Mol Of Zinc Research Articles

Study of Addition Metal (Ti, Zn) Dopan on the Structure of NASICON as Solid Electrolyte Batteries

This study aims to analysized the effect of addition doped metal (Ti and Zn) on NASICON structure to morphology, materials structure, and electrochemical performance especially ionic conductivity properties. NASICON is a sodium super ionic conductor that it could be as solid electrolyte batteries. One of the problems that exist in the secondary battery is the low working temperature of the electrolyte, which makes it easy to explode when exposed to free air. The common electrolyte in liquid phase, so NASICON as replacement alternative. The synthesis method used is the solid-state reaction method by mixing sodium carbonate, silicon dioxide, zirconium oxide, ammonium dihydrogen phosphate, doped metal (titanium oxide and zinc oxide) and some anhydrous ethanol into a planetary ball mill, dried then calcined. Then the material is pressed to produce pellets and the sintered. The doping used varies from 0 to 5 mol% of titanium and zinc. XRD results showed that all variations in titanium doped had found rhombohedral and monoclinic. whereas in zinc doping also have those phase. The highest ionic conductivity is 7.8x10-3 S/m on 2% mol Zinc Addition

Identification of 7-Deoxy-desulfo-argino-cylindrospermopsin, the Missing Piece in Cylindrospermopsin Biosynthesis.

Cylindrospermopsin, a major cyanotoxin, is produced by freshwater cyanobacteria. Its biosynthesis starts from arginine and glycine and involves five polyketide synthases and several tailoring enzymes. We report the identification of 7-deoxy-desulfo-argino-cylindrospermopsin in several cylindrospermopsin-producing cyanobacteria using mass spectrometry experiments. We have purified this new metabolite and established its structure by 1D and 2D NMR spectroscopy using scalar-based 1H-1H, 1H-13C, and 1H-15N as well as 2D 1H-1H ROESY correlation experiments. Using labeled arginines in isotopic incorporation experiments, we have shown that arginine is fully incorporated into 7-deoxy-desulfo-argino-cylindrospermopsin and that the uracil ring of cylindrospermopsin originates from the guanidino moiety of arginine, thus solving a long-standing puzzling question. CyrG and CyrH from the cylindrospermopsin-producing Oscillatoria sp. PCC 6506 were overproduced in Escherichia coli and purified to homogeneity. We showed that CyrG is a zinc-dependent hydrolase, homologous to adenosine deaminases, that transforms 7-deoxy-desulfo-argino-cylindrospermopsin into 7-deoxy-desulfo-cylindrospermopsin and ornithine, with the following kinetic parameters: KM = 0.21 ± 0.05 μM and kcat = 0.19 ± 0.02 min-1. CyrG contained 0.55 mol of zinc per mol of monomer but could be activated by FeII or CoII. CyrH contained almost no metal and showed no such activity even in the presence of excess metal. Using structure-based alignments and secondary structure predictions, we propose that the fifth and last polyketide synthase CyrF in cylindrospermopsin biosynthesis contains an unprecedented C-terminal domain homologous to N-acetyltransferases. We suggest that this domain catalyzes the condensation of the CyrF product with arginine to give 7-deoxy-desulfo-argino-cylindrospermopsin. This would be an unprecedented termination step for a polyketide synthase.

A Study on Pulping of Paper Mulberry Bast Fiber Using Zinc Sulfate-Based Deep Eutectic Solvents

In this study, it was investigated whether deep eutectic solvents (DESs) can be produced through the eutectic reaction experiment between zinc sulfate (an inorganic compound) and three compounds including a choline chloride, histidine, and proline. As the results, it was found that the following two DESs can be manufactured: (1) the DES from the mixing of 1 mol zinc sulfate and 0.7 mol histidine (Zs-H DES) and (2) the DES from the mixing of 0.5 mol zinc sulfate and 1 mol proline (Zs-P DES). Also, in the further analysis, it was evaluated whether the pulping of paper mulberry bast fibers can be made using these two DESs. In particular, the effect of the mixing ratio of water in the two DESs on the pulping efficiency was investigated. As the results, it was found that the pulping efficiency by Zs-P DES was higher than that by Zs-H DES and that the pulping efficiency by two DESs was increased by the mixing water, It is notable that the efficiency was the highest when the mixing ratio of water was 30% for Zs-H DES and 50% for Zs-P DES.

A green approach to the microwave-assisted synthesis of flower-like ZnO nanostructures for reduction of Cr(VI)

Zinc oxide nanostructures with different morphologies were prepared via a microwave-assisted method and used as photocatalysts for reduction of chromium(VI) under ultraviolet-B illumination. X-ray diffraction revealed that as-made ZnO samples have wurtzite hexagonal structure. Scanning electron microscopy demonstrated that the morphology of the ZnO crystals could be easily modified from star-shaped to chrysanthemum flower-like structure by changing the ratio of zinc nitrate and sodium hydroxide. X-ray photoelectron spectroscopy and energy dispersive X-ray analysis confirmed the stoichiometry of pure ZnO. Its optical properties were also investigated by UV-Vis diffuse reflectance spectroscopy, which indicated that the prepared ZnO possessed a band gap value of 3.2 eV. Photocatalytic activity studies revealed that the ZnO (1:6) (1 × 10−3 mol zinc nitrate: 6 × 10−3 mol sodium hydroxide) catalyst exhibited a significantly faster reduction rate of Cr(VI) compared to ZnO (1:5) and ZnO (1:4). More than 95% of Cr(VI) at a concentration of 10 mg/L has been reduced to Cr(III).

Crosslinked Polyarylene Ether Nitrile Interpenetrating with Zinc Ion Bridged Graphene Sheet and Carbon Nanotube Network.

We report the fabrication and improved properties of crosslinked polyarylene ether nitrile (CPEN) interpenetrating with a zinc ion bridged graphene sheet (GS) and carbon nanotube (CNT) network (GS-Zn-CNT) (CPEN/GS-Zn-CNT). Graphene oxide (GO) and acidulated CNT were firstly prepared and then coordinated with zinc ions to form the zinc ion bridged GO and CNT network (GO-Zn-CNT). The mass ratio of GO and acidulated CNT in GO-Zn-CNT was controlled to be 1:3 and the optimized content of Zn2+ was Zn2+/C = 0.01 mmol/mg (mole of zinc acetate/total weight of GO and acidulated CNT). Phthalonitrile end-capped polyarylene ether nitrile (PEN-Ph) permeated into the GO-Zn-CNT in N-methyl-2-pyrrolidone (NMP) and the corresponding composite PEN/GO-Zn-CNT was fabricated through the solution-casting method. After thermal annealing at 230 °C for 1 h and further curing at 320 °C for 2 h, the GO in GO-Zn-CNT was partly reduced into GS, and PEN-Ph was crosslinked, offering the CPEN/GS-Zn-CNT. The mechanical, thermal, and electrical properties of the obtained CPEN/GS-Zn-CNT were investigated in detail. The glass transition temperature, relative permittivity, and tensile strength of CPEN/GS-Zn-CNT with 2.0 wt % GS-Zn-CNT, compared to that of PEN, were increased by 18%, 181%, and 27%, respectively. The CPEN/GS-Zn-CNT based composite is a potential candidate as material in high performance electronic devices.

The effect of citric acid on morphology and photoluminescence properties of white light emitting ZnO–SiO2 nanocomposites

The white light emitting ZnO–SiO2 nanocomposites were synthesized by sol-gel combustion method using zinc nitrate, citric acid and tetraethoxysilane. To analyze the effect of fuel content on the photoluminescence properties of ZnO–SiO2 nanocomposites, the citric acid content was varied as 1, 5, and 10 moles with respect to one mole of zinc. The SEM images of the nanocomposites revealed the spherical, flower and platelet like morphology with variation in citric acid content and annealing temperatures. The ZnO–SiO2 nanocomposites prepared with various Zn:CA ratio excited at UV (280nm), near UV (365nm), violet (405nm) and blue (465nm) wavelength showed blue and greenish–yellow emission. Among all ratios, the ZnO–SiO2 nanocomposites with Zn:CA – 1:1 ratio showed the intense broad band emission compared to Zn:CA – 1:5 and 1:10 values. This particular composition of sample excited under violet (405nm) LED source shows white light, as confirmed by the CIE chromaticity coordinates (x=0.342, y=0.318).

A novel thiolated human-like collage zinc complex as a promising zinc supplement: Physicochemical characteristics and biocompatibility

To improve zinc binding ability to human-like collagen (HLC) and stability of metal complex, HLC was thiolated by mercaptosuccinylation reaction with S-acetylmercaptosuccinic anhydride (S-AMSA) at pH8.0. One mole of thiolated HLC–Zn (SHLC–Zn) complex possessed 24.3mol zinc ions when pH was 8.0 and zinc concentration was 15mM. The physicochemical properties and biocompatibility of thiolated HLC–Zn (SHLC–Zn) complex were investigated by UV–vis, CD, electrophoresis analysis, differential scanning calorimetry (DSC) and cell viability assay, respectively. The results showed that SHLC–Zn complex(1) exhibited higher zinc ions than that of native HLC and still maintained the secondary structure of HLC though interaction occurred between SHLC and zinc ions, (2) increased the apparent molecular weight when compared with native HLC, (3) exhibited greater thermal stability than native HLC, and (4) presented toxicity free for BHK cells. This study suggests that the SHLC–Zn complex is a potential nutrition as well as zinc supplement in the medical application.

A 13C-NMR study of azacryptand complexes.

An azacryptand has been solubilised in aqueous media containing 50% (v/v) dimethyl sulphoxide. (13)C-NMR has been used to determine how the azacryptand is affected by zinc binding at pH 10. Using (13)C-NMR and (13)C-enriched bicarbonate we have been able to observe the formation of 4 different carbamate derivatives of the azacryptand at pH 10. The azacryptand was shown to solubilise zinc or cadmium at alkaline pHs. Two moles of zinc are bound per mole of azacryptand and this complex binds 1 mole of carbonate. By replacing the zinc with cadmium-113 we have shown that the (13)C-NMR signal of the (13)C-enriched carbon of the bound carbonate is split into two triplets at 2.2 °C. This shows that two cadmium complexes are formed and in each of these complexes the carbonate group is bound by two magnetically equivalent metal ions. It also demonstrates that these cadmium complexes are not in fast exchange. From temperature studies we show that in the zinc complexes both complexes are in fast exchange with each other but are in slow exchange with free bicarbonate. HOESY is used to determine the position of the carbonate carbon in the complex. The solution and crystal structures of the zinc-carbonate-azacryptand complexes are compared.

Effect of zinc chloride on the growth and characterization of L-proline cadmium chloride monohydrate semiorganic NLO single crystals

Single crystals of zinc doped L-proline cadmium chloride monohydrate were successfully grown from aqueous solution by slow evaporation method at room temperature for different molar concentration of zinc chloride. The structural properties of grown crystals have been studied by single crystal X-ray diffraction, powder X-ray diffraction studies and Fourier transform infrared spectral analysis. The incorporation of the dopant (zinc chloride) into L-proline cadmium chloride monohydrate crystal lattice has been confirmed by EDAX analysis. UV–Vis spectral analyses showed that the doped crystals have lower UV cut-off wavelength at 200nm combined with very good transparency about 85% in a very wide range. The second harmonic generation efficiency test has been carried out and results are discussed. The 0.2 and 0.4mol Zinc chloride doped crystals were thermally stable up to 208.9°C and 211.9°C respectively. The electrical properties have been studied by dielectric constant studies. All results are compared with the results of pure L-PCCM crystals.

Methane Gas Sensing Properties of the Zinc Oxide Nanowhisker-derived Gas Sensor

A low power methane gas sensor with microheater was fabricated by silicon bulk micromachining technology. In order to heat up the sensing layer to operating temperature, a platinum (Pt) micro heater was embedded in the gas sensor. The line width and gap of the microheater was 20 <TEX>${\mu}m$</TEX> and 4.5 <TEX>${\mu}m$</TEX>, respectively. Zinc oxide (ZnO) nanowhisker arrays were grown on a sensor from a ZnO seed layer using a hydrothermal method. A 200 ml aqueous solution of 0.1 mol zinc nitrate hexahydrate, 0.1 mol hexamethylenetetramine, and 0.02 mol polyethylenimine was used for growing ZnO nanowhiskers. Temperature distribution of the sensor was analyzed by infrared thermal camera. The optimum temperature for highest sensitivity was found to be <TEX>$250^{\circ}C$</TEX> although relatively high (64%) sensitivity was obtained even at as low a temperature as <TEX>$150^{\circ}C$</TEX>. The power consumption was 72 mW at <TEX>$250^{\circ}C$</TEX>, and only 25 mW at <TEX>$150^{\circ}C$</TEX>.

ZnO 나노휘스커 소재를 이용한 MEMS가스센서의 소비전력과 메탄 감응 특성 연구

A low power gas sensor with microheater was fabricated by MEMS technology. In order to heat up the gas sensing material to a operating temperature, a platinum(Pt) micro heater was built on to the micromachined Si substrate. The width and gap of microheater were and , respectively. ZnO nanowhisker arrays were fabricated on a sensor device by hydrothermal method. The sensor device was deposited with ZnO seeds using PLD systems. A 200 ml aqueous solution of 0.1 mol zinc nitrate hexahydrate, 0.1 mol hexamethylenetetramine, and 0.02 mol polyethylenimine was used for growthing ZnO nanowhiskers. The power consumption to heat up the gas sensor to a operating temperature was measured and temperature distribution of sensor was analyzed by a Infrared Thermal Camera. The optimum temperature for highest sensitivity was found to be although relatively high(64 %) sensitivity was obtained even at as low as . The power consumption was 72 mW at and was only 25 mW at .

Essential role of copper in the activity and regular periodicity of a recombinant, tumor-associated, cell surface, growth-related and time-keeping hydroquinone (NADH) oxidase with protein disulfide-thiol interchange activity (ENOX2)

ECTO-NOX proteins are growth-related cell surface proteins that catalyze both hydroquinone or NADH oxidation and protein disulfide interchange and exhibit time-keeping and prion-like properties. A bacterially expressed truncated recombinant 46kDa ENOX2 with full ENOX2 activity bound ca 2 moles copper and 2 moles of zinc per mole of protein. Unfolding of the protein in trifluoroacetic acid in the presence of the copper chelator bathocuproine resulted in reversible loss of both enzymatic activities and of a characteristic pattern in the Amide I to Amide II ratios determined by FTIR with restoration by added copper. The H546-V-H together with His 562 form one copper binding site and H582 represents a second copper site as determined from site-directed mutagenesis. Bound copper emerges as having an essential role in ENOX2 both for enzymatic activity and for the structural changes that underly the periodic alternations in activity that define the time-keeping cycle of the protein.

Purification and characterization of thermostable organic solvent-stable protease from Aeromonas veronii PG01

A mesophilic bacterium, Aeromonas veronii PG01, isolated from industrial wastes produced an extracellular thermostable organic solvent tolerant protease. The optimum condition for cell growth and protease production was pH 7.0 and 30 °C. The protease produced was purified 53-fold to homogeneity with overall yield of 32%, through ammonium sulphate precipitation, ion-exchange and gel permeation chromatography (GPC). The molecular weight, as determined by GPC–HPLC, was found to be about 67 kDa. SDS-PAGE revealed that the enzyme consisted of two subunits, with molecular weight of 33 kDa. The protease was active in broad range of pH from 6.0 to 10.0 with optimum activity at pH 7.5. The optimum temperature for this protease was 60 °C. The enzyme remained active after incubation at 50–60 °C for 1 h. This enzyme was stable and active after incubation with benzene and it was activated 1.3- and 1.5-fold by n-hexane and n-dodecane, respectively. This protease was inhibited completely by the classic metalloprotease inhibitor 1,10-phenanthroline and partially by the metal chelator EDTA but not by the serine protease inhibitor PMSF. The PG01 protease was found to contain 1.901 mol of zinc per mole of enzyme upon analysis by Inductively coupled plasma-optical emission spectroscopy. The thermostable and solvent tolerance property make it an attractive and promising biocatalyst for enzyme mediated synthesis.

Fibrolase: Trials and Tribulations

Fibrolase is the fibrinolytic enzyme isolated from Agkistrodon contortrix contortrix (southern copperhead snake) venom. The enzyme was purified by a three-step HPLC procedure and was shown to be homogeneous by standard criteria including reverse phase HPLC, molecular sieve chromatography and SDS-PAGE. The purified enzyme is a zinc metalloproteinase containing one mole of zinc. It is composed of 203 amino acids with a blocked amino-terminus due to cyclization of the terminal Gln residue. Fibrolase shares a significant degree of homology with enzymes of the reprolysin sub-family of metalloproteinases including an active site homology of close to 100%; it is rapidly inhibited by chelating agents such as EDTA, and by alpha2-macroglobulin (α2Μ). The enzyme is a direct-acting thrombolytic agent and does not rely on plasminogen for clot dissolution. Fibrolase rapidly cleaves the A(α)-chain of fibrinogen and the B(β)-chain at a slower rate; it has no activity on the γ-chain. The enzyme exhibits the same specificity with fibrin, cleaving the α-chain more rapidly than the β-chain. Fibrolase was shown to have very effective thrombolytic activity in a reoccluding carotid arterial thrombosis model in the canine. A recombinant version of the enzyme was made in yeast by Amgen, Inc. (Thousand Oaks, CA, USA) and called alfimeprase. Alfimeprase is identical to fibrolase except for a two amino acid truncation at the amino-terminus and the insertion of a new amino-terminal amino acid in the truncated protein; these changes lead to a more stable enzyme for prolonged storage. Alfimeprase was taken into clinical trials by Nuvelo, Inc. (San Carlos, CA), which licensed the enzyme from Amgen. Alfimeprase was successful in Phase I and II clinical trials for peripheral arterial occlusion (PAO) and central venous access device (CVAD) occlusion. However, in Phase III trials alfimeprase did not meet the expected end points in either PAO or CVAD occlusion and in a Phaase II stroke trial, and Nuvelo dropped further development in 2008.

An Fe2+-Dependent Cyclic Phosphodiesterase Catalyzes the Hydrolysis of 7,8-Dihydro-d-neopterin 2′,3′-Cyclic Phosphate in Methanopterin Biosynthesis

7,8-Dihydro-D-neopterin 2',3'-cyclic phosphate (H(2)N-cP) is the first intermediate in biosynthesis of the pterin portion of tetrahydromethanopterin (H(4)MPT), a C(1) carrier coenzyme first identified in the methanogenic archaea. This intermediate is produced from GTP by MptA (MJ0775 gene product), a new class of GTP cyclohydrolase I [Grochowski, L. L., Xu, H., Leung, K., and White, R. H. (2007) Biochemistry 46, 6658-6667]. Here we report the identification of a cyclic phosphodiesterase that hydrolyzes the cyclic phosphate of H(2)N-cP and converts it to a mixture of 7,8-dihydro-D-neopterin 2'-monophosphate and 7,8-dihydro-d-neopterin 3'-monophosphate. The enzyme from Methanocaldococcus jannachii is designated MptB (MJ0837 gene product) to indicate that it catalyzes the second step of the biosynthesis of methanopterin. MptB is a member of the HD domain superfamily of enzymes, which require divalent metals for activity. Direct metal analysis of the recombinant enzyme demonstrated that MptB contained 1.0 mol of zinc and 0.8 mol of iron per protomer. MptB requires Fe(2+) for activity, the same as observed for MptA. Thus the first two enzymes involved in H(4)MPT biosynthesis in the archaea are Fe(2+) dependent.

Trans-Dienelactone hydrolase from Pseudomonas reinekei MT1, a novel zinc-dependent hydrolase

Pseudomonas reinekei MT1 is capable of growing on 4- and 5-chlorosalicylate, involving a pathway with trans-dienelactone hydrolase ( trans-DLH) as a key enzyme. It acts on 4-chloromuconolactone formed during cycloisomerization of 3-chloromuconate by hydrolyzing it to maleylacetate. The gene encoding this activity was localized, sequenced and expressed in Escherichia coli. Inductively coupled plasma mass spectrometry showed that both the wild-type as well as recombinant enzymes contained 2 moles of zinc but variable amounts of manganese/mol of protein subunit. The inactive metal-free apoenzyme could be reactivated by Zn 2+ or Mn 2+. Thus, trans-DLH is a Zn 2+-dependent hydrolase using halosubstituted muconolactones and trans-dienelactone as substrates, where Mn 2+ can substitute for Zn 2+. It is the first member of COG1878 and PF04199 for which a direct physiological function has been reported.

Significance of Zinc in a Regulatory Protein, CCM1, Which Regulates the Carbon-Concentrating Mechanism in Chlamydomonas reinhardtii

In conditions with the poor availability of inorganic carbon (CO(2) and HCO(3) (-): Ci) for photosynthesis, aquatic photosynthetic organisms induce active Ci uptake systems that allow accumulation of Ci within the cell, the so-called carbon-concentrating mechanism (CCM). In a unicellular green alga, Chlamydomonas reinhardtii, a regulatory factor CCM1 is indispensable for the regulation of the CCM by sensing CO(2) availability. CCM1 has two putative zinc-binding domains with several conserved cysteine and histidine residues in its N-terminal region. To determine whether the domains actually bind zinc atoms, the N-terminal parts of CCM1 were expressed as glutathione S-transferase fusion proteins and subjected to atomic absorption spectrometry. It was found that 1 mol of zinc is bound to 1 mol of amino acid regions 1-71 and 72-101 of CCM1, respectively. In the case of the site-directed mutant proteins, H54Y, C77V and C80V, the zinc-binding ability was lost. Physiological analyses of the transgenic Chlamydomonas cells harboring a mutated Ccm1 gene revealed that amino acid residues such as C36, C41, H54, C77, C80, H90 and C93 were indispensable for induction of the CCM in response to Ci-limiting stress conditions. Size exclusion chromatography followed by immunoblot analyses indicated that CCM1 is present as a protein complex of approximately 290-580 kDa independent of Ci availability.

Synthesis of Azolyl Carboximidamides as Ligands for Zn(II) and Cu(II): Application of the Zn(II) Complexes as Catalysts for the Copolymerization of Carbon Dioxide and Epoxides1

A series of novel S,N-heterocyclic (thiazolyl) substituted carboximidamides 3 and 4 was synthesized in yields up to 82% from specific triazinium salts 1 and primary or secondary amines 2 which additionally bear pyridine or imidazole units. These carboximidamides are used as tailor-made ligands for the complexation of Cu(II) and Zn(II). The coordination behavior of 3 and 4 and the properties of the resulting metal complexes are affected a significant extent especially by the nature of these amine substituents. The most important structural feature of the novel complexes is that the ligation of the metal cations is achieved by a 1,3,5-triazapentadienyl anion system, compare the X-ray structure of the model complex Cu-4d. Analogous Zn(II) complexes 5, 6a, 6b, 6c, 7a, and 7b were obtained from carboximidamides 3, 4a, 4b, 4c, 4d, and 4e after reaction with diethylzinc. Interestingly, these Zn(II) complexes possess an intrinsic activity to catalyze the copolymerization of cyclohexene oxide and carbon dioxide to give polycarbonates 15 (TON up to 113; Turn Over Number: moles of substrate 14 consumed per moles of zinc. Molecular weights: up to 206.10(3) Da). Contaminations of 15 by polyethers are produced only in remarkably small amounts.

Glycolytic Aminolysis of Poly (Ethylene Terephthalate) Waste at Atmospheric Pressure for Recovery of a Value Added Insecticide

: Reaction of poly (ethylene terephthalate) waste (PETW) powder with ethylene glycol (EG) in the presence of tetrahydrofurane (THF) using 0.003 mol zinc acetate as a catalyst was carried out in a batch reactor at 470 K and at atmospheric pressure. Reactions were undertaken with various particle size ranges from 50–512.5 µm, and reaction time from 30–70 min at 10 min intervals for reaction of PETW. A low molecular weight product of PETW was obtained during this step. In the next stage hydroxylamine hydrochloride (HAHC), cyclohexylamine (CHA), and potasium hydroxide solution were introduced to convert the low molecular weight product of PETW into terephthalohydroxamic acid (TPHA) by introduction of HCl as per stoichiometric requirement. The TPHA is used as insecticide, and it is highly expensive. To increase the PETW conversion rate, an external catalyst (zinc acetate) was introduced during the reaction. The product was deposited on the surface of unreacted PETW that was removed from the surface by introducing dimethyl sulfoxide (DMSO). To accelerate the reaction rate DMSO, CHA, and THF were introduced during the reaction, which had an industrial significance. Depolymerization of PETW was proportional to the reaction time. Depolymerization of PETW was inversely proportional to the particle size of PETW. Analyses of value added products (TPHA and EG) as well as PETW were undertaken. A kinetic model was developed and experimental data simulated with it that were consistent with the model. A thermodynamic study was undertaken as this is required during transfer of laboratory data through a pilot plant for commercialization.

Redox-dependent Changes in RsrA, an Anti-sigma Factor in Streptomyces coelicolor: Zinc Release and Disulfide Bond Formation

σ R is a sigma factor for transcribing genes to defend cells against oxidative stresses in the antibiotic-producing bacterium Streptomyces coelicolor. The availability of σ R is regulated by RsrA, an anti-sigma factor, whose σ R-binding activity is regulated by redox changes in the environment, via thiol–disulfide exchange. We found that reduced RsrA contains zinc in a stoichiometric amount, whereas oxidized form has very little: 1 mol of zinc per mol of RsrA was released upon oxidation as monitored by a chromogenic Zn-chelator, 4-(2-pyridylazo)-resorcinol (PAR). Measurement of zinc bound in several RsrA mutants of various cysteine and histidine substitutions suggested that C3, H7, C41, and C44 serve as zinc-binding sites. The zinc-binding and σ R-binding activities of mutant proteins did not coincide, suggesting that zinc might not be absolutely required for the anti-sigma activity of RsrA. Zn-free apo-RsrA bound σ R and inhibited σ R-dependent transcription in vitro. Compared with Zn-RsrA, the anti-transcription activity of apo-RsrA was about threefold lower and its σ R-binding affinity decreased by about ninefold when measured by surface plasmon resonance analysis. Apo-RsrA was more sensitive to protease, suggesting that zinc allows RsrA to maintain a more compact structure, optimized for binding σ R. The cysteine pairs that form disulfide bonds were determined by MALDI-TOF mass spectrometry, revealing formation of the critical disulfide bond between C11 and one of the essential cysteine residues C41 or 44, most likely C44. An improved model for the mechanism of redox-modulation of RsrA was presented.