either using specific systems like the copper efflux pump (CopA) or arsenate reductase (ArsC) and the arsenite efflux pump. Alternatively, more general multidrug transport systems of the RND family involving an efflux pump (RND), membrane fusions proteins (MFP) and an outer membrane protein (OMP) could be used to extrude metal ions. Oxidative stress (C, orange) is apparently managed by the detoxification of reactive oxygen species (ROS) using a superoxide dismutase (SOD) or thiol peroxidases and ferritin. Damaged proteins could be repaired in the cytoplasm using a thioredoxin (TrxA)/ thioredoxin reductase (TrxB)-dependent system or <i>via</i> the thiol:disulfide interchange proteins (DsbA, DsbD) in the periplasm. Damaged lipids may be restored by the peroxiredoxin (AhpC)/ alkyl hydroperoxide reductase (AhpF)-dependent system or by the glutathione peroxidase (Gpx). “<i>Ferrovum</i>” strain JA12 could use glutathione (GSH)-dependent systems to restore the redox balance, though it remains unclear how glutathione disulfide (GSSG) is recycled. A wide repertoire of DNA repair systems is predicted serve in the repair of damaged DNA (see also <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0146832#pone.0146832.s008" target="_blank">S3F Table</a>).</p