Role of Divalent Cations in Infections in Host-Pathogen Interaction.

With increasing numbers of patients worldwide diagnosed with diabetes mellitus, renal disease, and iatrogenic immune deficiencies, an increased understanding of the role of electrolyte interactions in mitigating pathogen virulence is necessary. The levels of divalent cations affect host susceptibility and pathogen survival in persons with relative immune insufficiency. For instance, when host cellular levels of calcium are high compared to magnesium, this relationship contributes to insulin resistance and triples the risk of clinical tuberculosis. The movement of divalent cations within intracellular spaces contributes to the host defense, causing apoptosis or autophagy of the pathogen. The control of divalent cation flow is dependent in part upon the mammalian natural resistance-associated macrophage protein (NRAMP) in the host. Survival of pathogens such as M tuberculosis within the bronchoalveolar macrophage is also dependent upon NRAMP. Pathogens evolve mutations to control the movement of calcium through external and internal channels. The host NRAMP as a metal transporter competes for divalent cations with the pathogen NRAMP in M tuberculosis (whether in latent, dormant, or active phase). This review paper summarizes mechanisms of pathogen offense and patient defense using inflow and efflux through divalent cation channels under the influence of parathyroid hormone vitamin D and calcitonin.

Divalent and multivalent cations control liquid-like assembly of poly(ADP-ribosyl)ated PARP1 into multimolecular associates in vitro.

The formation of nuclear biomolecular condensates is often associated with local accumulation of proteins at a site of DNA damage. The key role in the formation of DNA repair foci belongs to PARP1, which is a sensor of DNA damage and catalyzes the synthesis of poly(ADP-ribose) attracting repair factors. We show here that biogenic cations such as Mg2+, Ca2+, Mn2+, spermidine3+, or spermine4+ can induce liquid-like assembly of poly(ADP-ribosyl)ated [PARylated] PARP1 into multimolecular associates (hereafter: self-assembly). The self-assembly of PARylated PARP1 affects the level of its automodification and hydrolysis of poly(ADP-ribose) by poly(ADP-ribose) glycohydrolase (PARG). Furthermore, association of PARylated PARP1 with repair proteins strongly stimulates strand displacement DNA synthesis by DNA polymerase ß (Pol ß) but has no noticeable effect on DNA ligase III activity. Thus, liquid-like self-assembly of PARylated PARP1 may play a critical part in the regulation of i) its own activity, ii) PARG-dependent hydrolysis of poly(ADP-ribose), and iii) Pol ß-mediated DNA synthesis. The latter can be considered an additional factor influencing the choice between long-patch and short-patch DNA synthesis during repair.

Observing one-divalent-metal-ion-dependent and histidine-promoted His-Me family I-PpoI nuclease catalysis in crystallo.

Metal-ion-dependent nucleases play crucial roles in cellular defense and biotechnological applications. Time-resolved crystallography has resolved catalytic details of metal-ion-dependent DNA hydrolysis and synthesis, uncovering the essential roles of multiple metal ions during catalysis. The histidine-metal (His-Me) superfamily nucleases are renowned for binding one divalent metal ion and requiring a conserved histidine to promote catalysis. Many His-Me family nucleases, including homing endonucleases and Cas9 nuclease, have been adapted for biotechnological and biomedical applications. However, it remains unclear how the single metal ion in His-Me nucleases, together with the histidine, promotes water deprotonation, nucleophilic attack, and phosphodiester bond breakage. By observing DNA hydrolysis in crystallo with His-Me I-PpoI nuclease as a model system, we proved that only one divalent metal ion is required during its catalysis. Moreover, we uncovered several possible deprotonation pathways for the nucleophilic water. Interestingly, binding of the single metal ion and water deprotonation are concerted during catalysis. Our results reveal catalytic details of His-Me nucleases, which is distinct from multi-metal-ion-dependent DNA polymerases and nucleases.

Two-metal ion mechanism of DNA cleavage by activated, filamentous SgrAI.

Enzymes that form filamentous assemblies with modulated enzymatic activities have gained increasing attention in recent years. SgrAI is a sequence specific type II restriction endonuclease that forms polymeric filaments with accelerated DNA cleavage activity and expanded DNA sequence specificity. Prior studies have suggested a mechanistic model linking the structural changes accompanying SgrAI filamentation to its accelerated DNA cleavage activity. In this model, the conformational changes that are specific to filamentous SgrAI maximize contacts between different copies of the enzyme within the filament and create a second divalent cation binding site in each subunit, which in turn facilitates the DNA cleavage reaction. However, our understanding of the atomic mechanism of catalysis is incomplete. Herein, we present two new structures of filamentous SgrAI solved using cryo-EM. The first structure, resolved to 3.3 Å, is of filamentous SgrAI containing an active site mutation that is designed to stall the DNA cleavage reaction, which reveals the enzymatic configuration prior to DNA cleavage. The second structure, resolved to 3.1 Å, is of WT filamentous SgrAI containing cleaved substrate DNA, which reveals the enzymatic configuration at the end of the enzymatic cleavage reaction. Both structures contain the phosphate moiety at the cleavage site and the biologically relevant divalent cation cofactor Mg2+ and define how the Mg2+ cation reconfigures during enzymatic catalysis. The data support a model for the activation mechanism that involves binding of a second Mg2+ in the SgrAI active site as a direct result of filamentation induced conformational changes.

A Divalent Metal Cation-Metabolite Interaction Model Reveals Cation Buffering and Speciation.

I present the perspective that the divalent metalome and the metabolome can be modeled as a network of chelating interactions instead of separate entities. I review progress in understanding the complex cellular environment, in particular recent contributions to modeling metabolite-Mg2+ interactions. I then demonstrate a simple extension of these strategies based approximately on intracellular Escherichia coli concentrations. This model is composed of four divalent metal cations with a range of cellular concentrations and physical properties (Mg2+, Ca2+, Mn2+, and Zn2+), eight representative metabolites, and interaction constants. I applied this model to predict the speciation of divalent metal cations between free and metabolite-chelated species. This approach reveals potentially beneficial properties, including maintenance of free divalent metal cations at biologically relevant concentrations, buffering of free divalent metal cations, and enrichment of functional metabolite-chelated species. While currently limited by available interaction coefficients, this modeling strategy can be generalized to more complex systems. In summary, biochemists should consider the potential of cellular metabolites to form chelating interactions with divalent metal cations.

Structural and biochemical analysis of the unique interactions of the Campylobacter jejuni CorA channel protein with divalent cations.

CorA is a Mg2+ channel that plays a key role in the homeostasis of intracellular Mg2+ in bacteria and archaea. CorA consists of a cytoplasmic domain and a transmembrane domain and generates a Mg2+ pathway by forming a pentamer in the cell membrane. CorA gating is regulated via negative feedback by Mg2+, which is accommodated by the pentamerization interface of the CorA cytoplasmic domain (CorACD). The Mg2+-binding sites of CorACD differ depending on the species, suggesting that the Mg2+-binding modes and Mg2+-mediated gating mechanisms of CorA vary across prokaryotes. To define the Mg2+-binding mechanism of CorA in the Campylobacter jejuni pathogen, we structurally and biochemically characterized C. jejuni CorACD (cjCorACD). cjCorACD adopts a three-layered α/ß/α structure as observed in other CorA orthologs. Interestingly, cjCorACD exhibited enhanced thermostability in the presence of Ca2+, Ni2+, Zn2+, or Mn2+ in addition to Mg2+, indicating that cjCorACD interacts with diverse divalent cations. This cjCorACD stabilization is mediated by divalent cation accommodation by negatively charged residues located at the bottom of the cjCorACD structure away from the pentamerization interface. Consistently, cjCorACD exists as a monomer irrespective of the presence of divalent cations. We concluded that cjCorACD binds divalent cations in a unique pentamerization-independent manner.

Divalent cations promote huntingtin fibril formation on endoplasmic reticulum derived and model membranes.

Huntington's Disease (HD) is caused by an abnormal expansion of the polyglutamine (polyQ) domain within the first exon of the huntingtin protein (htt). This expansion promotes disease-related htt aggregation into amyloid fibrils and the formation of proteinaceous inclusion bodies within neurons. Fibril formation is a complex heterogenous process involving an array of aggregate species such as oligomers, protofibrils, and fibrils. In HD, structural abnormalities of membranes of several organelles develop. In particular, the accumulation of htt fibrils near the endoplasmic reticulum (ER) impinges upon the membrane, resulting in ER damage, altered dynamics, and leakage of Ca2+. Here, the aggregation of htt at a bilayer interface assembled from ER-derived liposomes was investigated, and fibril formation directly on these membranes was enhanced. Based on these observations, simplified model systems were used to investigate mechanisms associated with htt aggregation on ER membranes. As the ER-derived liposome fractions contained residual Ca2+, the role of divalent cations was also investigated. In the absence of lipids, divalent cations had minimal impact on htt structure and aggregation. However, the presence of Ca2+ or Mg2+ played a key role in promoting fibril formation on lipid membranes despite reduced htt insertion into and association with lipid interfaces, suggesting that the ability of divalent cations to promote fibril formation on membranes is mediated by induced changes to the lipid membrane physicochemical properties. With enhanced concentrations of intracellular calcium being a hallmark of HD, the ability of divalent cations to influence htt aggregation at lipid membranes may play a role in aggregation events that lead to organelle abnormalities associated with disease.

New Insights into the Dependence of CPEB3 Ribozyme Cleavage on Mn2+ and Mg2.

CPEB3 ribozyme is a self-cleaving RNA that occurs naturally in mammals and requires divalent metal ions for efficient activity. Ribozymes exhibit preferences for specific metal ions, but the exact differences in the catalytic mechanisms of various metal ions on the CPEB3 ribozyme remain unclear. Our findings reveal that Mn2+ functions as a more effective cofactor for CPEB3 ribozyme catalysis compared to Mg2+, as confirmed by its stronger binding affinity to CPEB3 by EPR. Cleavage assays of CPEB3 mutants and molecular docking analyses further showed that excessive Mn2+ ions can bind to a second binding site near the catalytic site, hindering CPEB3 catalytic efficiency and contributing to the Mn2+ bell-shaped curve. These results implicate a pivotal role for the local nucleobase-Mn2+ interactions in facilitating RNA folding and modulating the directed attack of nucleophilic reagents. Our study provides new insights and experimental evidence for exploring the divalent cation dependent cleavage mechanism of the CPEB3 ribozyme.

Competition between Stacking and Divalent Cation-Mediated Electrostatic Interactions Determines the Conformations of Short DNA Sequences.

Interplay between divalent cations (Mg2+ and Ca2+) and single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA), as well as stacking interactions, is important in nucleosome stability and phase separation in nucleic acids. Quantitative techniques accounting for ion-DNA interactions are needed to obtain insights into these and related problems. Toward this end, we created a sequence-dependent computational TIS-ION model that explicitly accounts for monovalent and divalent ions. Simulations of the rigid 24 base-pair (bp) dsDNA and flexible ssDNA sequences, dT30 and dA30, with varying amounts of the divalent cations show that the calculated excess number of ions around the dsDNA and ssDNA agree quantitatively with ion-counting experiments. Using an ensemble of all-atom structures generated from coarse-grained simulations, we calculated the small-angle X-ray scattering profiles, which are in excellent agreement with experiments. Although ion-counting experiments mask the differences between Mg2+ and Ca2+, we find that Mg2+ binds to the minor grooves and phosphate groups, whereas Ca2+ binds specifically to the minor groove. Both Mg2+ and Ca2+ exhibit a tendency to bind to the minor groove of DNA as opposed to the major groove. The dA30 conformations are dominated by stacking interactions, resulting in structures with considerable helical order. The near cancellation of the favorable stacking and unfavorable electrostatic interactions leads to dT30 populating an ensemble of heterogeneous conformations. The successful applications of the TIS-ION model are poised to confront many problems in DNA biophysics.

Klebsiella pneumoniae DedA family proteins have redundant roles in divalent cation homeostasis and resistance to phagocytosis.

The DedA superfamily is a highly conserved family of membrane proteins. Deletion of Escherichia coli yqjA and yghB, encoding related DedA family proteins, results in sensitivity to elevated temperature, antibiotics, and alkaline pH. The human pathogen Klebsiella pneumoniae possesses genes encoding DedA family proteins with >90% amino acid identity to E. coli YqjA and YghB. We hypothesized that the deletion of K. pneumoniae yqjA and yghB will impact its physiology and may reduce its virulence. The K. pneumoniae ΔyqjA ΔyghB mutant (strain VT101) displayed a growth defect at 42°C and alkaline pH sensitivity, not unlike its E. coli counterpart. However, VT101 retained mostly wild-type resistance to antibiotics. We found VT101 was sensitive to the chelating agent EDTA, the anionic detergent SDS, and agents capable of alkalizing the bacterial cytoplasm such as bicarbonate or chloroquine. We could restore growth at alkaline pH and at elevated temperature by addition of 0.5-2 mM Ca2+ or Mg2+ to the culture media. VT101 displayed a slower uptake of calcium, which was dependent upon calcium channel activity. VT201, with similar deletions as VT101 but derived from a virulent K. pneumoniae strain, was highly susceptible to phagocytosis by alveolar macrophages and displayed a defect in the production of capsule. These findings suggest divalent cation homeostasis and virulence are interlinked by common functions of the DedA family.IMPORTANCEKlebsiella pneumoniae is a dangerous human pathogen. The DedA protein family is found in all bacteria and is a membrane transporter often required for virulence and antibiotic resistance. K. pneumoniae possesses homologs of E. coli YqjA and YghB, with 60% amino acid identity and redundant functions, which we have previously shown to be required for tolerance to biocides and alkaline pH. A K. pneumoniae strain lacking yqjA and yghB was found to be sensitive to alkaline pH, elevated temperature, and EDTA/SDS and displayed a defect in calcium uptake. Sensitivity to these conditions was reversed by addition of calcium or magnesium to the growth medium. Introduction of ΔyqjA and ΔyghB mutations into virulent K. pneumoniae resulted in the loss of capsule, increased phagocytosis by macrophages, and a partial loss of virulence. These results show that targeting the Klebsiella DedA family results in impaired divalent cation transport and, in turn, loss of virulence.

Molecular determinants of ASIC1 modulation by divalent cations.

Acid-sensing ion channels (ASICs) are proton-gated cation channels widely expressed in the nervous system. ASIC gating is modulated by divalent cations as well as small molecules; however, the molecular determinants of gating modulation by divalent cations are not well understood. Previously, we identified two small molecules that bind to ASIC1a at a novel site in the acidic pocket and modulate ASIC1 gating in a manner broadly resembling divalent cations, raising the possibility that these small molecules may help to illuminate the molecular determinants of gating modulation by divalent cations. Here, we examined how these two groups of modulators might interact as well as mutational effects on ASIC1a gating and its modulation by divalent cations. Our results indicate that binding of divalent cations to an acidic pocket site plays a key role in gating modulation of the channel.

Group A Streptococcus cation diffusion facilitator proteins contribute to immune evasion by regulating intracellular metal concentrations.

Cation diffusion facilitators (CDFs) are a large family of divalent metal transporters with broad specificities that contribute to intracellular metal homeostasis and toxicity in bacterial pathogens. Streptococcus pyogenes (Group A Streptococcus [GAS]) expresses two homologous CDF efflux transporters, MntE and CzcD, which selectively transport Mn and Zn, respectively. We discovered that the MntE- and CzcD-deficient strains exhibited a marked decrease in the viability of macrophage-differentiated THP-1 cells and neutrophils. In addition, the viability of mice infected with both deficient strains markedly increased. Consistent with a previous study, our results suggest that MntE regulates the PerR-dependent oxidative stress response by maintaining intracellular Mn levels and contributing to the growth of GAS. The maturation and proteolytic activity of streptococcal cysteine protease (SpeB), an important virulence factor in GAS, has been reported to be abrogated by zinc and copper. Zn inhibited the maturation and proteolytic activity of SpeB in the culture supernatant of the CzcD-deficient strain. Furthermore, Mn inhibited SpeB maturation and proteolytic activity in a MntE-deficient strain. Since the host pathogenicity of the SpeB-deficient strain was significantly reduced, maintenance of intracellular manganese and zinc levels in the GAS via MntE and CzcD may not only confer metal resistance to the bacterium, but may also play an essential role in its virulence. These findings provide new insights into the molecular mechanisms of pathogenicity, which allow pathogens to survive under stressful conditions associated with elevated metal ion concentrations during host infection.

The structural basis of divalent cation block in a tetrameric prokaryotic sodium channel.

Divalent cation block is observed in various tetrameric ion channels. For blocking, a divalent cation is thought to bind in the ion pathway of the channel, but such block has not yet been directly observed. So, the behaviour of these blocking divalent cations remains still uncertain. Here, we elucidated the mechanism of the divalent cation block by reproducing the blocking effect into NavAb, a well-studied tetrameric sodium channel. Our crystal structures of NavAb mutants show that the mutations increasing the hydrophilicity of the inner vestibule of the pore domain enable a divalent cation to stack on the ion pathway. Furthermore, non-equilibrium molecular dynamics simulation showed that the stacking calcium ion repel sodium ion at the bottom of the selectivity filter. These results suggest the primary process of the divalent cation block mechanism in tetrameric cation channels.

Commentary on: Divalent metal cofactors differentially modulate RadA-mediated strand invasion and exchange in Saccharolobus solfataricus.

RecA ATPases are a family of proteins that catalyzes the exchange of complementary DNA regions via homologous recombination. They are conserved from bacteria to humans and are crucial for DNA damage repair and genetic diversity. In this work, Knadler et al. examine how ATP hydrolysis and divalent cations impact the recombinase activity of Saccharolobus solfataricus RadA protein (ssoRadA). They find that the ssoRadA-mediated strand exchange depends on ATPase activity. The presence of Manganese reduces ATPase activity and enhances strand exchange, while calcium inhibits ATPase activity by preventing ATP binding to the protein, yet destabilizes the nucleoprotein ssoRadA filaments, allowing strand exchange regardless of the ATPase activity. Although RecA ATPases are highly conserved, this research offers intriguing new evidence that each member of the family requires individual evaluation.

Free ferrous ions sustain activity of mammalian stearoyl-CoA desaturase-1.

Mammalian stearoyl-CoA desaturase-1 (SCD1) introduces a double-bond to a saturated long-chain fatty acid in a reaction catalyzed by a diiron center. The diiron center is well-coordinated by conserved histidine residues and is thought to remain with the enzyme. However, we find here that SCD1 progressively loses its activity during catalysis and becomes fully inactive after about nine turnovers. Further studies show that the inactivation of SCD1 is due to the loss of an iron (Fe) ion in the diiron center and that the addition of free ferrous ions (Fe2+) sustains the enzymatic activity. Using SCD1 labeled with Fe isotope, we further show that free Fe2+ is incorporated into the diiron center only during catalysis. We also discover that the diiron center in SCD1 has prominent electron paramagnetic resonance signals in its diferric state, indicative of distinct coupling between the two ferric ions. These results reveal that the diiron center in SCD1 is structurally dynamic during catalysis and that labile Fe2+ in cells could regulate SCD1 activity and hence lipid metabolism.

Assembly properties of bacterial actin MreB involved in Spiroplasma swimming motility.

Bacterial actin MreB forms filaments composed of antiparallel double-stranded units. The wall-less helical bacterium Spiroplasma has five MreB homologs (MreB1-5), some of which are involved in an intracellular ribbon for driving the bacterium's swimming motility. Although the interaction between MreB units is important for understanding Spiroplasma swimming, the interaction modes of each ribbon component are unclear. Here, we examined the assembly properties of Spiroplasma eriocheiris MreB5 (SpeMreB5), one of the ribbon component proteins that forms sheets. Electron microscopy revealed that sheet formation was inhibited under acidic conditions and bundle structures were formed under acidic and neutral conditions with low ionic strength. We also used solution assays and identified four properties of SpeMreB5 bundles as follows: (I) bundle formation followed sheet formation; (II) electrostatic interactions were required for bundle formation; (III) the positively charged and unstructured C-terminal region contributed to promoting lateral interactions for bundle formation; and (IV) bundle formation required Mg2+ at neutral pH but was inhibited by divalent cations under acidic pH conditions. During these studies, we also characterized two aggregation modes of SpeMreB5 with distinct responses to ATP. These properties will shed light on SpeMreB5 assembly dynamics at the molecular level.

Usnic Acid-Mediated Exchange of Protons for Divalent Metal Cations across Lipid Membranes: Relevance to Mitochondrial Uncoupling.

Usnic acid (UA), a unique lichen metabolite, is a protonophoric uncoupler of oxidative phosphorylation, widely known as a weight-loss dietary supplement. In contrast to conventional proton-shuttling mitochondrial uncouplers, UA was found to carry protons across lipid membranes via the induction of an electrogenic proton exchange for calcium or magnesium cations. Here, we evaluated the ability of various divalent metal cations to stimulate a proton transport through both planar and vesicular bilayer lipid membranes by measuring the transmembrane electrical current and fluorescence-detected pH gradient dissipation in pyranine-loaded liposomes, respectively. Thus, we obtained the following selectivity series of calcium, magnesium, zinc, manganese and copper cations: Zn2+ > Mn2+ > Mg2+ > Ca2+ >> Cu2+. Remarkably, Cu2+ appeared to suppress the UA-mediated proton transport in both lipid membrane systems. The data on the divalent metal cation/proton exchange were supported by circular dichroism spectroscopy of UA in the presence of the corresponding cations.

Molecular mechanisms underlying paracellular calcium and magnesium reabsorption in the proximal tubule and thick ascending limb.

Calcium and magnesium are the most abundant divalent cations in the body. The plasma level is controlled by coordinated interaction between intestinal absorption, reabsorption in the kidney, and, for calcium at least, bone storage and exchange. The kidney adjusts urinary excretion of these ions in response to alterations in their systemic concentration. Free ionized and anion-complexed calcium and magnesium are filtered at the glomerulus. The majority (i.e., >85%) of filtered divalent cations are reabsorbed via paracellular pathways from the proximal tubule and thick ascending limb (TAL) of the loop of Henle. Interestingly, the largest fraction of filtered calcium is reabsorbed from the proximal tubule (65%), while the largest fraction of filtered magnesium is reclaimed from the TAL (60%). The paracellular pathways mediating these fluxes are composed of tight junctional pores formed by claudins. In the proximal tubule, claudin-2 and claudin-12 confer calcium permeability, while the exact identity of the magnesium pore remains to be determined. Claudin-16 and claudin-19 contribute to the calcium and magnesium permeable pathway in the TAL. In this review, we discuss the data supporting these conclusions and speculate as to why there is greater fractional calcium reabsorption from the proximal tubule and greater fractional magnesium reabsorption from the TAL.

Calcium decreases cell wall swelling in sweet cherry fruit.

Swelling of epidermal cell walls decreases cell-to-cell adhesion and increases cracking susceptibility in sweet cherry. Ca is suggested to decrease cracking susceptibility by crosslinking of cell wall components and, possibly, by decreasing swelling. The objective is to test this hypothesis. The effect of Ca on swelling of anticlinal epidermal cell walls was quantified microscopically in vivo using excised skin sections and in vitro using extracted cell walls. After removal of turgor, cell wall thickness increased. Incubation in CaCl2 decreased cell wall thickness up to 3 mM CaCl2. At higher concentrations thickness remained constant. Decreased cell wall swelling in vivo also occurred with other salts of divalent and trivalent cations, but not with those of monovalent cations. Decreased swelling was due to the Ca cation, the anions had no effect. Ca also decreased swelling of cell walls that were already swollen. CaCl2 also decreased swelling of extracted cell walls in vitro. There was no effect on swelling pressure. The effect on swelling increased as the CaCl2 concentration increased. Chlorides of divalent and trivalent cations, but not those of monovalent cations decreased swelling in vitro. The decrease in swelling among the divalent cations was linearly related to the radius of the cation. The results indicate that Ca decreases cracking susceptibility by decreasing swelling.

Refolding and biophysical characterization of the Caulobacter crescentus copper resistance protein, PcoB: An outer membrane protein containing an intrinsically disordered domain.

Copper cations play fundamental roles in biological systems, such as protein folding and stabilization, or enzymatic reactions. Although copper is essential to the cell, it can become cytotoxic if present in too high concentration. Organisms have therefore developed specific regulation mechanisms towards copper. This is the case of the Pco system present in the bacterium Caulobacter crescentus, which is composed of two proteins: a soluble periplasmic protein PcoA and an outer membrane protein PcoB. PcoA oxidizes Cu+ to Cu2+, whereas PcoB is thought to be an efflux pump for Cu2+. While the PcoA protein has already been studied, very little is known about the structure and function of PcoB. In the present work, PcoB has been overexpressed in high yield in E. coli strains and successfully refolded by the SDS-cosolvent method. Binding to divalent cations has also been studied using several spectroscopic techniques. In addition, a three-dimensional structure model of PcoB, experimentally supported by circular dichroism, has been constructed, showing a ß-barrel conformation with a N-terminal disordered chain. This peculiar intrinsic disorder property has also been confirmed by various bioinformatic tools.