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1.
Oshima T 《Amino acids》2007,33(2):367-372
Summary. Recent research progress on polyamines in extreme thermophiles is reviewed. Extreme thermophiles produce two types of unique
polyamines; one is longer polyamines such as caldopentamine and caldohexamine, and the other is branched polyamines such as
tetrakis(3-aminopropyl)ammonium. The protein synthesis catalyzed by a cell-free extract of Thermus thermophilus, an extreme thermophile, required the presence of a polyamine and the highest activity was found in the presence of tetrakis(3-aminopropyl)ammonium.
In vitro experiments, longer polyamines efficiently stabilized double stranded nucleic acids and a branched polyamine, tetrakis(3-aminropyl)ammonium,
stabilized stem-and-loop structures. In T. thermophilus, polyamines are synthesized from arginine by a new metabolic pathway; arginine is converted to agmatine and then agmatine
is aminopropylated to N1-aminopropylagmatine which is converted to spermidine by an enzyme coded by a gene homologous to speB (a gene for agmatinase). In this new pathway spermidine is not synthesized from putrescine. Reverse genetic studies indicated
that the unique polyamines are synthesized from spermidine. 相似文献
2.
The conserved two-domain ribosomal protein (r-protein) L1 is a structural part of the L1 stalk of the large ribosomal subunit and regulates the translation of the operon that comprises its own gene. The regulatory properties of the bacterial r-protein L1 have only been studied in detail for Escherichia coli; however, there were no such studies for other bacteria, in particular, Thermus thermophilus and Thermotoga maritima, which are more evolutionarily ancient. It is known that domain I of the r-protein L1 might have regulatory properties of the whole protein. The aim of this study was to identify regulatory sites on the mRNA of T. thermophilus and T. maritima that interact with r-proteins L1, as well as with their domains I from the same organisms. An analysis of the mRNA of the L11 operon T. thermophilus showed the presence of one potential binding site of the L1 r-protein, two such regions were found also in the mRNA sequence of the L11 operon of T. maritima. The dissociation constants for the L1 proteins from T. thermophilus and T. maritima and their domains I with mRNA fragments from the same organisms that contain the supposed L1-binding sites were determined by surface plasmon resonance. It has been shown that the ribosomal proteins L1 as their domains I bind specific fragments of mRNA from the same organisms that may suggest regulatory activity of the L1 protein in the T. thermophilus and T. maritima and conservatism of the principles of L1-RNA interactions. 相似文献
3.
Kottakis F Lamari F Matragkou Ch Zachariadis G Karamanos N Choli-Papadopoulou T 《Amino acids》2008,34(3):413-420
Summary. Arabino-Galactan Proteins (AGPs) were isolated from Chios mastic gum (CMG) by using a buffer containing 0.1 M NaCl, 20 mM
Tris–HCl, pH 7.5. Protein analytical methods, combined with specific procedures for carbohydrate characterization, indicated
the presence of highly glycosylated protein backbone. In particular, staining by Yariv reagent of the electrophoretically
separated molecules revealed the existence of arabinose and galactose and such a modification is characteristic for AGPs.
After experiments involving extensive dialysis of the isolated extracts against water and atomic absorption, there was evidence
of the existence of zinc ions that are probably covalently bound to the AGPs. By using anion-exchange chromatography, capillary
electrophoresis, colorimetric methods and GC-MS, it was found that the extracts were separated into three major populations
(A, B, and C), which were consistent with their respective negative charge content namely, uronic acid. The characterization
of neutral sugars that was investigated with GC-MS showed the existence of arabinose and galactose in different amounts for
each group.
Experiments concerning the inhibition of growth of Helicobacter pylori in the presence of AGPs, as is shown for other CMG constituents, showed that the extracts of at least 1.4 g CMG affected
the viability of the bacterium. There is no evidence as to whether the AGPs provoke abnormal morphologies of H. pylori, as is reported for the total CMG, or for O-glycans that possess terminal α1, 4-linked N-acetylglucosamine and are expressed
in the human gastric mucosa; this has to be further investigated.
Authors’ address: Theodora Choli-Papadopoulou, Laboratory of Biochemistry, School of Chemistry, Aristotle University of Thessaloniki,
TK 54124 Thessaloniki, Greece 相似文献
4.
M. Helanto K. Kiviharju T. Granström M. Leisola A. Nyyssölä 《Applied microbiology and biotechnology》2009,83(1):77-83
l-Ribose is a rare and expensive sugar that can be used as a precursor for the production of l-nucleoside analogues, which are used as antiviral drugs. In this work, we describe a novel way of producing l-ribose from the readily available raw material l-arabinose. This was achieved by introducing l-ribose isomerase activity into l-ribulokinase-deficient Escherichia coli UP1110 and Lactobacillus plantarum BPT197 strains. The process for l-ribose production by resting cells was investigated. The initial l-ribose production rates at 39°C and pH 8 were 0.46 ± 0.01 g g−1 h−1 (1.84 ± 0.03 g l−1 h−1) and 0.27 ± 0.01 g g−1 h−1 (1.91 ± 0.1 g l−1 h−1) for E. coli and for L. plantarum, respectively. Conversions were around 20% at their highest in the experiments. Also partially purified protein precipitates
having both l-arabinose isomerase and l-ribose isomerase activity were successfully used for converting l-arabinose to l-ribose. 相似文献
5.
V. N. Verbenko L. V. Kuznetsova E. P. Krupyan V. I. Shalguev 《Russian Journal of Genetics》2009,45(10):1192-1199
Plasmids pKS5 and pKSrec30 carrying normal and mutant alleles of the Deinococcus recA gene controlled by the lactose promoter slightly increase radioresistance of Escherichia coli cells with mutations in genes recA and ssb. The RecA protein of D. radiodurans is expressed in E. coli cells, and its synthesis can be supplementary induced. The radioprotective effect of the xenologic protein does not exceed
1.5 fold and yields essentially to the contribution of plasmid pUC19-recA1.1 harboring the E. coli recA
+ gene in the recovery of resistance of the ΔrecA deletion mutant. These data suggest that the expression of D. radiodurans recA gene in E. coli cells does not complement mutations at gene recA in the chromosome possibly due to structural and functional peculiarities of the D. radiodurans RecA protein. 相似文献
6.
Ozaki S Imai H Iwakiri T Sato T Shimoda K Nakayama T Hamada H 《Biotechnology letters》2012,34(3):475-481
A glucosyltransferase (GT) of Phytolacca americana (PaGT3) was expressed in Escherichia coli and purified for the synthesis of two O-β-glucoside products of trans-resveratrol. The reaction was moderately regioselective with a ratio of 4′-O-β-glucoside: 3-O-β-glucoside at 10:3. We used not only the purified enzyme but also the E. coli cells containing the PaGT3 gene for the synthesis of glycoconjugates. E. coli cell cultures also have other advantages, such as a shorter incubation time compared with cultured plant cells, no need for
the addition of exogenous glucosyl donor compounds such as UDP-glucose, and almost complete conversion of the aglycone to
the glucoside products. Furthermore, a homology model of PaGT3 and mutagenesis studies suggested that His-20 would be a catalytically
important residue. 相似文献
7.
Thermostable Mn-dependent catalases are promising enzymes in biotechnological applications. In the present study, a Mn-containing
superoxide dismutase of the hyperthermophilic Thermus thermophilus HB27 had been purified and characterized by a two-stage ultrafiltration process after being expressed in E. coli. The enzyme was highly stable at 90°C and retained 57% activity after heat treatment at 100°C for 1 h. The native form of
the enzyme was determined as a homotetramer by analytical size exclusion chromatography and sodium dodecyl sulfate–polyacrylamide
gel electrophoresis. The final purified enzyme had an isoelectric point of 6.2 and a high α-helical content of 70%, consistent
with the theoretical values. This showed that the purified SOD folded with a reasonable secondary structure. 相似文献
8.
Su-Lim Choi Sang Jun Lee Jae-Seok Ha Jae Jun Song Young Ha Rhee Seung-Goo Lee 《Biotechnology and Bioprocess Engineering》2011,16(6):1173-1179
The cellulose-binding domain (CBD) of a Cellulomonas fimi exo-glucanase was translationally fused with β-glucuronidase (GusA) from Escherichia coli and β-glycosidase (BglA) from Thermus caldophilus, respectively. Two fusion proteins (GusA-CBD and BglA-CBD) were expressed as insoluble aggregates in cells and isolated by
centrifugation of the cell lysates. Interestingly, activity assays revealed that > 90% of the catalytic activity of both proteins
was localized in the insoluble fractions. For example, the GusA-CBD particles exhibited 21 units per mg protein, which corresponded
to 19% specific activity of the highly purified soluble GusA. The specific activity increased further up to 42 units per mg
protein when treated with either sonication or chaotropic L-arginine. These results demonstrate that fusion with CBD family
II may activate catalytic protein particles in E. coli cells, and that internal proteins of the particles are also active. Finally, the protein particles were tested in repeated
batch operations after being cross-linked with chemicals, indicating that they have potential as a new preparation for immobilized
biocatalysts. 相似文献
9.
The genome of Lactococcus lactis encodes a single long chain 3-ketoacyl-acyl carrier protein synthase. This is in contrast to its close relative, Enterococcus faecalis, and to Escherichia coli, both of which have two such enzymes. In E. faecalis and E. coli, one of the two long chain synthases (FabO and FabB, respectively) has a role in unsaturated fatty acid synthesis that cannot
be satisfied by FabF, the other long chain synthase. Since L. lactis has only a single long chain 3-ketoacyl-acyl carrier protein synthase (annotated as FabF), it seemed likely that this enzyme
must function both in unsaturated fatty acid synthesis and in elongation of short chain acyl carrier protein substrates to
the C18 fatty acids found in the cellular phospholipids. We report that this is the case. Expression of L. lactis FabF can functionally replace both FabB and FabF in E. coli, although it does not restore thermal regulation of phospholipid fatty acid composition to E. coli
fabF mutant strains. The lack of thermal regulation was predictable because wild-type L. lactis was found not to show any significant change in fatty acid composition with growth temperature. We also report that overproduction
of L. lactis FabF allows growth of an L. lactis mutant strain that lacks the FabH short chain 3-ketoacyl-acyl carrier protein synthase. The strain tested was a derivative
(called the ∆fabH bypass strain) of the original fabH deletion strain that had acquired the ability to grow when supplemented with octanoate. Upon introduction of a FabF overexpression
plasmid into this strain, growth proceeded normally in the absence of fatty acid supplementation. Moreover, this strain had
a normal rate of fatty acid synthesis and a normal fatty acid composition. Both the ∆fabH bypass strain that overproduced FabF and the wild type strain incorporated much less exogenous octanoate into long chain
phospholipid fatty acids than did the ∆fabH bypass strain. Incorporation of octanoate and decanoate labeled with deuterium showed that these acids were incorporated
intact as the distal methyl and methylene groups of the long chain fatty acids. 相似文献
10.
Summary. A novel practical method for the synthesis of N-methyl-DL-aspartic acid 1 (NMA) and new syntheses for N-methyl-aspartic acid derivatives are described. NMA 1, the natural amino acid was synthesized by Michael addition of methylamine to dimethyl fumarate 5. Fumaric or maleic acid mono-ester and -amide were regioselectively transformed into beta-substituted aspartic acid derivatives.
In the cases of maleamic 11a or fumaramic esters 11b, the α-amide derivative 13 was formed, but hydrolysis of the product provided N-methyl-DL-asparagine 9 via base catalyzed ring closure to DL-α-methylamino-succinimide 4, followed by selective ring opening. Efficient methods were developed for the preparation of NMA-α-amide 13 from unprotected NMA via sulphinamide anhydride 15 and aspartic anhydride 3 intermediate products. NMA diamide 16 was prepared from NMA dimethyl ester 6 and methylamino-succinimide 4 by ammonolysis. Temperature-dependent side reactions of methylamino-succinimide 4 led to diazocinone 18, resulted from self-condensation of methylamino-succinimide via nucleophyl ring opening and the subsequent ring-transformation. 相似文献
11.
Smirnova IA Zaslavsky D Fee JA Gennis RB Brzezinski P 《Journal of bioenergetics and biomembranes》2008,40(4):281-287
The ba
3-type cytochrome c oxidase from Thermus thermophilus is phylogenetically very distant from the aa
3–type cytochrome c oxidases. Nevertheless, both types of oxidases have the same number of redox-active metal sites and the reduction of O2 to water is catalysed at a haem a
3-CuB catalytic site. The three-dimensional structure of the ba
3 oxidase reveals three possible proton-conducting pathways showing very low homology compared to those of the mitochondrial,
Rhodobacter sphaeroides and Paracoccus denitrificans aa
3 oxidases. In this study we investigated the oxidative part of the catalytic cycle of the ba
3
-cytochrome c oxidase using the flow-flash method. After flash-induced dissociation of CO from the fully reduced enzyme in the presence
of oxygen we observed rapid oxidation of cytochrome b (k ≅ 6.8 × 104 s−1) and formation of the peroxy (PR) intermediate. In the next step a proton was taken up from solution with a rate constant of ~1.7 × 104 s−1, associated with formation of the ferryl (F) intermediate, simultaneous with transient reduction of haem b. Finally, the enzyme was oxidized with a rate constant of ~1,100 s−1, accompanied by additional proton uptake. The total proton uptake stoichiometry in the oxidative part of the catalytic cycle
was ~1.5 protons per enzyme molecule. The results support the earlier proposal that the PR and F intermediate spectra are similar (Siletsky et al. Biochim Biophys Acta 1767:138, 2007) and show that even though the architecture of the proton-conducting pathways is different in the ba
3 oxidases, the proton-uptake reactions occur over the same time scales as in the aa
3-type oxidases.
Smirnova and Zaslavsky contributed equally to the work described in this paper. 相似文献
12.
A convenient synthetic strategy of the common acidic pentasaccharide repeating unit corresponding to the O-antigen of enterotoxigenic E. coli O168 and Shigella dysenteriae type 4 has been successfully developed. A stereoselective [2 + 3] block glycosylation method has been exploited to get the
target pentasaccharide derivative. Most of the synthetic intermediates were solid and prepared in high yields from commercially
available reducing sugars following a series of protection-deprotection reactions. A α-D-mannose moiety has been used as the
source of α-D-glucosamine moiety. A late-stage TEMPO mediated selective oxidation reaction finally resulted in the pentasaccharide
containing a glucuronic acid unit. 相似文献
13.
Summary. To mutant ddY/DAO− mice lacking D-amino-acid oxidase activity and normal ddY/DAO+ mice, five D-amino acids (D-Asp, D-Ser, D-Ala, D-Leu and D-Pro) were orally administered for two weeks, and the D-amino acid levels were examined in seven brain regions. The levels of D-Asp markedly increased in the pituitary and pineal glands in both strains. In the ddY/DAO+ mice, the levels of the other D-amino acids did not significantly change in most of the brain regions. While in the ddY/DAO− mice the levels of D-Ser significantly increased in most of the brain regions except for the cerebrum and hippocampus. The levels of D-Ala and D-Leu increased in all regions but the levels of D-Pro did not significantly change. The same five D-amino acids were intravenously injected into Wistar rats and the D-amino acid levels in their brains were examined for 60 min after the administration. The levels of D-Asp markedly increased in the pineal gland 3 min after the administration, while the levels of D-Ser, D-Ala, and D-Pro increased both in the pineal and pituitary glands, the levels of D-Leu increased in all brain regions. These results are useful for the elucidation of the origins and regulation of D-amino acids in the mammalian body. 相似文献
14.
A quantitative fluorescent in situ hybridization method was employed to evaluate the competitive inhibitory effect of three Lactobacillus strains (Lactobacillus reuteri, Lactobacillus gasseri, and Lactobacillus plantarum) against Escherichia coli internalization in a model system of HT 29 cells. Furthermore, aggregation and adhesion abilities of the Lactobacillus strains were examined. All lactobacilli were able to attach to the HT 29 cells and aggregate with pathogens; however, the adhesion and aggregation degree was strain-dependent. L. reuteri possessed a high capacity of adhesion (6.80 ± 0.63; log CFU ± SEM per well), whereas lower capacities were expressed by L. gasseri (4.52 ± 0.55) and L. plantarum (4.90 ± 0.98). Additionally, L. reuteri showed the rapid or normal ability to aggregate with selected E. coli in comparison with remaining two lactobacilli, which showed only slow or negative aggregative reaction. Internalization of E. coli into the cell lines was markedly suppressed by L. reuteri, while L. gasseri and L. plantarum caused only a minimum anti-invasion effect. The fact that L. reuteri in our experiments showed an outstanding potential for adhering to the colon epithelial cell line, compared with the rest strains, suggested that one of the possible mechanisms of preventing pathogen adhesion and invasion is simple competitions at certain receptors and capability to block receptor binding sites, or that an avid interaction between L. reuteri and the host cell might be modulating intracellular events responsible for the E. coli internalization. Moreover, L. reuteri exhibited a strong ability to aggregate with E. coli, which could be another limiting factor of pathogen invasion. 相似文献
15.
Hayakawa T Howlader MT Yamagiwa M Sakai H 《Applied microbiology and biotechnology》2008,80(6):1033-1037
Cry4Aa produced by Bacillus thuringiensis is a dipteran-specific toxin and is, therefore, of great interest for developing a bioinsecticide to control mosquitoes.
However, the expression of Cry4Aa in Escherichia coli is relatively low, which is a major disadvantage in its development as a bioinsecticide. In this study, to establish an effective
production system, a 1,914-bp modified gene (cry4Aa-S1) encoding Cry4Aa was designed and synthesized in accordance with the G + C content and codon preference of E. coli genes without altering the encoded amino acid sequence. The cry4Aa-S1 gene allowed a significant improvement in expression level, over five-fold, compared to that of the original cry4Aa gene. The product of the cry4Aa-S1 gene showed the same level of insecticidal activity against Culex pipiens larvae as that from cry4Aa. This suggested that unfavorable codon usage was one of the reasons for poor expression of cry4Aa in E. coli, and, therefore, changing the cry4Aa codons to accord with the codon usage in E. coli led to efficient production of Cry4Aa. Efficient production of Cry4Aa in E. coli can be a powerful measure to prepare a sufficient amount of Cry4Aa protein for both basic analytical and applied researches. 相似文献
16.
Hai-Nan Su Bin-Bin Xie Xiu-Lan Chen Jin-Xia Wang Xi-Ying Zhang Bai-Cheng Zhou Yu-Zhong Zhang 《Journal of applied phycology》2010,22(1):65-70
Allophycocyanin (APC) is a minor component of phycobiliproteins in cyanobacteria and red algae. This paper describes a simple
and inexpensive extracting method for isolating APC from Spirulina (Arthrospira) platensis with high efficiency. The crude phycobiliprotein extract was pretreated by ammonium sulfate fractionation. Then, by adding
hydroxylapatite into crude phycobiliprotein extract dissolved in 20 mM phosphate buffer (pH 7.0), APC was selectively adsorbed
by hydroxylapatite but C-phycocyanin (C-PC) was not. The hydroxylapatite was collected and APC was extracted from the crude
phycobiliprotein extract. Then, the enriched APC was washed off from the hydroxylapatite using 100 mM phosphate buffer (pH 7.0).
In this simple extracting method it was easy to remove C-PC and isolate APC in large amounts. The absorbance ratio A
650/A
280 of extracted APC reached 2.0. The recovery yield was 70%, representing 4.61 mg · g−1 wet weight. The extracted APC could be further purified by a simple anion-exchange chromatography with a pH gradient from
5.6 to 4.0. The absorbance ratio A
650/A
280 of the purified APC reached 5.0, and the overall recovery yield was 43%, representing 2.83 mg · g−1 wet weight. Its purity was confirmed by native polyacrylamide gel electrophoresis (PAGE) and sodium dodecyl sulfate-PAGE. 相似文献
17.
Cupriavidus metallidurans CH34 genome contains an ortholog of Atm1p named AtmA (Rmet_0391, YP_582546). In Saccharomyces cerevisiae, the ABC-type transport system Atm1p is involved in export of iron–sulfur clusters from mitochondria into the cytoplasm for
assembly of cytoplasmic iron–sulfur containing proteins. An ∆atmA mutant of C. metallidurans was sensitive to nickel and cobalt but not iron cations. AtmA increased also resistance to these cations in Escherichia coli strains that carry deletions of the genes for other nickel and cobalt transport systems. In C. metallidurans, atmA expression was not significantly induced by nickel and cobalt, but repressed by zinc. AtmA was purified as a 70 kDa protein
after expression in E. coli. ATPase activity of AtmA was stimulated by nickel and cobalt. 相似文献
18.
Takaomi Arai Aya Kotake P. Mark Lokman Katsumi Tsukamoto 《Ichthyological Research》2003,50(2):190-194
The migratory history of Anguilla dieffenbachii and A. australis, collected from a coastal lake of New Zealand, was examined using analysis of strontium (Sr) and calcium (Ca) concentrations.
Line analysis of Sr : Ca ratios along the life history transect of each otolith showed a peak (Ca. 16–20 × 10−3) between the core and elver mark, which corresponded to the period of their leptocephalus and early glass eel stages in the
ocean. The mean Sr : Ca ratios from the elver mark to the otolith edge indicated that eels had different migratory histories,
which included freshwater residency in some eels (average Sr : Ca ratios, 1.7 × 10−3–2.4 × 10−3) but not in others (average Sr : Ca ratios, 3.1 × 10−3–6.5 × 10−3). These findings suggest that New Zealand freshwater eels have a flexible migration strategy and an ability to adapt to various
habitats and salinities.
Received: November 25, 2002 / Revised: January 17, 2003 / Accepted: January 17, 2003 相似文献
19.
Summary. Ornithine decarboxylase (ODC) has a very fast turnover in mammalian cells, but is a stable enzyme in T. brucei and other trypanosmatid parasites like Leishmania donovani. However, Crithidia fasciculata, which is a phylogenetically closely related trypanosomatid to L. donovani, has an ODC with a rapid turnover. Interestingly, C. fasciculata ODC, but not L. donovani ODC, is rapidly degraded also in mammalian systems. In order to obtain information on what sequences are important for the
rapid degradation of C. fasciculata ODC, we produced a variety of C. fasciculata/L. donovani ODC hybrid proteins and characterized their turnover using two different mammalian expression systems. The results obtained
indicate that C. fasciculata ODC contains several sequence elements essential for the rapid turnover of the protein and that these regions are mainly
located in the central part of the enzyme.
Present address: Department of Microbiology, Moyne Institute of Preventive Medicine, University of Dublin – Trinity College,
Dublin, Ireland
Authors’ address: Lo Persson, Department of Experimental Medical Science, Lund University, BMC D-12, S-221 84 Lund, Sweden 相似文献
20.
Daisuke Sugimori 《Applied microbiology and biotechnology》2009,82(2):351-357
To develop a microbial treatment of edible oil-contaminated wastewater, microorganisms capable of rapidly degrading edible
oil were screened. The screening study yielded a yeast coculture comprising Rhodotorula pacifica strain ST3411 and Cryptococcus laurentii strain ST3412. The coculture was able to degrade efficiently even at low contents of nitrogen ([NH4–N] = 240 mg/L) and phosphorus sources ([PO4–P] = 90 mg/L). The 24-h degradation rate of 3,000 ppm mixed oils (salad oil/lard/beef tallow, 1:1 w/w) at 20°C was 39.8% ± 9.9% (means ± standard deviations of eight replicates). The highest degradation rate was observed at
20°C and pH 8. In a scaled-up experiment, the salad oil was rapidly degraded by the coculture from 671 ± 52.0 to 143 ± 96.7 ppm
in 24 h, and the degradation rate was 79.4% ± 13.8% (means ± standard deviations of three replicates). In addition, a repetitive
degradation was observed with the cell growth by only pH adjustment without addition of the cells. 相似文献