RESUMO
The removal of isoleucine from valine has been a key issue in the stage of valine crystallization, which is the final step in the valine production process in industry. To address this issue, a three-zone simulated moving-bed (SMB) process for the separation of valine and isoleucine has been developed previously. However, the previous process, which was based on a classical port-location mode, had some limitations in throughput and valine product concentration. In this study, a three-zone SMB process based on a modified port-location mode was applied to the separation of valine and isoleucine for the purpose of making a marked improvement in throughput and valine product concentration. Computer simulations and a lab-scale process experiment showed that the modified three-zone SMB for valine separation led to >65% higher throughput and >160% higher valine concentration compared to the previous three-zone SMB for the same separation.
Assuntos
Cromatografia Líquida/métodos , Isoleucina/isolamento & purificação , Valina/isolamento & purificação , Adsorção , Cromatografia Líquida/instrumentação , Simulação por ComputadorRESUMO
The cattle tick Rhipicephalus (Boophilus) microplus lays eggs in the soil near the roots of grass, or in similar highly moist environments that are prone to biofilm formation. Tick eggs have a protective wax coating that may be a source of nutrients for microorganisms. However, as the eggs remain viable and show no visible signs of microbial colonization, we hypothesized that the coating might have anti-biofilm properties. We show here that the coating inhibits biofilm formation by both Gram-negative and Gram-positive bacteria, though by different mechanisms. We have identified the anti-biofilm molecule as N-(3-sulfooxy-25-cholest-5-en-26-oyl)-L-isoleucine (boophiline), and we show that it inhibits the expression of fliC (flagellin) and cdrA (biofilm scaffold), whose products are necessary for biofilm formation in Pseudomonas aeruginosa. Boophiline is a novel biofilm inhibitor being also effective against Staphylococcus epidermidis biofilm. In our study we show evidences of the boophiline mode of action in the protection of arthropod eggs against biofilm colonization.
Assuntos
Bactérias/efeitos dos fármacos , Biofilmes/efeitos dos fármacos , Colestenos/farmacologia , Isoleucina/análogos & derivados , Rhipicephalus/química , Rhipicephalus/microbiologia , Animais , Antibacterianos/isolamento & purificação , Antibacterianos/farmacologia , Colestenos/isolamento & purificação , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Isoleucina/isolamento & purificação , Isoleucina/farmacologia , Óvulo/química , Óvulo/microbiologia , Pseudomonas aeruginosa/efeitos dos fármacosRESUMO
An X-ray crystal structural analysis revealed that (2S,3S)-N-acetyl-2-amino-3-methylpentanoic acid (N-acetyl-L-isoleucine; Ac-L-Ile) and (2R,3S)-N-acetyl-2-amino-3-methylpentanoic acid (N-acetyl-D-alloisoleucine; Ac-D-aIle) formed a molecular compound containing one Ac-L-Ile molecule and one Ac-D-aIle molecule as an unsymmetrical unit. This molecular compound is packed with strong hydrogen bonds forming homogeneous chains consisting of Ac-L-Ile molecules or Ac-D-aIle molecules and weak hydrogen bonds connecting these homogeneous chains in a fashion similar to that observed for Ac-L-Ile and Ac-D-aIle. Recrystallization of an approximately 1:1 mixture of Ac-L-Ile and Ac-D-aIle from water gave an equimolar molecular compound due to its lower solubility than that of Ac-D-aIle or especially Ac-L-Ile. The results suggest that the equimolar mixture of Ac-L-Ile and Ac-D-aIle could be obtained from an Ac-L-Ile-excess mixture by recystallization from water.
Assuntos
Isoleucina/análogos & derivados , Ácidos Pentanoicos/química , Cristalografia por Raios X , Ligação de Hidrogênio , Isoleucina/química , Isoleucina/isolamento & purificação , Modelos Moleculares , Conformação Molecular , Ácidos Pentanoicos/isolamento & purificação , Compostos de Amônio Quaternário/química , Sais/química , Solubilidade , EstereoisomerismoRESUMO
BACKGROUND: The novel natural product cinnacidin was isolated from a fungal fermentation extract of Nectria sp. DA060097. The compound was found to contain a cyclopentalenone ring system with an isoleucine subunit linked through an amide bond. Initial biological characterization of cinnacidin suggested promising herbicidal activity. RESULTS: Two synthetic analogs, (2S,3S)-2-[(3RS,3aSR,6aRS)-3-methoxy-4-oxo-3,3a,4,5,6,6a-hexahydropentalen-1-ylcarbamoyl]-3-methylvaleric acid and benzyl (2S,3S)-2-[(3RS,3aSR,6aRS)-3-methoxy-4-oxo-3,3a,4, 5,6,6a-hexahydropentalen-1-ylcarbamoyl]-3-methylvalerate, were prepared for further characterization, and their herbicidal activities were compared with that of cinnacidin. CONCLUSIONS: The synthetic compounds were highly phytotoxic on a range of weeds. Based on the symptoms in treated plants, the mode of action of these compounds is suggested to be similar to that of coronatine and jasmonic acid. Coronatine was more active against warm-season grasses, while the cinnacidin benzyl ester analog was more effective on cool-season grasses. In a seedling growth bioassay conducted on bentgrass, the cinnacidin analog was equivalent in activity to coronatine.
Assuntos
Herbicidas/química , Herbicidas/farmacologia , Hypocreales/química , Isoleucina/análogos & derivados , Toxinas Biológicas/química , Toxinas Biológicas/farmacologia , Agrostis/efeitos dos fármacos , Aminoácidos/farmacologia , Arabidopsis/efeitos dos fármacos , Herbicidas/síntese química , Herbicidas/isolamento & purificação , Hypocreales/genética , Hypocreales/isolamento & purificação , Hypocreales/metabolismo , Indenos/farmacologia , Isoleucina/síntese química , Isoleucina/química , Isoleucina/isolamento & purificação , Isoleucina/farmacologia , Toxinas Biológicas/síntese química , Toxinas Biológicas/isolamento & purificaçãoRESUMO
The enantiomeric separation of unmodified D,L-isoleucine was achieved in citric acid-zinc(â ¡) medium by capillary electrophoresis (CE) with contactless conductivity detector (C4D). In the conventional chiral separation methods of amino acid, a chiral complex used as the chiral selector was added into the eluent in order to yield a chiral environment. However, in this study a non-chiral solution, i. e. 2.8 mmol/L NaOH+0.8 mmol/L citric acid+2.0 mmol/L zinc acetate was used as the running buffer, and the citric acid-zinc(â ¡) acted the role of a chiral selector. Under the optimum experimental conditions:uncoated fused-silica capillary (45 cm×50 µm, Leff=40 cm), separation voltage of +13 kV, electrokinetic injection of 11 kV×8 s, the enantiomers of D,L-isoleucine were baseline separated within 8 min with the resolutions (Rs) of 2.0. The calibration curve of each enantiomer showed good linearity in the range from 1.0 mg/L to 20 mg/L, with the limits of detection of 0.40 mg/L. The intra-and inter-day precisions were examined. The RSDs of peak area and migration time were found to be below 5.0% and 2.5% (n=6), respectively, indicating good repeatability (intra-day) and reproducibility (inter-day) of the method. Interference experiment was also tested. As a result, other common amino acids did not interfere with the detection. The proposed method provided a potential new way to further investigate the enantioseparation of unmodified or native amino acids.
Assuntos
Ácido Cítrico , Eletroforese Capilar , Isoleucina/isolamento & purificação , Zinco , Soluções Tampão , Reprodutibilidade dos Testes , EstereoisomerismoRESUMO
BACKGROUND: Fenugreek (Trigonella foenum-graecum) is globally recognized for its medicinal properties and hypoglycemic effects. The seed extract as well as its active compound, 4-hydroxyisoleucine (4-OH-Ile), have been shown to reduce hyperglycemic insulin resistance. The mechanism by which this occurs has not been investigated in human liver cells (HepG2) in comparison to the antihyperglycemic drug, metformin. METHODS: We investigated the effects of an aqueous fenugreek seed extract (FSE), 4-OH-Ile, and metformin in HepG2 cells relative to insulin as a positive control. Cells were treated with FSE and 4-OH-Ile at 100 ng/mL under normoglycemic (5 mM glucose) and hyperglycemic (30 mM glucose) conditions for 72 hr. Tyrosine phosphorylation of insulin receptor-ß (IR-ß), protein kinase B (Akt), glycogen synthase kinase-3α/ß (GSK-3α/ß), and glucose transporter 2 (GLUT2) was determined by western blotting. Gene expression of sterol regulatory element-binding protein 1c (SREBP1c), GLUT2, glycogen synthase (GS), and glucokinase (GK) was evaluated by quantitative polymerase chain reaction, and supernatant glucose levels were measured using the Piccolo biochemistry analyzer. RESULTS: Under normo- and hyperglycemic conditions, FSE, 4-OH-Ile, insulin (100 ng/mL), and metformin (2 mM) caused a significant increase in phosphorylation of IR-ß, Akt, GSK-3α/ß, and GLUT2. Glucose uptake, however, was most significantly increased in FSE-treated cells during both conditions. FSE induced the most significant changes in downstream insulin signaling, GS, GK, SREBP1c, and GLUT2 expression compared to 4-OH-Ile, metformin, and insulin. In addition, FSE significantly increased glucose uptake. CONCLUSIONS: Collectively, these findings provide a mechanism by which FSE exerts antihyperglycemic effects similar to metformin and insulin that occurs via enhanced insulin signaling, gene expression, and increasing glucose uptake.
Assuntos
Enzimas/metabolismo , Transportador de Glucose Tipo 2/metabolismo , Hepatócitos/efeitos dos fármacos , Hipoglicemiantes/farmacologia , Insulina/metabolismo , Isoleucina/análogos & derivados , Metformina/farmacologia , Extratos Vegetais/farmacologia , Transdução de Sinais/efeitos dos fármacos , Antígenos CD/metabolismo , Enzimas/genética , Transportador de Glucose Tipo 2/genética , Quinase 3 da Glicogênio Sintase/metabolismo , Glicogênio Sintase Quinase 3 beta/metabolismo , Células Hep G2 , Hepatócitos/enzimologia , Humanos , Isoleucina/isolamento & purificação , Isoleucina/farmacologia , Fosforilação , Fitoterapia , Plantas Medicinais , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor de Insulina/metabolismo , Trigonella , Regulação para CimaRESUMO
We report the characterization of a new insulinotropic compound, 4-hydroxyisoleucine. This amino acid has been extracted and purified from fenugreek seeds, which are known in traditional medicine for their antidiabetic properties. 4-Hydroxyisoleucine increases glucose-induced insulin release, in the concentration range of 100 micromol/l to 1 mmol/l, through a direct effect on isolated islets of Langerhans from both rats and humans. The stimulating effect of 4-hydroxyisoleucine was strictly glucose dependent; indeed, ineffective at low (3 mmol/l) or basal (5 mmol/l) glucose concentrations, the amino acid potentiated the insulin secretion induced by supranormal (6.6-16.7 mmol/l) concentrations of glucose. In addition, in the isolated perfused rat pancreas, we could show 1) that the pattern of insulin secretion induced by 4-hydroxyisoleucine was biphasic, 2) that this effect occurred in the absence of any change in pancreatic alpha- and delta-cell activity, and 3) that the more glucose concentration was increased, the more insulin response was amplified. Moreover, 4-hydroxyisoleucine did not interact with other agonists of insulin secretion (leucine, arginine, tolbutamide, glyceraldehyde). Therefore, we conclude that 4-hydroxyisoleucine insulinotropic activity might, at least in part, account for fenugreek seeds' antidiabetic properties. This secretagogue may be considered as a novel drug with potential interest for the treatment of NIDDM.
Assuntos
Hipoglicemiantes , Insulina/metabolismo , Isoleucina/análogos & derivados , Extratos Vegetais/química , Animais , Glucose/farmacologia , Humanos , Secreção de Insulina , Ilhotas Pancreáticas/efeitos dos fármacos , Ilhotas Pancreáticas/metabolismo , Isoleucina/administração & dosagem , Isoleucina/isolamento & purificação , Isoleucina/farmacologia , Cinética , Masculino , Plantas Medicinais , Ratos , Ratos Wistar , TrigonellaRESUMO
Previous studies have indicated that 4hydroxyisoleucine (4HIL) improves insulin resistance, however, the underlying mechanisms remain to be elucidated. In the present study, the molecular mechanisms underlying how 4HIL improves insulin resistance in hepatocytes were examined. HepG2 cells were cocultured with insulin and a high glucose concentration to obtain insulinresistant (IR) HepG2 cells. Insulin sensitivity was determined by measuring the glucose uptake rate. The IR HepG2 cells were treated with different concentrations of 4HIL to determine its effect on IR Hep2 cells. The levels of tumor necrosis factorα (TNFα) were measured by an enzymelinked immunosorbent assay and protein levels of TNFα converting enzyme (TACE)/tissue inhibitor of metalloproteinase 3 (TIMP3), insulin receptor substrate (IRS)1, IRS2, phosphorylated (p)IRS1 (Ser307) and glucose transporter type 4 (GLUT4) were measured by western blot analysis. The results of the present study demonstrated that insulininduced glucose uptake was reduced in IR HepG2 cells; however, this reduction was reversed by 4HIL in a dosedependent manner. 4HIL achieved this effect by downregulating the expression of TNFα and TACE, and upregulating the expression of TIMP3 in IR HepG2 cells. In addition, 4HIL increased the expression of the insulin transduction regulators IRS1 and GLUT4, and decreased the expression of pIRS1 (Ser307), without affecting the expression of IRS2. The present study suggests that 4HIL improved insulin resistance in HepG2 cells by the following mechanisms: 4HIL reduced TNFα levels by affecting the protein expression of the TACE/TIMP3 system and 4HIL stimulated the expression of IRS1 and GLUT4, but inhibited the expression of pIRS1 (Ser307).
Assuntos
Glucose/farmacologia , Resistência à Insulina , Insulina/farmacologia , Isoleucina/análogos & derivados , Transdução de Sinais/efeitos dos fármacos , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Proteínas ADAM/genética , Proteínas ADAM/metabolismo , Proteína ADAM17 , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica , Glucose/metabolismo , Transportador de Glucose Tipo 4/agonistas , Transportador de Glucose Tipo 4/genética , Transportador de Glucose Tipo 4/metabolismo , Células Hep G2 , Humanos , Insulina/metabolismo , Proteínas Substratos do Receptor de Insulina/agonistas , Proteínas Substratos do Receptor de Insulina/genética , Proteínas Substratos do Receptor de Insulina/metabolismo , Isoleucina/isolamento & purificação , Isoleucina/farmacologia , Fosforilação/efeitos dos fármacos , Transdução de Sinais/genética , Inibidor Tecidual de Metaloproteinase-3/genética , Inibidor Tecidual de Metaloproteinase-3/metabolismo , Trigonella/química , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismoRESUMO
N-Bromosuccinimide cleavage reaction on cerexin A liberated allo-isoleucine. Treatment with conc. hydrochloric acid cleaved the antibiotic into two peptide fragments selectively at gamma-hydroxylysine residue. Deacylation with an enzyme preparation from Pseudomonas sp. afforded deacyl cerexin A. The amino acid sequences of these peptide fragments were examined by Edman degradation. From all the results, the entire amino acid sequence of cerexin A was deduced.
Assuntos
Antibacterianos , Bacillus cereus/química , Sequência de Aminoácidos , Antibacterianos/isolamento & purificação , Bromosuccinimida , Fenômenos Químicos , Química , Ácido Clorídrico , Hidrólise , Isoleucina/isolamento & purificação , Peptídeos/análise , Treonina/análise , Triptofano/isolamento & purificaçãoRESUMO
WF14865A and B, novel cathepsins B and L inhibitors, were produced and isolated separately from the culture mycelium of a fungal strain Aphanoascus fulvescens No. 14865. Spectroscopic analysis revealed that both WF14865A and B were composed of trans-epoxysuccinyl moieties, 1-H-imidazole-2-ylamine, and isoleucine or leucine. These compounds inhibited human cathepsins B and L with IC50 values in the range of 8.4 approximately 72nM in vitro. Though their in vitro properties were typical as trans-epoxysuccinyl type inhibitors, they exerted strong bone resorption inhibitory effects in low-calcium-diet-fed mouse model at 3.2 approximately 10 mg/kg.
Assuntos
Ascomicetos/metabolismo , Catepsina B/antagonistas & inibidores , Catepsinas/antagonistas & inibidores , Inibidores de Cisteína Proteinase/classificação , Endopeptidases , Compostos de Epóxi/química , Isoleucina/análogos & derivados , Animais , Catepsina L , Cromatografia Líquida , Cisteína Endopeptidases , Inibidores de Cisteína Proteinase/isolamento & purificação , Inibidores de Cisteína Proteinase/farmacologia , Compostos de Epóxi/isolamento & purificação , Compostos de Epóxi/farmacologia , Feminino , Humanos , Isoleucina/química , Isoleucina/isolamento & purificação , Isoleucina/farmacologia , Fígado/enzimologia , Camundongos , Camundongos Endogâmicos ICRRESUMO
The conditions of L-leucine and L-isoleucine esterification were investigated. It was found that chromatographically homogeneous L-isoleucine and L-leucine could be isolated from their mixture during its incomplete esterification. As a result, L-leucine transformed into L-leucine and ester chlorohydrate L-isoleucine into L-isoleucine chlorohydrate. By means of further neutralization of the reaction mixture L-isoleucine with a 80-85% yield was obtained, and with the aid of L-leucine ester hydrolysis L-leucine with a 80-85% yield was produced. The content of the major amino acid following its crystallization from the water-ethanol mixture was 99%.
Assuntos
Isoleucina/isolamento & purificação , Leucina/isolamento & purificação , Cromatografia em Camada Fina , Combinação de Medicamentos , Esterificação , Concentração de Íons de Hidrogênio , Temperatura , Fatores de TempoRESUMO
Staphylococcus aureus is responsible for many types of infections related to biofilm presence. As the early diagnostics remains the best option for prevention of biofilm infections, the aim of the work presented was to search for differences in metabolite patterns of S. aureus ATCC6538 biofilm vs. free-swimming S. aureus planktonic forms. For this purpose, Nuclear Magnetic Resonance (NMR) spectroscopy was applied. Data obtained were supported by means of Scanning Electron Microscopy, quantitative cultures and X-ray computed microtomography. Metabolic trends accompanying S. aureus biofilm formation were found using Principal Component Analysis (PCA). Levels of isoleucine, alanine and 2,3-butanediol were significantly higher in biofilm than in planktonic forms, whereas level of osmoprotectant glycine-betaine was significantly higher in planktonic forms of S. aureus. Results obtained may find future application in clinical diagnostics of S. aureus biofilm-related infections.
Assuntos
Biofilmes/crescimento & desenvolvimento , Metaboloma , Infecções Estafilocócicas/diagnóstico , Staphylococcus aureus/metabolismo , Alanina/isolamento & purificação , Alanina/metabolismo , Biomarcadores/metabolismo , Butileno Glicóis/isolamento & purificação , Butileno Glicóis/metabolismo , Hidrogênio , Isoleucina/isolamento & purificação , Isoleucina/metabolismo , Espectroscopia de Ressonância Magnética , Radioisótopos , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/crescimento & desenvolvimento , Staphylococcus aureus/isolamento & purificação , Tomografia Computadorizada por Raios XRESUMO
Trigonella foenum-graecum, commonly known as fenugreek, is an annual herbaceous plant. From the seeds of T. foenum-graecum an unusual amino acid, 4-hydroxyisoleucine 5, has been isolated, which significantly decreased the plasma triglyceride levels by 33% (P<0.002), total cholesterol (TC) by 22% (P<0.02), and free fatty acids by 14%, accompanied by an increase in HDL-C/TC ratio by 39% in the dyslipidemic hamster model.
Assuntos
Hipoglicemiantes/uso terapêutico , Hipolipemiantes/uso terapêutico , Isoleucina/análogos & derivados , Administração Oral , Animais , Colesterol/sangue , Colesterol/metabolismo , HDL-Colesterol/sangue , HDL-Colesterol/efeitos dos fármacos , Cricetinae , Diabetes Mellitus Tipo 2/tratamento farmacológico , Gorduras na Dieta/administração & dosagem , Gorduras na Dieta/efeitos adversos , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Ácidos Graxos/sangue , Ácidos Graxos/metabolismo , Hipoglicemiantes/administração & dosagem , Hipoglicemiantes/isolamento & purificação , Hipolipemiantes/administração & dosagem , Hipolipemiantes/isolamento & purificação , Isoleucina/administração & dosagem , Isoleucina/isolamento & purificação , Isoleucina/uso terapêutico , Masculino , Estrutura Molecular , Sementes/química , Relação Estrutura-Atividade , Triglicerídeos/sangue , Triglicerídeos/metabolismo , Trigonella/químicaRESUMO
In an attempt to improve the sensitivity of detection in capillary electrophoresis (CE), a novel online sample-concentration method, full-capillary sample stacking (FCSS)/sweeping-micellar electrokinetic chromatography (sweeping-MEKC) mode, is proposed. Naphthalene-2,3-dicarboxaldehyde (NDA)-derivatized tryptophan and isoleucine were selected as model compounds. In the initial step, the weakly acidic compounds, dissolved in a low-conductivity buffer (35.1 microS/cm; apparent ph (pH*) in a mixed solution of acetonitrile/methanol/water, 4.6), fill the entire capillary, two vials of a high-conductivity buffer (2.06 mS/cm; pH* 2.0) are placed on each end, and a negative polarity is then applied. Under these conditions, the direction of the electroosmotic flow (EOF) is toward the inlet. Meanwhile, the anionic analytes move in the reverse direction and are neutralized and stacked at the boundary of a dynamic pH-junction (between the sample matrix and the nonmicellar background solution (BGS)). When the sample concentration is completed, the BGS is quickly changed to solutions containing SDS-BGS for the subsequent separation. Since the mobility of SDS-analytes is then greater than the EOF, the following steps occur by the sweeping (for focusing) and MEKC (for separation) mode. Using these steps, a full-capillary sample injection/separation can be achieved.
Assuntos
Cromatografia Capilar Eletrocinética Micelar/métodos , Isoleucina/análogos & derivados , Isoleucina/isolamento & purificação , Naftalenos/química , Triptofano/análogos & derivados , Triptofano/isolamento & purificação , Sensibilidade e EspecificidadeRESUMO
Some underivatized DL-amino acid enantiomers were separated on L-isoleucine-type chiral-ligand-exchange stationary phase (EILE). The effects of contents of copper ion and methanol in mobile phase, temperature of column and flow rate of eluent on the efficiency and resolution of DL-amino acids were investigated in detail. The results showed that Cu(II) ion played an important role in the separation. When there was no Cu(II) ion in mobile phase, the DL-amino acids could not be enantio-separated, while the content of Cu(II) increased from 0.3 mmol/L to 1.2 mmol/L, the efficiency of resolution and retention time decreased. The increases of the content of methanol and column temperature are in favor of the resolution for most of DL-amino acids. The flow rate also affected the resolution results. Good results could be obtained in reducing flow rate but it prolongs the retention time. The mechanisms of different chiral separation results on the new stationary phase under different conditions were also investigated.
Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Isoleucina/isolamento & purificação , Cromatografia Líquida de Alta Pressão/instrumentação , Cobre/química , Metanol/química , Estrutura Molecular , Estereoisomerismo , TemperaturaRESUMO
The simplest RNA that can meet a column affinity selection for isoleucine was previously defined using selection amplification with decreasing numbers of randomized nucleotides. This simplest UAUU motif was a small asymmetric internal loop. Conserved positions of the loop include isoleucine codon and anticodon triplets (Lozupone C., Changayil, S., Majerfeld, I., and Yarus, M. (2003) RNA (N. Y.) 9, 1315-1322). Using new primer sequences, we now select a somewhat more complex isoleucine binding RNA, requiring 4.7 more bits of information to describe. The newly selected structure is a terminal or hairpin loop of 20 nucleotides, 15 being invariant. An information profile shows that the new binding site contains five short functional loop regions joined by less significant single nucleotide positions. Among the important nucleotides is a conserved isoleucine anticodon, supporting the escaped triplet theory, which posits a stereochemical genetic code originating in RNA amino acid binding sites.
Assuntos
Isoleucina/química , RNA/química , Anticódon/química , Anticódon/genética , Sequência de Bases , Sítios de Ligação/genética , Cromatografia de Afinidade , Códon/química , Códon/genética , Isoleucina/genética , Isoleucina/isolamento & purificação , Conformação de Ácido Nucleico , Oligorribonucleotídeos/química , Oligorribonucleotídeos/genética , RNA/genéticaRESUMO
Alloisoleucine, prepared from a mixture of racemic isoleucine-alloisoleucine diastereoisomers, can be heavily contaminated with isoleucine if the recrystallization of acetyl-dl-alloisoleucine is controlled by melting point only. Two commercial specimens were found to be thus contaminated.
Assuntos
Isoleucina/isolamento & purificação , Cristalização , EstereoisomerismoRESUMO
High-voltage paper electrophoresis and analytical and preparative amino acid ion-exchange chromatography were used to investigate the products resulting from strong acid treatment and strong acid treatment and subsequent alkalinization of alpha-amino-delta-hydroxyvaleric acid. After strong acid treatment, alpha-amino-delta-hydroxyvaleric acid and proline, as well as several unidentified components, presumably lactones, were recovered. No definitive identification of the putative unstable lactones was accomplished either by physical-chemical means or by ion-exchange chromatography or high-voltage paper electrophoresis. Nevertheless, after acid treatment and alkalinization, only alpha-amino-delta-hydroxyvaleric acid and proline were recovered, in approximately equal amounts. The formation of such large amounts of proline from the acid treatment and alkalinization of alpha-amino-delta-hydroxyvaleric acid is therefore an important consideration in quantitating the number of gamma-glutamyl phosphate residues present in various proteins.
Assuntos
Aminoácidos/isolamento & purificação , Ácidos , Cromatografia por Troca Iônica , Eletroforese em Papel , Isoleucina/isolamento & purificação , Lactonas/isolamento & purificação , Prolina/isolamento & purificaçãoRESUMO
Of the four possible stereoisomers of isoleucine, only L-alloisoleucine and L-isoleucine were found by capillary gas chromatography in the plasma of two maple syrup urine disease (MSUD) patients, one with classical and one with variant MSUD. The relative plasma concentration ratios of L-alloisoleucine/L-isoleucine were 0.795 +/- 0.025 (+/- 95% confidence limits) and 0.637 +/- 0.016 in the classical- and variant-MSUD patients, respectively. The patients were also studied in the postabsorptive state with a 6-hr continuous infusion of L-[15N]leucine. In each patient plasma leucine 15N enrichment approximated plateau after 150 min, and there was rapid appearance of [15N]isoleucine and [15N]alloisoleucine which were identical at plateau, although in variant-MSUD patient [15N]alloisoleucine enrichment did not equal that of [15N]isoleucine until 240 min of infusion. These results offer direct in vivo evidence for the rapid equilibrium of plasma isoleucine and alloisoleucine through keto-enol tautomerization of alpha-keto-beta-methylvalerate.