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1.
High-speed counter-current chromatography (HSCCC)—a support free all liquid–liquid chromatography technique—has been successfully used for the preparative isolation of isorhamnetin 3-O-β-d-glucoside, isorhamnetin 3-O-β-rutinoside, quercetin 3-O-β-d-glucoside, syringetin 3-O-β-d-glucoside and protocatechuic acid from sea buckthorn juice concentrate (Hippophaë rhamnoides L. ssp. rhamnoides, Elaeagnaceae). The preparative HSCCC instrument was a multilayer coil planet centrifuge equipped with three preparative coils. Separation was performed with a two phase solvent system (n-hexane–n-butanol–water, 1:1:2 v/v/v) in ‘head-to-tail’ mode. Each injection of 4.1 g crude ethyl acetate extract yielded isorhamnetin 3-O-β-d-glucoside (95 mg), isorhamnetin 3-O-β-rutinoside (10 mg), quercetin 3-O-β-d-glucoside (5 mg), and protocatechuic acid (34 mg) with purities >98%. The flavonoid syringetin 3-O-β-d-glucoside (2 mg) was a novel compound for H. rhamnoides. Chemical structures of all compounds were determined by HPLC–ESI–MS–MS, 1D-NMR (1H, 13C, DEPT 135) spectroscopy and for elucidation of glycosidic linkages 2D-NMR (HMBC) spectroscopy was used. 相似文献
2.
Nobuko Mibu Hatsumi Aki Hirohito Ikeda Ai Saito Wataru Uchida Kazumi Yokomizo Jianrong Zhou Takeshi Miyata Kunihiro Sumoto 《Journal of Thermal Analysis and Calorimetry》2013,113(3):1015-1018
Carbohydrate recognition of some bioactive symmetrical tripodal receptor type tris(2-aminoethyl)amine (TAEA) derivatives was investigated. In calorimetric experiments, the highest binding constant (Ka) of compound C (C35H49N5O4S) with methyl α-d-mannopyranoside was Ka = 858 M?1 with 1:1 stoichiometry. Formation of hydrogen bonds in binding between symmetrical tripodal receptor type compound C and sugars was suggested by the large negative values of ?H° (=?34 to ?511 kJ mol?1). In a comparison of each set of α- and β-anomers of some monosaccharides (methyl α/β-d-galactopyranoside, methyl α/β-d-glucopyranoside, and methyl α/β-l-fucopyranoside), compound C showed that the binding constant of β-anomer was larger than that of the corresponding α-anomer, indicating higher β-anomer selectivity. The calculated energy-minimized structure of the complex of compound C with guest methyl α-d-mannopyranoside is also presented. The experimental results obtained from this work indicated that symmetrical tripodal receptor type TAEA derivative C has a lectin-like carbohydrate recognition property. 相似文献
3.
L. M. Likhosherstov O. S. Novikova N. G. Kolotyrkina I. A. Yamskov V. E. Piskarev 《Russian Chemical Bulletin》2014,63(2):507-510
Treatment of the natural tri-, tetra-, and pentasaccharides, β-d-Galp-(1→4)-[α-l-Fucp-(1→3)]-d-Glcp, α-l-Fucp-(1→2)-β-d-Galp-(1→4)-[α-l-Fucp-(1→3)]-d-Glcp, and α-l-Fucp-(1→2)-[α-d-GalNAcp-(1→3)]-β-d-Galp-(1→4)-[α-l-Fucp-(1→3)]-d-Glcp, which are glucose analogs of Lex, with ammonium carbamate in aqueous methanol gave the corresponding β-glycopyranosyl amines. After their N-acylation with N-Z-glycine N-hydroxysuccinimidyl ester (Z is benzyloxycarbonyl) with subsequent hydrogenolytic removal of Z-group, corresponding N-glycyl-β-glycopyranosyl amines were obtained in yields up to 70%. 相似文献
4.
T. V. Malyarenko A. A. Kicha N. V. Ivanchina A. I. Kalinovskii P. S. Dmitrenok S. P. Ermakova Ch. V. Minkh 《Russian Chemical Bulletin》2012,61(10):1986-1991
Three asterosaponins were isolated from the tropical starfish Asteropsis carinifera: a new one, asteropsiside A, and two known ones, regularoside A and thornasteroside A. The structure of the new compound was established using 2D NMR spectroscopy and ESI mass spectrometry as the sodium salt of 3-O-sulfonato-(20E)-6-O-{β-d-fucopyranosyl-(1→2)-β-d-galactopyranosyl-(1→4)-[β-d-quinovopyranosyl-(1→2)]-β-d-xylopyranosyl-(1→3)-β-d-quinovopyranosyl}-3β,6α-dihydroxy-5α-cholesta-9(11),20(22)-dien-23-one. Regularoside A and thornasteroside A were shown to display the ability to inhibit the growth of the T-47D and RPMI-7951 tumor cell colonies in vitro. 相似文献
5.
Mayumi Matsushita-Morita Hiroyuki Nakagawa Sawaki Tada Junichiro Marui Ryota Hattori Satoshi Suzuki Youhei Yamagata Hitoshi Amano Hiroki Ishida Michio Takeuchi Ken-Ichi Kusumoto 《Applied biochemistry and biotechnology》2013,171(1):145-164
β-Aminopeptidases exhibit both hydrolytic and aminolytic (peptide bond formation) activities and have only been reported in bacteria. We identified a gene encoding the β-aminopeptidase homolog from a genome database of the filamentous fungus Aspergillus oryzae. The gene was overexpressed in A. oryzae, and the resulting recombinant enzyme was purified. Apart from bacterial homologs [β-Ala-para-nitroanilide (pNA)], the enzyme preferred d-Leu-pNA and d-Phe-pNA as substrates. Therefore, we designated this gene as d-stereoselective aminopeptidase A (damA). The purified recombinant DamA was estimated to be a hexamer and was composed of two subunits with molecular masses of 29.5 and 11.5 kDa, respectively. Optimal hydrolytic activity of DamA toward d-Leu-pNA was observed at 50 °C and pH 8.0. The enzyme was stable up to 60 °C and from pH 4.0–11.0. DamA also exhibited aminolytic activity, producing d-Leu-d-Leu-NH2 from d-Leu-NH2 as a substrate. In the presence of 3.0 M NaCl, the amount of pNA liberated from d-Leu-pNA by DamA was 3.1-fold higher than that in the absence of NaCl. Thus, DamA is a halophilic enzyme. The enzyme was utilized to synthesize several hetero-dipeptides containing a d-amino acid at the N-terminus as well as physiologically active peptides. 相似文献
6.
Fifteen carbohydrates (d-mannose, d-glucose, d-galactose, methyl-α-d-glucose, l-rhamnose, d-xylose, d-fructose, d-arabinose, dulcitol, mannitol, β-maltose, α-lactose, melibiose, sucrose, and raffinose) and four cyclitols [l-(+)-bornesitol, myo-inositol, per-O-acetyl-1-l-(+)-bornesitol, and quinic acid] were assayed for in vitro ACE inhibition. Of these molecules, per-O-Acetyl-1-l-(+)-bornesitol, quinic acid, methyl-α-d-glucose, d-rhamnose, raffinose, and the disaccharides were determined to be either inactive or weak ACE inhibitors, whereas l-(+)-bornesitol, d-galactose, d-glucose, and myo-inositol exhibited significant ACE inhibition. Molecular docking studies were performed to investigate interactions between active compounds and human ACE (Protein Data Bank, PDB 1O83). The results of various calculations showed that all active sugars bind to the same enzyme region, which is a tunnel directed towards the active site. With the exception of myo-inositol (K i = 13.95 μM, IC50 = 449.2 μM), the active compounds presented similar K i and IC50 values. d-Galactose (K i = 19.6 μM, IC50 = 35.7 μM) and l-(+)-bornesitol (K i = 25.3 μM, IC50 = 41.4 μM) were the most active compounds, followed by d-glucose (K i = 32.9 μM, IC50 = 85.7 μM). Our docking calculations are in agreement with the experimental data and show a new binding region for sugar-like molecules, which may be explored for the development of new ACE inhibitors. 相似文献
7.
Mariateresa Maldini Paola Montoro Ireneusz Kapusta Wieslaw Oleszek Sonia Piacente Cosimo Pizza 《Chromatographia》2009,70(11-12):1621-1626
Six secondary metabolites from the methanolic extract of Sweetia panamensis (Fabaceae) bark were isolated and characterised. Along with the pyrones desmethylangonine β-d-O-glucopyranoside and desmethylangonine β-d-O-glucopyranosyl-(1→6)-O-β-d-glucopyranoside, already reported in this species, 5-O-caffeoylquinic acid (chlorogenic acid), 4-O-caffeoylquinic acid, 3-O-caffeoylquinic acid and the isoflavonoid 5-O-methylgenistein 7-O-β-d-glucopyranoside were isolated for the first time from S. panamensis. Additionally, an LC-ESI-MS qualitative analysis was performed and an ultra performance liquid chromatography (UPLC) method was developed and validated for the determination of these compounds. The UPLC method was applied to the quantitative analysis of plant samples. Pyrones and caffeoylquinic acids resulted to be the main compounds in the extract; in particular desmethylangonine β-d-O-glucopyranosyl-(1→6)-O-β-d-glucopyranoside was the most abundant compound. 相似文献
8.
Xiao-Dan Wang Kyle J. Horning Francesca M. Notarangelo Robert Schwarcz 《Analytical and bioanalytical chemistry》2013,405(30):9747-9754
d-Kynurenine (d-KYN), a metabolite of d-tryptophan, can serve as the bioprecursor of kynurenic acid (KYNA) and 3-hydroxykynurenine, two neuroactive compounds that are believed to play a role in the pathophysiology of several neurological and psychiatric diseases. In order to investigate the possible presence of d-KYN in biological tissues, we developed a novel assay based on the conversion of d-KYN to KYNA by purified d-amino acid oxidase (d-AAO). Samples were incubated with d-AAO under optimal conditions for measuring d-AAO activity (100 mM borate buffer, pH 9.0), and newly produced KYNA was detected by high-performance liquid chromatography (HPLC) with fluorimetric detection. The detection limit for d-KYN was 300 fmol, and linearity of the assay was ascertained up to 300 pmol. No assay interference was noted when other d-amino acids, including d-serine and d-aspartate, were present in the incubation mixture at 50-fold higher concentrations than d-KYN. Using this new method, d-KYN was readily detected in the brain, liver, and plasma of mice treated systemically with d-KYN (300 mg/kg). In these experiments, enantioselectivity was confirmed by determining total kynurenine levels in the same samples using a conventional HPLC assay. Availability of a sensitive, specific, and simple method for d-KYN measurement will be instrumental for evaluating whether d-KYN should be considered for a role in physiology and pathology. 相似文献
9.
β-d-Xylosidase/α-l-arabinofuranosidase from Selenomonas ruminantium is the most active enzyme known for catalyzing hydrolysis of 1,4-β-d-xylooligosaccharides to d-xylose. Catalysis and inhibitor binding by the GH43 β-xylosidase are governed by the protonation states of catalytic base (D14, pK a 5.0) and catalytic acid (E186, pK a 7.2). Biphasic inhibition by triethanolamine of E186A preparations reveals minor contamination by wild-type-like enzyme, the contaminant likely originating from translational misreading. Titration of E186A preparations with triethanolamine allows resolution of binding and kinetic parameters of the E186A mutant from those of the contaminant. The E186A mutation abolishes the pK a assigned to E186; mutant enzyme binds only the neutral aminoalcohol $ \left( {{\text{pH}} - {\text{independent}}\;K_{\text{i}}^{\text{triethanolamine}} = 19\,{\text{mM}}} \right) $ , whereas wild-type enzyme binds only the cationic aminoalcohol $ \left( {{\text{pH}} - {\text{independent}}\;K_{\text{i}}^{\text{triethanolamine}} = 0.065\,{\text{mM}}} \right) $ . At pH 7.0 and 25°C, relative kinetic parameter, $ k_{\text{cat}}^{\text{4NPX}}/k_{\text{cat}}^{\text{4NPA}} $ , for substrates 4-nitrophenyl-β-d-xylopyranoside (4NPX) and 4-nitrophenyl-α-l-arabinofuranoside (4NPA) of E186A is 100-fold that of wild-type enzyme, consistent with the view that, on the enzyme, protonation is of greater importance to the transition state of 4NPA whereas ring deformation dominates the transition state of 4NPX. 相似文献
10.
Danica Mislovičová Vladimír Pätoprstý Alica Vikartovská 《Applied biochemistry and biotechnology》2010,162(6):1669-1677
Glucose oxidase from Aspergillus niger, the specific enzyme for β-d-glucose oxidation, can also oxidize other related saccharides at very slow or negligible rates. The present study aimed to compare the kinetics of d-glucose oxidation using immobilized glucose oxidase on bead cellulose for the oxidation of related saccharides using the same biocatalyst. The significant differences were observed between the reaction rates for d-glucose and other saccharides examined. As a result, k cat/K M ratio for d-glucose was determined to be 42 times higher than d-mannose, 61.6 times higher than d-galactose, 279 times higher than d-xylose, and 254 times higher than for d-fructose and d-cellobiose. On the basis of these differences, the ability of immobilized glucose oxidase to remove d-glucose from d-cellobiose, d-glucose from d-xylose, and d-xylose from d-lyxose was examined. Immobilized catalase on Eupergit and mixed with immobilized glucose oxidase on bead cellulose or co-immobilized with glucose oxidase on bead cellulose was used for elimination of hydrogen peroxide from the reaction mixture. The accelerated elimination of d-glucose and d-xylose in the presence of co-immobilized catalase was observed. The co-immobilized glucose oxidase and catalase were able to decrease d-glucose or d-xylose content to 0–0.005% of their initial concentrations, while a minimum decrease of low oxidized saccharides d-xylose, d-cellobiose, and d-lyxose, respectively, was observed. 相似文献
11.
Toru Nagasawa Hidekazu Hosono Haruyuki Ohkishi Hideaki Yamada 《Applied biochemistry and biotechnology》1983,8(6):481-489
S-(Carboxymethyl)-d-cysteine, which is an important component of semisynthetic cephalosporin, MT-141, was enzymatically synthesized.S-(Ethoxy-carbonyl-methyl)-d-cystein was synthesized from 3-chloro-d-alanine and ethyl thioglycolate by the β-replacement reaction of 3-chloro-d-alanine chloride-lyase fromPseudomonas putida CR 1-1 and subsequently hydrolyzed by alkali. The synthesizedS-(carboxymethyl)-d-cysteine was isolated from a large scale reaction mixture and identified physicochemically. The reaction conditions for the synthesis ofS-(ethoxycarbonylmethyl)-d-cysteine were optimized using resting cells ofP. putida CR 1-1. 相似文献
12.
I. V. Kulakov A. I. Il’in Zh. A. Kabyl A. M. Gazaliev 《Russian Chemical Bulletin》2008,57(11):2443-2445
The possibility of coupling of d-glucose and d-galactose with 4-bromo-3-methylaniline, 2,4,6-tribromoaniline, and 2-amino-5-bromopyridine was studied. The substituent in the aromatic ring was found to influence the conditions and possibility of the reaction. The yields of β-d-glucopyranosyl- and β-d-galactopyranosylamines from 4-bromo-3-methylaniline and 2-amino-5-bromopyridine were 50–65%; 2,4,6-tribromoaniline did not react at all. 相似文献
13.
Annalinda Contino Giuseppe Maccarrone Maurizio Remelli 《Analytical and bioanalytical chemistry》2013,405(2-3):951-959
Stereoselective amino acid analysis has increasingly moved into the scope of interest of the scientific community. In this work, we report a study on the chiral separation of underivatized d,l-His by ligand exchange capillary electrophoresis (LECE), utilizing accurate ex ante calculations. This has been obtained by the addition to the background electrolytes (BGE) of NaClO4 which renders the separations “all in solution processes”, allowing to accurately calculate in advance the concentrations of the species present in solution and to optimize the system performances. To this aim, the formation of ternary complexes of Cu2+ ion and l-lysine (l-Lys) or l-ornithine (l-Orn) with l- and d-histidine (His), and histamine (Hm) have been studied by potentiometry and calorimetry at 25 °C and with 0.1 mol dm?3 (KNO3) in aqueous solution. The ternary species [Cu(L)(l-His)H]+ and [Cu(L)(d-His)H]+ (where L?=?l-Lys or l-Orn) show a slight but still detectable stereoselectivity, and the determination of ΔH° and ΔS° values allowed the understanding of the factors which determine this phenomenon. The stereoselectivity showed by the protonated ternary species has been exploited to chirally separate d,l-His in LECE, by using the binary complexes of copper(II) with l-Lys or l-Orn as background electrolytes added with the appropriate amounts of NaClO4. Figure
Schematic view of the separation process 相似文献
14.
For the first time an RP-LC method with diode-array detection has been developed for simultaneous analysis of three flavonoids [rhamnocitrin-3-O-β-d-galactopyranoside-4′-O-β-d-glucospyranoside (RGG), rhamnocitrin-3-O-β-d-galactopyranoside (RG), and 10-methoxymedicarpin (MC)] in a methanol extract of Oxytropis kansuensis Bunge whole plant. Separation was achieved on an ODS column within 18 min. The effect of mobile phase pH on separation of the three flavonoids was investigated. Compared with relative errors obtained by use of least-squares linear regression and logarithmic regression for data processing, weighted least-squares linear regression was more accurate. Response was a linear function of concentration in the ranges 0.0091–3.4, 0.013–4.9, and 0.0085–3.2 mg mL?1 for RGG, RG, and MC, respectively, with correlation coefficients >0.9997. The amounts of the three flavonoids in O. kansuensis Bunge were successfully analyzed with satisfactory repeatability and recovery. 相似文献
15.
Wioleta Byszewska Marianna Kańska 《Journal of Radioanalytical and Nuclear Chemistry》2014,299(3):1373-1378
We report the studies on the mechanism of oxidation of 3′,4′-dihydroxy-l-phenylalanine (l-DOPA) to neurotoxic dopachrome catalyzed by enzyme horseradish peroxidase (EC 1.11.1.7) using the kinetic (KIE), and solvent (SIE), isotope effect methods. For kinetic studies two specifically deuterated isotopomers: [2′,5′,6′-2H3]-l -DOPA was synthesized by the acid catalyzed isotopic exchange between native l-DOPA and heavy water, and [5′-2H]-l-DOPA was synthesized in two step reaction. The first step involved acid catalyzed isotopic exchange between l-tyrosine and deuterated water and resulting product [3′,5′-2H2]-l-tyrosine was hydroxylated by enzyme tyrosinase (EC 1.14.18.1). The values of deuterium KIEs and SIE’s in the enzymatic oxidation of l-DOPA and its isotopomers are determined using non-competitive spectrophotometric method. The measured values were: KIE on V max (1.1 and 2.2) and KIE on V max/K M (1.7 and 3.2) for [2′,5′,6′-2H3]-l-DOPA and [5′-2H]-l-DOPA, respectively, while the corresponding values of SIE were: SIE on V max (2.1, 2.4, and 2.1) and SIE on V max/K M (1.3. 1.6, and 1.1) for l-DOPA, [2′,5′,6′-2H3]-l-DOPA, and [5′-2H]-l-DOPA, respectively. The size of KIE and SIE, typical for secondary isotope effects indicate that both the solvent and presence of deuterium at the 2′-, 5′, and 6′-positions of l-DOPA has the little impact on the enzymatic oxidation of this compound. 相似文献
16.
Affinity chromatography of different natural biopolymers, including enzymes, is perhaps the most widely used modern technique for isolation and purification of these molecules. We have synthesized three biospecific sorbents with weak ion exchange properties by coupling ligands to carriers through hydrazide groups. These sorbents have no ion exchange or hydrophobic groups, thereby minimizing the influence of nonspecific binding on the process of affinity chromatography. These biospecific sorbents have been used for purification ofN-acetyl-β-d-hexosaminidase (EC 3.2.1.52), and for study of its active site and of its sorption and elution mechanisms in affinity chromatography. Biospecificity of sorbents was suggested by adsorption ofN-acetyl-β-d-hexosaminidase at optimum catalytic pH and by elution of the enzyme with minimal variation of pH. By comparing the known data for mapping the active site ofN-acetyl-β-d-hexosaminidase and energetic contributions of functional groups of inhibitor molecules with the obtained results, one can infer that sorption of the enzyme on biospecific sorbents is realized only by hydrogen bonds between the ligand used and ionizable groups ofN-acetyl-β-d-hexosaminidase (pK 5.5). 相似文献
17.
The reactions of a racemic four-coordinate Ni(II) complex [Ni(rac-L)](ClO4)2 with l- and d-alanine in acetonitrile/water gave two six-coordinate enantiomers formulated as [Ni(RR-L)(l-Ala)](ClO4)·2CH3CN (1) and [Ni(SS-L)(d-Ala)](ClO4) (2) (L = 5,5,7,12,12,14-hexamethyl-1,4,8,11-tetraazacyclo-tetradecane, Ala? = alanine anion), respectively. Evaporation from the remaining solutions gave two four-coordinate enantiomers characterized as [Ni(SS-L)](ClO4)2 (S-3) and [Ni(RR-L)](ClO4)2 (R-3), respectively. Single-crystal X-ray diffraction analyses of complexes 1 and 2 revealed that the Ni(II) atom has a distorted octahedral coordination geometry, being coordinated by four nitrogen atoms of L in a folded configuration, plus one carboxylate oxygen atom and one nitrogen atom of l- or d-Ala? in mutually cis-positions. Complexes 1 and 2 are supramolecular stereoisomers, constructed via hydrogen bonding between [Ni(RR-L)(l-Ala)]+ or [Ni(SS-L)(d-Ala)]+ monomers to form 1D hydrogen-bonded zigzag chains. The homochiral natures of complexes 1 and 2 have been confirmed by CD spectroscopy. 相似文献
18.
Ran Yin Xiaohui Chen Fei Han Zhenduo Shen Weiming Cheng Kaishun Bi 《Chromatographia》2008,67(11-12):961-965
A rapid and sensitive LC-MS method has been developed for the determination of luteolin-7-O-β-d-glucoside in rat plasma after solvent extraction. Separation was on an Elite Hypersil ODS2 column (250 mm × 4.6 mm i.d., 5 μm) with a mobile phase of acetonitrile-0.3% acetic acid (26:74, v/v). The samples were analyzed by using positive electrospray ionization MS in selected ion monitoring mode. The selected ions for luteolin-7-O-β-d-glucoside and the internal standard, isoquercitrin, were m/z 448.95 and m/z 464.95. Good linearity was observed over the range of 20–2,000 ng mL?1 with a lower limit of quantification of 20 ng mL?1. No interference peaks or matrix effects were observed. The validated method was applied to the pharmacokinetic study of luteolin-7-O-β-d-glucoside in rat plasma after intravenous administration of Kudiezi Injection. 相似文献
19.
N. V. Komarova I. V. Golubev S. V. Khoronenkova V. I. Tishkov 《Russian Chemical Bulletin》2012,61(7):1489-1496
d-Amino acid oxidase from the yeast Trigonopsis variabilis (TvDAAO) is widely used in fine organic synthesis, including the preparation of unnatural l-amino acids and α-keto acids. The analysis of the three-dimensional structure of TvDAAO was carried out with the aim of producing the enzyme specific to d-amino acids with bulky side chains. The analysis revealed the residue Phe54 at the entrance to the active site, which controls the substrate access to this site. The residue Phe54 was replaced by residues Ala, Ser, and Tyr. The cultivation of recombinant E. coli strains expressing TvDAAO mutants showed that the mutein with the Phe54Ala substitution had very low stability. Thus, the inactivation of the enzyme occured within 10 min after the cell disruption. The Phe54Ser TvDAAO and Phe54Tyr TvDAAO mutants were obtained as homogeneous preparations, and their thermal stability and catalytic properties were investigated. The introduction of Phe54Ser and Phe54Tyr substitutions resulted in additional stabilization of the protein macromolecule compared to the wild-type TvDAAO. Thus, the half-inactivation time for the mutant enzymes at 54 °C increased by a factor of 1.5 and 2, respectively. As in the case of wild-type TvDAAO, the thermal inactivation of the muteins proceeds via a two-step dissociative mechanism. The introduction of mutations led to a strong change in the substrate specificity profile. The mutants have no activity toward a series of d-amino acids (Phe54Ser TvDAAO toward d-Ala, d-Ser, d-Val, and d-Thr; Phe54Tyr TvDAAO toward d-Ser, d-Tyr, d-Thr, and d-Lys). The catalytic efficiency (the k cat/K M ratio) of the Phe54Ser TvDAAO mutant toward d-amino acids with bulky side chains (d-Lys, d-Asn, d-Phe, d-Tyr, d-Trp, and d-Leu) increased from 2.4 to 7.3 times. 相似文献
20.
Xiao Xia Liu Lan Wang Yu Jie Wang Li Ling Cai 《Applied biochemistry and biotechnology》2010,160(3):822-830
In this study, we introduced a new strategy, feeding d-glucose, to overproduce extracellular 5-aminolevulinic acid (ALA) in the recombinant Escherichia coli. We investigated that the d-glucose concentration is dependent on extracellular ALA production. The results indicated that increasing d-glucose concentration in bacteria culture enhanced final cell density and ALA yield and simultaneously decreased the activities of ALA synthase (ALAS) and ALA dehydratase (ALAD); then, the inhibitory effect of d-glucose on ALAS activity was relieved with the metabolism of d-glucose. when 4.0 g/L d-glucose was added at late exponential phase; 1.46 g/L ALA was achieved in shaking culture, which is 47% or 109% higher than the ALA yields with 30 mM levulinic acid of ALAD inhibitor or no inhibitor. In jar fermenter, final extracellular ALA concentration reached 3.1 g/L by feeding with d-glucose. 相似文献