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1.
Robert W. Hogg 《Journal of cellular biochemistry》1977,6(3):411-417
The active accumulation of L-arabinose by arabinose induced cultures of Escherichia coli is mediated by 2 independent transport mechanisms. One, specified by the gene locus araE, is membrane bound and possesses a relatively “low affinity.” The other, specified in part by the genetic locus araF, contains as a functional component the L-arabinose binding protein and functions with a “high affinity” for the substrate. The L-arabinose binding protein has been purified, partially characterized, crystallized, and sequenced. 相似文献
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3.
Miho Sasaki Toru Jojima Hideo Kawaguchi Masayuki Inui Hideaki Yukawa 《Applied microbiology and biotechnology》2009,85(1):105-115
Corynebacterium glutamicum strains CRA1 and CRX2 are able to grow on l-arabinose and d-xylose, respectively, as sole carbon sources. Nevertheless, they exhibit the major shortcoming that their sugar consumption
appreciably declines at lower concentrations of these substrates. To address this, the C. glutamicum ATCC31831 l-arabinose transporter gene, araE, was independently integrated into both strains. Unlike its parental strain, resultant CRA1-araE was able to aerobically
grow at low (3.6 g·l−1) l-arabinose concentrations. Interestingly, strain CRX2-araE grew 2.9-fold faster than parental CRX2 at low (3.6 g·l−1) d-xylose concentrations. The corresponding substrate consumption rates of CRA1-araE and CRX2-araE under oxygen-deprived conditions
were 2.8- and 2.7-fold, respectively, higher than those of their respective parental strains. Moreover, CRA1-araE and CRX2-araE
utilized their respective substrates simultaneously with d-glucose under both aerobic and oxygen-deprived conditions. Based on these observations, a platform strain, ACX-araE, for
C. glutamicum-based mixed sugar utilization was designed. It harbored araBAD for l-arabinose metabolism, xylAB for d-xylose metabolism, d-cellobiose permease-encoding bglF
317A
, β-glucosidase-encoding bglA and araE in its chromosomal DNA. In mineral medium containing a sugar mixture of d-glucose, d-xylose, l-arabinose, and d-cellobiose under oxygen-deprived conditions, strain ACX-araE simultaneously and completely consumed all sugars. 相似文献
4.
Khlebnikov A Skaug T Keasling JD 《Journal of industrial microbiology & biotechnology》2002,29(1):34-37
The arabinose-inducible PBAD promoter suffers from all-or-none gene expression in which cells harboring the natively controlled arabinose transport gene
(araE) are either induced or uninduced, the relative fraction of which is controlled by the concentration of arabinose. The population-averaged
variation in expression from PBAD as a function of inducer concentration is proportional to the percentage of cells that are fully induced (vs. uninduced)
rather than the level of expression in individual cells. Because of its undesirable effects on the expression of heterologous
genes, the all-or-none phenomenon was eliminated in Escherichia coli by expression of araE from arabinose-independent (either the Lactococcus lactis constitutive or IPTG-inducible lac) promoters. In these arabinose-transport engineered cells, variation in PBAD expression with arabinose concentration was a result of variation of the expression level in individual cells with all cells
in the population having approximately the same induction level. Journal of Industrial Microbiology & Biotechnology (2002) 29, 34–37 doi:10.1038/sj.jim.7000259
Received 20 November 2001/ Accepted in revised form 29 March 2002 相似文献
5.
Xiaochao Xiong Xi Wang Shulin Chen 《Journal of industrial microbiology & biotechnology》2016,43(7):1017-1025
The oleaginous bacterium, Rhodococcus jostii RHA1 has attracted considerable attention due to its capability to accumulate significant levels of triacylglycerol as renewable hydrocarbon. To enable the strain to utilize arabinose derived from lignocellulosic biomass, the metabolic pathway of L-arabinose utilization was introduced into R. jostii RHA1 by heterogenous expression of the operon, araBAD from Escherichia coli. The results showed that recombinant bearing araBAD could grow on L-arabinose as the sole carbon source, and additional expression of araFGH encoding the arabinose transporter from E. coli could improve the cell biomass yield from high contents of arabinose. We further increased the content of lipid produced from arabinose in the recombinants from 47.9 to 56.8 % of the cell dry weight (CDW) by overexpression of a gene, atf1 encoding a diglyceride acyltransferase from R. opacus PD630. This work demonstrated the feasibility of producing lipid from arabinose by genetic modification of the rhodococci strain. 相似文献
6.
Sachie Senoo Sebastian Tommi Tandar Sayaka Kitamura Yoshihiro Toya Hiroshi Shimizu 《Biotechnology and bioengineering》2019,116(12):3292-3300
An engineering tool for controlling flux distribution on metabolic pathways to an appropriate state is highly desirable in bioproduction. An optogenetic switch, which regulates gene expression by light illumination is an attractive on/off switchable system, and is a promising way for flux control with an external stimulus. We demonstrated a light-inducible flux control between glycolysis and the methylglyoxal (MGO) pathway in Escherichia coli using a CcaS/CcaR system. CcaR is phosphorylated by green light and is dephosphorylated by red light. Phosphorylated CcaR induces gene expression under the cpcG2 promoter. The tpiA gene was expressed under the cpcG2 promoter in a genomic tpiA deletion strain. The strain was then cultured with glucose minimum medium under green or red light. We found that tpiA messenger RNA level under green light was four times higher than that under red light. The repression of tpiA expression led to a decrease in glycolytic flux, resulting in slower growth under red light (0.25 hr −1) when compared to green light (0.37 hr −1). The maximum extracellular MGO concentration under red light (0.2 mM) was higher than that under green light (0.05 mM). These phenotypes confirm that the MGO pathway flux was enhanced under red light. 相似文献
7.
The inhibitory effects of 2-hydroxybiphenyl on various electron transport reactions of isolated membranes and growth in the presence of malate of either phototrophic or chemotrophic cells of Rhodospirillum rubrum were studied. 50% inhibition of both oxygen uptake of whole cells and growth under chemotrophic conditions (i.e. aerobiosis in the dark) was achieved in the presence of 0.09 mM 2-hydroxybiphenyl. With isolated membranes the same effect on NADH oxidase was obtained with 0.08 mM of inhibitor. Succinate dependent respiratory reactions were inhibited by 50% at a concentration of 0.36 mM. Growth under phototrophic conditions (i.e. anaerobiosis in the light) was inhibited by 50% in the presence of 0.17 mM (wild type strain) or 0.21 mM (blue-green mutant, strain VI) of 2-hydroxybiphenyl. Photophosphorylation and light dependent NAD+ reduction by succinate were inhibited by 50% at concentrations of 0.21 mM and 0.03 mM of inhibitor, respectively. After phototrophic growth of the organisms for about five doublings of cell mass in the presence of 0.18 mM of 2-hydroxybiphenyl coloured carotenoids could no longer be detected. Membrane fractions of such cultures exhibited normal activities of succinate cytochrome c reductase but activities of NADH cytochrome c reductase were decreased by 80%. In comparison with a blue green mutant, strain VI, of R. rubrum light induced absorbance changes at 865 nm as well as activities of photophosphorylation were unaffected. However, no activity of light dependent NAD+ reduction with succinate could be detected. The data indicate that cellular respiration as well as chemotrophic growth depend largely on NADH dependent respiration. Phototrophic growth, on the other hand, is limited by photophosphorylation while energy dependent reversed electron flow to NAD+, if at all, is of rathe minor importance.Abbreviation BChl
bacteriochlorophyll 相似文献
8.
L-arabinose isomerase (EC 5.3.1.4) mediates the isomerization of D-galactose into D-tagatose as well as the conversion of L-arabinose into L-ribulose. To investigate the properties of L-arabinose isomerase as a biocatalyst for the conversion of galactose to tagatose, the L-arabinose isomerase of Escherichia coli was characterized. The substrate specificity for L-arabinose was 166-fold higher than that for D-galactose. The optimal pH and temperature for the galactose isomerization reaction were 8.0 and 30 °C, respectively. The enzyme activity was stable for 1 h at temperatures below 35 °C and within a pH range of 8–10. The Michaelis constant, K
m, for galactose was 1480 mM, which is 25-fold higher than that for arabinose. The addition of Fe2+ and Mn2+ ions enhanced the conversion of galactose to tagatose, whereas the addition of Cu2+, Zn2+, Hg2+, and Fe3+ ions inhibited the reaction completely. In the presence of 1 mM Fe2+ ions, the K
m for galactose was found to be 300 mM. 相似文献
9.
Xiaohong Zheng Tony D'Amore Inge Russell Graham G. Stewart 《Journal of industrial microbiology & biotechnology》1994,13(3):159-166
Summary Maltotriose transport was studied in two brewer's yeast strains, an ale strain 3001 and a lager strain 3021, using laboratory-synthesized14C-maltotriose. The maltotriose transport systems preferred a lower pH (pH 4.3) to a higher pH (pH 6.6). Two maltotriose transport affinity systems have been indentified. The high affinity system hasK
m values of 1.3 mM for strain 3021 and 1.4 mM for strain 3001. The low affinity competitively inhibited by maltose and glucose withK
i values of 58 mM and 177 mM. respectively, for strain 3021, and 55 mM and 147 mM, respectively, for strain 3001. Cells grown in maltotriose and maltose had higher maltotriose and maltose transport rates, and cells grown in glucose had lower maltortriose and maltose transport rates. Early-logarithmic phase cells transported glucose faster than either maltose or maltotriose. Cells harvested later in the growth phase had increased maltotriose and maltose transport activity. Neither strain exhibited significant differences with respect to maltose and maltotriose transport activity. 相似文献
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11.
The uptake of l-phenylalanine into brush border microvilli vesicles and basolateral plasma membrane vesicles isolated from rat kidney cortex by differential centrifugation and free flow electrophoresis was investigated using filtration techniques.Brush border microvilli but not basolateral plasma membrane vesicles take up l-phenylalanine by an Na+-dependent, saturable transport system. The apparent affinity of the transport system for l-phenylalanine is 6.1 mM at 100 mM Na+ and for Na+ 13 mM at 1 mM l-phenylalanine. Reduction of the Na+ concentration reduces the apparent affinity of the transport system for l-phenylalanine but does not alter the maximum velocity.In the presence of an electrochemical potential difference for Na+ across the membrane (ηNa0 >ηNa1) the brush border microvilli accumulate transiently l-phenylalanine over the concentration in the incubation medium (overshoot phenomenon). This overshoot and the initial rate of uptake are markedly increased when the intravesicular space is rendered electrically more negative by membrane diffusion potentials induced by the use of highly permeant anions, of valinomycin in the presence of an outwardly directed K+ gradient and of carbonyl cyanide p-trifluoromethoxyphenylhydrazone in the presence of an outward-directed proton gradient.These results indicate that the entry of l-phenylalanine across the brush border membrane into the proximal tubular epithelial cells involves cotransport with Na+ and is dependent on the concentration difference of the amino acid, on the concentration difference of Na+ and on the electrical potential difference. The exit of l-phenylalanine across the basolateral plasma membranes is Na+-independent and probably involves facilitated diffusion. 相似文献
12.
Isabel G. Flórez Pedro S. Laro Amparo G. ochoa Santiago Gascón 《Biochimica et Biophysica Acta (BBA)/General Subjects》1981,674(1):71-77
A number of sugars and derivatives have been tested for their ability to induce the synthesis of α-galactosidase from Saccharomyces carlbergensis. Besides galactose and the substrates of the enzyme melibiose, raffinose and stachyose, D-galacturonic acid, L-arabinose, D-tagatose, methyl-α-D-galactoside, lactose and isopropyl-β-D-thiogalactoside were able to act as inducers. Of these, metyl-α-D-galactoside, lactose, isopropyl-β-D-thiogalactoside and L-arabinose have been shown to be gratuitous inducers with which kinetic studies of induction have been carried out. Lactose was the most efficient inducer, giving a maximal differential rate of synthesis of the enzyme of 110 mU/107 cells at a concentration of 190 mM, followed by L-arabinose (60 mU/107 cells at a concentration of 180 mM, followed by L-arabinose (60 mU/107 cells at 40 mM), isopropyl-β-D-thiogalactoside (43 mU/107 cells at 60 mM) and metyl-α-D-galactoside (25 mU/107 cells at 150 mM). The concentration of inducer required to obtain half-maximal induction was similar for lactose, L-arabinose and isopropyl-β-D-thiogalactoside and about 5-fold higher for methyl-α-D-galactoside. The property of the compounds to act as inducers was compared to their ability to interact with the enzyme and the results discussed in terms of the molecular structures which are recognized by the enzyme and by the induction machinery. 相似文献
13.
Hydrophobic protein (H protein) was isolated from membrane fractions of Bacillus subtilis and constituted into artificial membrane vesicles with lipid of B. substilis. Glutamate was accumulated into the vesicle when a Na+ gradient across the membrane was imposed. The maximum effect of Na+ on the transport was achieved at a concentration of about 40 mM, while the apparent Km for Na+ was approximately 8 mM. On the other hand, Km for glutamate in the presence of 50 mM Na+ was about 8 μM. Increasing the concentration of Na+ resulted in a decrease in Km for glutamate, maximum velocity was not affected. The transport was sensitive to monensin (Na+ ionophore).Glutamate was also accumulated when pH gradient (interior alkaline) across the membrane was imposed or a membrane potential was induced with K+-diffusion potential. The pH gradient-driven glutamate transport was sensitive to carbonylcyanide m-chlorophenylhydrazone and the apparent Km for glutamate was approximately 25 μM.These results indicate that two kinds of glutamate transport system were present in H protein: one is Na+ dependent and the other is H+ dependent. 相似文献
14.
Lars Kristian Munck Marie Louise Grndahl Jens Erik Thorbll Erik Skadhauge Bjarne Gyldenlve Munck 《Comparative biochemistry and physiology. Part A, Molecular & integrative physiology》2000,126(4)
Amino acid influx across the brush border membrane of the intact pig ileal epithelium was studied. It was examine whether in addition to system B, systems ASC and bo,+ were involved in transport of bipolar amino acids. The kinetics of interactions between lysine and leucine demonstrates that system bo,+ is present and accessible also to
-glutamine.
-aspartate (K1/2 0.3 mM) and
-glutamate (Ki 0.5 mM) share a high affinity transporter with a maximum rate of 1.3 μmol cm−2 h−1, while only
-glutamate with a K1/2 of 14.4 mM uses a low affinity transporter with a maximum rate of 2.7 μmol cm−2 h−1, system ASC, against which serine has a Ki of 1.6 mM. In the presence of 100 mM lysine,
-glutamine (A), leucine (B), and methionine (C) fulfilled the criteria of the ABC test for transport by one and the same transporter. However, serine inhibits not only transport of
-glutamate but also of glutamine (Ki 0.5 mM), and
-glutamate inhibits part of the transport of glutamine. The test does, therefore, only indicate that the three bipolar amino acids have similar affinities for transport by systems B and ASC. Further study of the function of system B must be carried out under full inhibition by lysine and glutamate. 相似文献
15.
Yanlu Zhang Liehua Wu Jingai Yu JianFeng Mei Yu Yi JianShu Chen 《Preparative biochemistry & biotechnology》2017,47(9):934-938
Human thymic stromal lymphopoietin (hTSLP) protein plays a central role in inflammation. Characterizing properties of hTSLP requires a recombinant overexpression system that produces correctly folded, active hTSLP. In this report, an efficient overexpression system for the production of hTSLP was developed. We constructed expression plasmids of the full-length hTslp gene with or without the signal peptide and transformed the plasmids into Escherichia coli. The design of the recombinant proteins included an N-terminal His-tag, which facilitated purification. An affinity gradient elution method was used to improve recovery and concentration levels of denatured hTSLP, with 90% purity observed following affinity chromatography. Refolding of the denatured hTSLP was tested using four different protein refolding approaches. The optimal refolding conditions involved stepwise buffer exchanges to reduce the urea concentration from 4 to 0?M in 50?mM Tris (pH 8.0), 1?mM EDTA, 50?mM NaCl, 10% glycerol, 400?mM L-Arg, 0.2?mM oxidized glutathione, and 2?mM reduced glutathione. The activity of the refolded recombinant hTSLP protein was measured by an ELISA assay. Interestingly, the presence of N-terminal signal peptide inhibited the overexpression of hTSLP in E. coli. The amount of recombinant hTSLP protein purified reached a level of 2.52?×?10?3?mg/L. 相似文献
16.
Tatyana V. Votyakova Elena N. Bazhenova Renata A. Zvjagilskaya 《Journal of bioenergetics and biomembranes》1993,25(5):569-574
Spermine, spermidine, and magnesium ions modulate the kinetic parameters of the Ca2+ transport system ofEndomyces magnusii mitochondria. Mg2+ at concentrations up to 5 mM partially inhibits Ca2+ transport with a half-maximal inhibiting concentration of 0.5 mM. In the presence of 2 mM MgCl2, theS
0.5 value of the Ca2+ transport system increases from 220 to 490 µM, which indicates decreased affinity for the system. Spermine and spermidine exert an activating effect, having half-maximal concentrations of 12 and 50 µM, respectively. In the case of spermine, theS
0.5 value falls to 50–65 µM, which implies an increase in the transport system affinity for Ca2+. Both Mg2+ and spermine cause a decrease of the Hill coefficient, giving evidence for a smaller degree of cooperativity. Spermine and spermidine enable yeast mitochondria to remove Ca2+ from the media completely. In contrast, Mg2+ lowers the mitochondrial buffer capacity. When both Mg2+ and spermine are present in the medium, their effects on theS
0.5 value and the free extramitochondrial Ca2+ concentration are additive. The ability of spermine and Mg2+ to regulate yeast mitochondrial Ca2+ transport is discussed. 相似文献
17.
Bhaskar Bhadra Ashis Kumar Nanda Ranadhir Chakraborty 《World journal of microbiology & biotechnology》2006,22(3):225-232
Summary A gram negative, motile, short rod-shaped, and nickel resistant (tolerating 6.5 mM Ni2+) bacterium, strain BB1A, was isolated from the waters of the River Torsa in Hashimara, Jalpaiguri district, West Bengal,
India. The isolate BB1A was identified as a strain of Acinetobacter junii following detailed analysis of morphological, physio-biochemical and 16S rRNA gene sequence. The expression of nickel resistance
in BB1A was inducible by exposure to nickel chloride at a concentration as low as 50 μM Ni2+. The other metal ions, Cu2+, Zn2+, or Pb2+ at a concentration range of 20–30 μM, also induced the nickel resistance system in this bacterium. Southern hybridizations
of BB1A genomic DNA with digoxigenin-dUTP labeled DNA probes specific for well known nickel resistance determinants, cnr, ncc or nre, resulted in no detectable signal, but nir specific probe yielded weak hybridization signal with restricted genomic DNA of BB1A. The isolate BB1A, therefore, carries
out a novel induction phenomenon of nickel resistance and presumably with a nickel resistance genetic system different from
that previously characterized in other bacteria. 相似文献
18.
19.
Jian Guo Amber L. Annett Rebecca L. Taylor Suzanne Lapi Thomas J. Ruth Maria T. Maldonado 《Journal of phycology》2010,46(6):1218-1228
We investigated copper (Cu) acquisition mechanisms and uptake kinetics of the marine diatoms Thalassiosira oceanica Hasle, an oceanic strain, and Thalassiosira pseudonana Hasle et Heimdal, a coastal strain, grown under replete and limiting iron (Fe) and Cu availabilities. The Cu‐uptake kinetics of these two diatoms followed classical Michaelis–Menten kinetics. Biphasic uptake kinetics as a function of Cu concentration were observed, suggesting the presence of both high‐ and low‐affinity Cu‐transport systems. The half‐saturation constants (Km) and the maximum Cu‐uptake rates (Vmax) of the high‐affinity Cu‐transport systems (~7–350 nM and 1.5–17 zmol · μm?2 · h?1, respectively) were significantly lower than those of the low‐affinity systems (>800 nM and 30–250 zmol · μm?2 · h?1, respectively). The two Cu‐transport systems were controlled differently by low Fe and/or Cu. The high‐affinity Cu‐transport system of both diatoms was down‐regulated under Fe limitation. Under optimal‐Fe and low‐Cu growth conditions, the Km of the high‐affinity transport system of T. oceanica was lower (7.3 nM) than that of T. pseudonana (373 nM), indicating that T. oceanica had a better ability to acquire Cu at subsaturating concentrations. When Fe was sufficient, the low‐affinity Cu‐transport system of T. oceanica saturated at 2,000 nM Cu, while that of T. pseudonana did not saturate, indicating different Cu‐transport regulation by these two diatoms. Using CuEDTA as a model organic complex, our results also suggest that diatoms might be able to access Cu bound within organic Cu complexes. 相似文献
20.
Functional Expression of Sinorhizobium meliloti BetS, a High-Affinity Betaine Transporter, in Bradyrhizobium japonicum USDA110 总被引:1,自引:0,他引:1
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Alexandre Boscari Karine Mandon Marie-Christine Poggi Daniel Le Rudulier 《Applied microbiology》2004,70(10):5916-5922
Among the Rhizobiaceae, Bradyrhizobium japonicum strain USDA110 appears to be extremely salt sensitive, and the presence of glycine betaine cannot restore its growth in medium with an increased osmolarity (E. Boncompagni, M. Østerås, M. C. Poggi, and D. Le Rudulier, Appl. Environ. Microbiol. 65:2072-2077, 1999). In order to improve the salt tolerance of B. japonicum, cells were transformed with the betS gene of Sinorhizobium meliloti. This gene encodes a major glycine betaine/proline betaine transporter from the betaine choline carnitine transporter family and is required for early osmotic adjustment. Whereas betaine transport was absent in the USDA110 strain, such transformation induced glycine betaine and proline betaine uptake in an osmotically dependent manner. Salt-treated transformed cells accumulated large amounts of glycine betaine, which was not catabolized. However, the accumulation was reversed through rapid efflux during osmotic downshock. An increased tolerance of transformant cells to a moderate NaCl concentration (80 mM) was also observed in the presence of glycine betaine or proline betaine, whereas the growth of the wild-type strain was totally abolished at 80 mM NaCl. Surprisingly, the deleterious effect due to a higher salt concentration (100 mM) could not be overcome by glycine betaine, despite a significant accumulation of this compound. Cell viability was not significantly affected in the presence of 100 mM NaCl, whereas 75% cell death occurred at 150 mM NaCl. The absence of a potential gene encoding Na+/H+ antiporters in B. japonicum could explain its very high Na+ sensitivity. 相似文献