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1.
Eva Freisinger 《Inorganica chimica acta》2007,360(1):369-380
Metallothioneins (MTs) are ubiquitous low molecular mass, cysteine-rich proteins with the ability to bind d10 metal ions in the form of metal-thiolate clusters. In contrast to the vertebrate forms, knowledge about the properties of members of the plant metallothionein family is still scarce. The amino acid sequences of plant MTs are distinctively different to the sequences of other MT species. The protein under investigation, Musa acuminata (banana) MT3, belongs to the plant MT fruit-specific p3 subfamily. With a total of 10 cysteine residues, MT3 features a cysteine content and percentage that is more comparable to fungal and prokaryotic MTs than to the well characterized mammalian iso-forms. The gene sequence encoding MT3 was cloned into a suitable vector and the protein was recombinantly overexpressed in Escherichia coli. MT3 is able to coordinate a maximum of four divalent d10 metal ions under the formation of metal-thiolate clusters. The hitherto unknown spectroscopic behavior of MT3 in combination with the metal ions Zn2+, Cd2+, Pb2+, and Hg2+ will be presented and gives rise to the existence of a weaker metal ion coordination site. The pH stability of the investigated zinc and cadmium clusters is comparable to the values found for other plant metallothioneins though significantly lower than for the mammalian iso-forms. Possible metal-thiolate cluster structures will additionally be discussed. 相似文献
2.
Ming Guan Junmin Li Xiaocui Pan Suizi Wang Yuelin Li 《International journal of phytoremediation》2016,18(4):368-377
The aim of this study was to investigate the effects of temperature and Cu on the morphological and physiological traits of Elsholtzia haichowensis grown in soils amended with four Cu concentrations (0, 50, 500, and 1000 mg kg?1) under ambient temperature and slight warming. At the same Cu concentration, the height, shoot dry weight, total plant dry weight, and root morphological parameters such as length, surface area and tip number of E. haichowensis increased due to the slight warming. The net photosynthetic rate, stomatal conductance, transpiration, light use efficiency were also higher under the slight warming than under ambient temperature. The increased Cu concentrations, total Cu uptake, bioaccumulation factors and tolerance indexes of shoots and roots were also observed at the slight warming. The shoot dry weight, root dry weight, total plant dry weight and the bioaccumulation factors of shoots and roots at 50 mg Cu kg?1 were significantly higher than those at 500 and 1000 mg Cu kg?1 under the slight warming. Therefore, the climate warming may improve the ability of E. haichowensis to phytoremediate Cu-contaminated soil, and the ability improvement greatly depended on the Cu concentrations in soils. 相似文献
3.
Kobayashi Y Kuroda K Kimura K Southron-Francis JL Furuzawa A Kimura K Iuchi S Kobayashi M Taylor GJ Koyama H 《Plant physiology》2008,148(2):969-980
Copper (Cu) is an essential element in plant nutrition, but it inhibits the growth of roots at low concentrations. Accessions of Arabidopsis (Arabidopsis thaliana) vary in their tolerance to Cu. To understand the molecular mechanism of Cu tolerance in Arabidopsis, we performed quantitative trait locus (QTL) analysis and accession studies. One major QTL on chromosome 1 (QTL1) explained 52% of the phenotypic variation in Cu tolerance in roots in a Landsberg erecta/Cape Verde Islands (Ler/Cvi) recombinant inbred population. This QTL regulates Cu translocation capacity and involves a Cu-transporting P1B-1-type ATPase, HMA5. The Cvi allele carries two amino acid substitutions in comparison with the Ler allele and is less functional than the Ler allele in Cu tolerance when judged by complementation assays using a T-DNA insertion mutant. Complementation assays of the ccc2 mutant of yeast using chimeric HMA5 proteins revealed that N923T of the Cvi allele, which was identified in the tightly conserved domain N(x)6YN(x)4P (where the former asparagine was substituted by threonine), is a cause of dysfunction of the Cvi HMA5 allele. Another dysfunctional HMA5 allele was identified in Chisdra-2, which showed Cu sensitivity and low capacity of Cu translocation from roots to shoots. A unique amino acid substitution of Chisdra-2 was identified in another strictly conserved domain, CPC(x)6P, where the latter proline was replaced with leucine. These results indicate that a portion of the variation in Cu tolerance of Arabidopsis is regulated by the functional integrity of the Cu-translocating ATPase, HMA5, and in particular the amino acid sequence in several strictly conserved motifs. 相似文献
4.
5.
Copper (Cu) accumulation and tolerance mechanisms in Elsholtzia haichowensis, an indicator plant of Cu mines, were investigated under hydroponics supplied with different concentrations (0.32, 50.0,
100.0 and 200.0 μM) of Cu for 8 days. Cu at 100 and 200 μM significantly decreased the root dry weight, but had no significant
effect on shoot dry weight. The plants grown in the presence of 200 μM Cu accumulated 288 and 7626 μg g−1 DW total Cu in the shoots and roots, respectively. A greater proportion of accumulated Cu was water-soluble accounting for
42–93% of the total Cu content in the shoots. The concentrations of reduced glutathione (GSH) and protein thiols were significantly
enhanced under excess Cu supply. However, the concentrations of these compounds, particularly protein thiols, were much higher
in the leaves than that in the roots. Three UV-absorbing peaks could be eluted out through gel filtration chromatography on
Sephadex G-50. A large amount of Cu was detected in the UV-absorbing peaks in 40–50 and 70–90 ml elution fractions of the
root extract, and in 40–50 and 120–140 ml elution fractions of the leaf extract. The results suggested that the adaptive Cu
tolerance mechanism in E. haichowensis might involve the active participation of protein thiols which had a more important role in the leaves than in the roots. 相似文献
6.
Oliver Schicht 《Inorganica chimica acta》2009,362(3):714-7306
The plant metallothioneins differ distinctively from other metallothionein families with respect to the cysteine distribution patterns, the presence of aromatic amino acids in most and histidine in some forms, as well as long cysteine-free amino acid stretches between cysteine-rich regions. Although known for more than 25 years, research activity on plant metallothioneins has been low increasing only in the past few years. In the following, we will present the first characterization of Cicer arietinum (chickpea) MT1. In this root-specific protein two cysteine-rich regions with six cysteine residues each are separated by a 42 amino acids long linker region. A synthetic gene encoding MT1 was designed, cloned into a suitable vector, and the protein was over-expressed in Escherichia coli. We could show, that MT1 has the ability to coordinate up to five Zn2+ or Cd2+ ions and even higher amounts of Hg2+. According to titration experiments pH-dependent zinc- and cadmium-thiolate cluster stability in MT1 is considerably lower than in vertebrate metallothioneins. The approximate contribution of secondary structural elements to the overall structure was assessed with circular dichroism and infrared spectroscopy. Hypothetical metal-thiolate cluster structures will be presented. 相似文献
7.
Ingrid Falnoga Andreja Zelenik Pevec Zdenka ?lejkovec Magda Tu?ek ?nidari? Irena Zajc Simona Jurkovi? Mlakar Janja Marc 《Biological trace element research》2012,149(3):331-339
Arsenic trioxide (As2O3; ATO, TRISENOX?) is used to treat patients with refractory or relapsed acute promyelocytic leukaemia while its application for treatment of solid cancers like glioblastoma is still under evaluation. In the present study, we investigated the interaction of arsenic trioxide with metallothionein (MT) isoforms as a possible (protective response) resistance of glioblastoma cells to arsenic-induced cytotoxicity. Special attention was focused on MT3, the isoform expressed mainly in the brain. MT3 has low metal inducibility, fast metal binding/releasing properties and outstanding neuronal inhibitory activity. The human astrocytoma (glioblastoma) cell line U87 MG was treated with 0.6, 2 and 6?C7???M arsenic (equivalent to 0.3, 1 and 3?C3.5???M As2O3) for 12, 24 or 48?h and gene expression for different MT isoforms, namely MT2A, MT1A, MT1F, MT1X, MT1E and MT3, was measured by real time qPCR using SYBR Green I and Taqman? gene expression assays. TfR, 18S rRNA, GAPDH and AB were tested as reference genes, and the last two evaluated to be appropriate in conditions of low (GAPDH) and high (AB) arsenic exposure. The gene expression of MT3 gene was additionally tested and confirmed by restriction enzyme analysis with PvuII. In the given conditions the mRNAs of six MT isoforms were identified in human glioblastoma cell line U87 MG. Depending on arsenic exposure conditions, an increase or decrease of MT gene expression was observed for each isoform, with the highest increase for isoforms MT1X, MT1F and MT2A mRNA (up to 13-fold) and more persistent decreases for MT1A, MT1E and MT3 mRNA. Despite the common assumption of the noninducibility of MT3, the evident MT3 mRNA increase was observed during high As exposure (up to 4-fold). In conclusion, our results clearly demonstrate the influence of As on MT isoform gene expression. The MT1X, MT1F and MT2A increase could represent brain tumour acquired resistance to As cytotoxicity while the MT3 increase is more enigmatic, with its possible involvement in arsenic-related induction of type II cell death. 相似文献
8.
Taku Fujiwara Yutaka Terao Tomonori Hoshino Shigetada Kawabata Takashi Ooshima Shizuo Sobue Shigenobu Kimura Shigeyuki Hamada 《FEMS microbiology letters》1998,161(2):331-336
Three glucosyltransferase (GTase) genes (gtfB, gtfC and gtfD) were cloned and sequenced from clinically isolated strains of Streptococcus mutans MT8148 (serotype c), MT4239 (c), MT4245 (e), MT4467 (e) and MT4251 (f), respectively. Comparison of the gtf genes revealed that interstrain difference of gtfB and gtfD was limited, while gtfC showed significant interstrain variations. Similar to gtfB and gtfD, gtfC possessed five direct repeats composed of homologous unit in the carboxyl-terminal portion. The repeating unit consisted of 63–65 amino acid residues and is responsible for glucan binding. The gtfC gene from S. mutans MT4245 lacked the fourth unit. Multiple alignment with the gtf sequence of strain GS-5 (c) revealed several changes in these gtf genes due to frameshift mutations. The peptides encoded by the gtfB, gtfC and gtfD genes of GS-5 were 1, 80, and 32 amino acid residues shorter than those of the test strains except strain MT4245. 相似文献
9.
A ferredoxin of MW 11 000 was isolated from the marine alga Rhodymenia palmata (Palmaria palmata). In its oxidised form the ferredoxin had absorption maxima at 276, sh 281, 328, 423 and 465 nm, and contained a single [2Fe-2S] cluster. The midpoint potential of the ferredoxin was ?400 mV and it effectively mediated electron transport in NADP+-photoreduction by higher plant chloroplasts, and pyruvate decarboxylation by the phosphoroclastic system of an anacrobic bacterium. The amino acid composition was Lys3, His1, Arg1, Asx12, Thr9, Ser8, Glx13, Pro4, Gly8, Ala7, Cys5, Val8, Ile4, Leu9, Tyr4, Phe2; tryptophan and methionine were absent from the molecule. The N-terminal amino acid region consisting of ca half the total amino acid sequence was determined using an automatic sequencer. 相似文献
10.
Fred S. Larimore Ilmar Kuisk Phil M. Korkowski Robert J. Roon 《Archives of biochemistry and biophysics》1980,204(1):234-240
The general amino acid transport system of Saccharomyces cerevisiae functions in the uptake of neutral, basic, and acidic amino acids (M. Grenson, C. Hou, and M. Crabeel, 1970,J. Bacteriol. 103, 770–777; J. Rytka, 1975,J. Bacteriol.121, 562–570; C. Darte and M. Grenson, 1975,Biochem. Biophys. Res. Commun.67, 1028–1033). We have previously demonstrated that this transport system can be inhibited by the amino acid, N-δ-chloroacetyl-l-ornithine (NCAO) (F. S., Larimore and R.J. Roon, 1978,Biochemistry17, 431–436). In the present study radiolabeled NCAO was synthesized and its transport and metabolism studied. Under initial rate conditions: (a) NCAO was transported by the general amino acid transport system with a Km of 52 μm, a V of 32 nmol/min/mg cells, and a pH optimum of 5.0; (b) the V for NCAO transport in gap mutants, which lack the general amino acid transport system, was approximately 1% of that observed with wild-type cells; (c) the V for NCAO in cells deprived of glucose was less than 5% of that observed when glucose was present. NCAO was transiently concentrated more than 1000-fold by yeast cells when glucose served as an energy source. The internal pool of NCAO was metabolized by the yeast cells and the products were excreted. When 100 μm [14C]NCAO was incubated with a yeast cell suspension for 8 h, more than 95% of the compound was converted into two ninhydrin-negative excretory products. The effect of NCAO on the growth of yeast cells was determined. Wild-type strains did not grow when 1 mm NCAO was present in the medium. The growth of gap mutants was not inhibited by 1 mm NCAO. 相似文献
11.
JeanClare Seagrave John L. Hanners Wayne Taylor Harold A. O'Brien 《Biological trace element research》1986,10(3):163-173
Copper uptake and distribution with time among cytoplasmic proteins were followed in cultured cells under several conditions:
(1) CHO cells, which cannot synthesize metallothioneins, were labeled with67Cu in the presence of 100 μM ZnCl2; (2) Cdr30F9 cells, which contain some constitutive metallothionein (MT), were labeled in the absence of additional ZnCl2 and; (3) Cdr30F9 cells were labeled in the presence of ZnCl2, under which conditions they synthesized additional metallothioneins. The exogenous67Cu and ZnCl2, where present, were then removed, and the distributions of67Cu among size fractions of the cellular proteins were observed at intervals for 16 h. In addition, a culture identical to
condition (3) above was also treated with 100 μM ZnCl2 during the redistribution period. The67Cu was initially resolved into three peaks by Sephadex G-75 chromatography: high molecular weight, intermediate molecular
weight, and MT. The67Cu in the MT fraction decreased with at
1/2 of 10–12 h. In contrast to this, generally, in cells with a higher initial67Cu bound to metallothionein, there was a progressive increase in the amount of67Cu eluting with the high- and intermediate-molecular-weight fractions. Since no other source of67Cu was available, these experiments suggest that copper stored in MT can be transferred to other proteins in these cells. 相似文献
12.
13.
Anne Davy Ib Svendsen Susanne O. S?rensen Mikael Blom S?rensen Jacques Rouster Morten Meldal David J. Simpson Verena Cameron-Mills 《Plant physiology》1998,117(1):255-261
The cysteine endoproteases (EP)-A and EP-B were purified from green barley (Hordeum vulgare L.) malt, and their identity was confirmed by N-terminal amino acid sequencing. EP-B cleavage sites in recombinant type-C hordein were determined by N-terminal amino acid sequencing of the cleavage products, and were used to design internally quenched, fluorogenic peptide substrates. Tetrapeptide substrates of the general formula 2-aminobenzoyl-P2-P1-P1′-P2′-tyrosine(NO2)-aspartic acid, in which cleavage occurs between P1 and P1′, showed that the cysteine EPs preferred phenylalanine, leucine, or valine at P2. Arginine was preferred to glutamine at P1, whereas proline at P2, P1, or P1′ greatly reduced substrate kinetic specificity. Enzyme cleavage of C hordein was mainly determined by the primary sequence at the cleavage site, because elongation of substrates, based on the C hordein sequence, did not make them more suitable substrates. Site-directed mutagenesis of C hordein, in which serine or proline replaced leucine, destroyed primary cleavage sites. EP-A and EP-B were both more active than papain, mostly because of their much lower Km values. 相似文献
14.
The effects of excess copper (Cu) on the accumulation of hydrogen peroxide (H2O2) and antioxidant enzyme activities in roots of the Cu accumulator Elsholtzia haichowensis Sun were investigated. Copper at 100 and 300 μM significantly increased the concentrations of malondialdehyde and H2O2, and the activities of catalase (E.C. 1.11.1.6), ascorbate peroxidase (E.C. 1.11.1.11), guaiacol peroxidase (GPOD, E.C. 1.11.1.7)
and superoxide dismutase (SOD, E.C. 1.15.1.1). Isoenzyme pattern and inhibitor studies showed that, among SOD isoforms, only
copper–zinc superoxide dismutase (CuZn–SOD) increased. Excess Cu greatly increased the accumulation of superoxide anion (O2
·−) and H2O2 in E. haichowensis roots. This study also provides the first cytochemical evidence of an accumulation of H2O2 in the root cell walls as a consequence of Cu treatments. Experiments with diphenyleneiodonium as an inhibitor of NADPH oxidase,
1,2-dihydroxybenzene-3,5-disulphonic acid as an O2
·− scavenger, and N-N-diethyldithiocarbamate as an inhibitor of SOD showed that the source of H2O2 in the cell walls could partially be NADPH oxidase. The enzyme can use cytosolic NADPH to produce O2
·−, which rapidly dismutates to H2O2 by SOD. Apoplastic GPOD and CuZn–SOD activities were induced in roots of E. haichowensis with 100 μM Cu suggesting that these two antioxidant enzymes may be responsible for H2O2 accumulation in the root apoplast. 相似文献
15.
Isolation of a Novel Peroxisomal Catalase Gene from Sugarcane,Which Is Responsive to Biotic and Abiotic Stresses 总被引:1,自引:0,他引:1
Yachun Su Jinlong Guo Hui Ling Shanshan Chen Shanshan Wang Liping Xu Andrew C. Allan Youxiong Que 《PloS one》2014,9(1)
Catalase is an iron porphyrin enzyme, which serves as an efficient scavenger of reactive oxygen species (ROS) to avoid oxidative damage. In sugarcane, the enzymatic activity of catalase in a variety (Yacheng05–179) resistant to the smut pathogen Sporisorium scitamineum was always higher than that of the susceptible variety (Liucheng03–182), suggesting that catalase activity may have a positive correlation with smut resistance in sugarcane. To understand the function of catalase at the molecular level, a cDNA sequence of ScCAT1 (GenBank Accession No. ), was isolated from sugarcane infected by S. scitamineum. ScCAT1 was predicted to encode 492 amino acid residues, and its deduced amino acid sequence shared a high degree of homology with other plant catalases. Enhanced growth of ScCAT1 in recombinant Escherichia coli Rosetta cells under the stresses of CuCl2, CdCl2 and NaCl indicated its high tolerance. Q-PCR results showed that ScCAT1 was expressed at relatively high levels in the bud, whereas expression was moderate in stem epidermis and stem pith. Different kinds of stresses, including S. scitamineum challenge, plant hormones (SA, MeJA and ABA) treatments, oxidative (H2O2) stress, heavy metal (CuCl2) and hyper-osmotic (PEG and NaCl) stresses, triggered a significant induction of ScCAT1. The ScCAT1 protein appeared to localize in plasma membrane and cytoplasm. Furthermore, histochemical assays using DAB and trypan blue staining, as well as conductivity measurement, indicated that ScCAT1 may confer the sugarcane immunity. In conclusion, the positive response of ScCAT1 to biotic and abiotic stresses suggests that ScCAT1 is involved in protection of sugarcane against reactive oxidant-related environmental stimuli. KF664183相似文献
16.
De Novo Synthesis of Amino Acids by the Ruminal Bacteria Prevotella bryantii B14, Selenomonas ruminantium HD4, and Streptococcus bovis ES1
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Cengiz Atasoglu Carmen Valds Nicola D. Walker C. James Newbold R. John Wallace 《Applied microbiology》1998,64(8):2836-2843
The influence of peptides and amino acids on ammonia assimilation and de novo synthesis of amino acids by three predominant noncellulolytic species of ruminal bacteria, Prevotella bryantii B14, Selenomonas ruminantium HD4, and Streptococcus bovis ES1, was determined by growing these bacteria in media containing 15NH4Cl and various additions of pancreatic hydrolysates of casein (peptides) or amino acids. The proportion of cell N and amino acids formed de novo decreased as the concentration of peptides increased. At high concentrations of peptides (10 and 30 g/liter), the incorporation of ammonia accounted for less than 0.16 of bacterial amino acid N and less than 0.30 of total N. At 1 g/liter, which is more similar to peptide concentrations found in the rumen, 0.68, 0.87, and 0.46 of bacterial amino acid N and 0.83, 0.89, and 0.64 of total N were derived from ammonia by P. bryantii, S. ruminantium, and S. bovis, respectively. Concentration-dependent responses were also obtained with amino acids. No individual amino acid was exhausted in any incubation medium. For cultures of P. bryantii, peptides were incorporated and stimulated growth more effectively than amino acids, while cultures of the other species showed no preference for peptides or amino acids. Apparent growth yields increased by between 8 and 57%, depending on the species, when 1 g of peptides or amino acids per liter was added to the medium. Proline synthesis was greatly decreased when peptides or amino acids were added to the medium, while glutamate and aspartate were enriched to a greater extent than other amino acids under all conditions. Thus, the proportion of bacterial protein formed de novo in noncellulolytic ruminal bacteria varies according to species and the form and identity of the amino acid and in a concentration-dependent manner. 相似文献
17.
Superoxide dismutase (SOD) is an antioxidant enzyme protecting cells from oxidative stress. Ginger (Zingiber officinale) is known for its antioxidant properties, however, there are no data on SODs from ginger rhizomes. In this study, we purified SOD from the rhizome of Z. officinale (Zo-SOD) and determined its complete amino acid sequence using N terminal sequencing, amino acid analysis, and de novo sequencing by tandem mass spectrometry. Zo-SOD consists of 151 amino acids with two signature Cu/Zn-SOD motifs and has high similarity to other plant Cu/Zn-SODs. Multiple sequence alignment showed that Cu/Zn-binding residues and cysteines forming a disulfide bond, which are highly conserved in Cu/Zn-SODs, are also present in Zo-SOD. Phylogenetic analysis revealed that plant Cu/Zn-SODs clustered into distinct chloroplastic, cytoplasmic, and intermediate groups. Among them, only chloroplastic enzymes carried amino acid substitutions in the region functionally important for enzymatic activity, suggesting that chloroplastic SODs may have a function distinct from those of SODs localized in other subcellular compartments. The nucleotide sequence of the Zo-SOD coding region was obtained by reverse-translation, and the gene was synthesized, cloned, and expressed. The recombinant Zo-SOD demonstrated pH stability in the range of 5–10, which is similar to other reported Cu/Zn-SODs, and thermal stability in the range of 10–60?°C, which is higher than that for most plant Cu/Zn-SODs but lower compared to the enzyme from a Z. officinale relative Curcuma aromatica. 相似文献
18.
19.
Xue-Ning Zhang Zhen-Hai Han Li-Li Yin Jin Kong Xue-Feng Xu Xin-Zhong Zhang Yi Wang 《Molecular biology reports》2013,40(2):1499-1504
Malus xiaojinensis is an important, iron-efficient rootstock germplasm. Iron uptake is an elaborately controlled process in plant roots, involving specialized transporters. MxIRT1, a Fe(II) transporter gene of M. xiaojinensis, is homologous to other iron transporters at the amino acid level. In the current study, the plasmid pYES2.0-MxIRT1, containing MxIRT1 cDNA, was constructed and transformed into yeast mutants. The results indicated that it could reverse the phenotype of yeast strain DEY1453, an iron uptake mutant. Complementation tests suggested that it might not be a specific transporter, as it was able to restore the phenotypes of other yeast mutant strains, including Mn, Cu and Zn uptake mutants. The functions of the critical histidine residues in the His-box of MxIRT1 were tested by transforming mutant yeast strain DEY1453 with different His residues altered by directed mutagenesis. The His-box of MxIRT1 was found to be necessary for iron transport, with different histidine residues (H1–4) playing different roles in the transport. 相似文献