Acids Arginine Research Articles

Minerals and Amino Acids in Peach (Prunus persica L.) Cultivars and Hybrids Belonging to World Germoplasm Collection in the Conditions of West Romania

Abstract In this study carried out, the nutritional profile of 19 peach cultivars and hybrids from west side of Romania in terms of selected amino acids, macro and microelements content. Amino acids (arginine, lysine, alanine, phenylalanine, tyrosine, serine and glycine) were detected and quantified using DIONEX ICS-3000 system, while the macroelements (K, Ca, Mg) and microelements (Fe, Cu, Zn, Mn) were determined by atomic absorption spectroscopy (Varian 220 AAS equipment). The obtained results showed major differences in content of individual aminoacids in peach fruits. Alanine, responsible for the sweet taste of fruit, but also lysine and arginine are found in higher quantities. The cultivars ‘JULY ELBERTA’ and HB4/81 presented higher arginine content, ‘MARQUEEN’ and IFF853 lysine and ‘MARIANA’ variety has alanine and phenylalanine more than other cultivars. The macro and microelements profile depended on cultivar and increased in following order: K>Mg>Ca>Fe>Zn>Cu>Mn.

Comparative Analysis of Gene Expression Profiling Between Resistant and Susceptible Varieties Infected With Soybean Cyst Nematode Race 4 in Glycine max

Soybean cyst nematode (SCN) is one of the most devastating pathogen for soybean. Therefore, identification of resistant germplasm resources and resistant genes is needed to improve SCN resistance for soybean. Soybean varieties Huipizhiheidou and Wuzhaiheidou were distributed in China and exhibited broad spectrums of resistance to various SCN races. In this study, these two resistant varieties, combined with standard susceptible varieties (Lee and Essex), were utilized to identify the differentially expressed transcripts after infection with SCN race 4 between resistant and susceptible reactions by using the Affymetrix Soybean Genome GeneChip. Comparative analyses indicated that 21 common genes changed significantly in the resistant group, of which 16 increased and 5 decreased. However, 12 common genes changed significantly in the susceptible group, of which 9 increased and 3 decreased. Additionally, 27 genes were found in common between resistant and susceptible reactions. The 21 significantly changed genes in resistant reaction were associated with disease and defense, cell structure, transcription, metabolism, and signal transduction. The fold induction of 4 from the 21 genes was confirmed by quantitative RT-PCR (qRT-PCR) analysis. Moreover, the gene ontology (GO) enrichment analyses demonstrated the serine family amino acid metabolic process and arginine metabolic process may play important roles in SCN resistance. This study provided a new insight on the genetic basis of soybean resistance to SCN race 4, and the identified resistant or resistant-related genes are potentially useful for SCN-resistance breeding in soybean.

Karakteristik Kimiawi dan Potensi Pemanfaatan Dunaliella salina dan Nannochloropsis

The objective of this research was to know the chemical characteristics and the potential utilization of D.salina and Nannochloropsis sp. The research has been conducted from December 2010 until April 2012 in Technology Of Fishery Product Laboratory, Aquaculture Laboratory, Bioprocess Laboratory University of Sriwijaya and Integrated Laboratory Bogor Agricultural University. The parameters observed were biomass dry weight, proximate (moisture content, ash content, protein content, fat content, and total carbohydrate content), carotenoid, and amino acids. Proximate composition of Nannochloropsis sp. obtained in this research were 58.00% ash content, moisture content 12.39%, protein content 16.17%, fat content 0.30%, total carbohydrate content 19.08%, total carotenoids 0.27 ppm, and essential amino acids (histidine, threonine, arginine , methionine, phenylalanine, valine, isoleucine, and leucine) and non-essential amino acids (aspartic acid, glutamic acid, serine, glycine, alanine, and tyrosine). As for the sample D.salina were the ash content 58.29%, water content 15.58%, protein content 17.08%, fat content 0.003%, total carbohydrate content 15.07%, total carotenoids 0.19 ppm, and essential amino acids (histidine , threonine, arginine, methionine, phenylalanine, valine, isoleucine, leucine, and lysine) and non-essential amino acids (aspartic acid, glutamic acid, serine, glycine, alanine, and tyrosine). From the results of data analysis and chemical characteristics of dry weight D.salina and Nannochloropsis sp. obtained in this study, biomass produced this research has the chemical characteristics of lower quality than the quality requirements of microalgae as an industrial raw material of food, feed, biodiesel and bioethanol.

Two basic (hydrophilic) regions in the movement protein of Parietaria mottle virus have RNA binding activity and are required for cell-to-cell transport

The movement protein (MP) of parietaria mottle virus (PMoV) is required for virus cell-to-cell movement. Bioinformatics analysis identified two hydrophilic non-contiguous regions (R1 and R2) rich in the basic amino acids lysine and arginine and with the predicted secondary structure of an α-helix. Different approaches were used to determine the implication of the R1 and R2 regions in RNA binding, plasmodesmata (PD) targeting and cell-to-cell movement. EMSA (Electrophoretic Mobility Shift Assay) showed that both regions have RNA-binding activity whereas that mutational analysis reported that either deletion of any of these regions, or loss of the basic amino acids, interfered with the viral intercellular movement. Subcellular localization studies showed that PMoV MP locates at PD. Mutants designed to impeded cell-to-cell movement failed to accumulate at PD indicating that basic residues in both R1 and R2 are critical for binding the MP at PD.

A Novel Recombinant Human Granulocyte Colony-Stimulating Factor (G-CSFa) Enhances Peripheral Platelet Recovery in Mice Exposed to adiotherapy and Has No Immunogenicity in Rats

Using site-direct mutagenesis and recombinant DNA technology, we had previously obtained a structurally modified derivative of human G-CSF termed G-CSFa. G-CSFa contains alanine 17 (instead of cysteine 17 as in wildtype G-CSF) as well as four additional amino acids (methionine, arginine, glycine and serine) at the amino terminus. Previous studies showed that G-CSFa is more potent than the wild-type counterpart in stimulating proliferation and differentiation of myeloid cells of the granulocytic lineage, both in vitro and in vivo. Here, we show that G-CSFa can significantly accelerate peripheral platelet recovery in C57BL/6 mice exposed to radiotherapy. We further demonstrate that G-CSFa is not immunogenic in rats, by confirming the absence of any binding antibodies, analyzed using ELISA, or neutralizing antibodies, determined using the NFS-60 cell proliferation bioassay, to G-CSFa in the sera of Sprague-Dawley rats following repeated G-CSFa administration. Taken together, these findings further support the benefits of G-CSFa for clinical therapy.

Seasonal Variation on General Composition, Free Amino Acids and Fatty Acids in the Gonad of Taiwan's Sea Urchin Tripneustes gratilla

The seasonal variation of chemical composition in sea urchin Tripneustes gratilla gonad from Taiwan is investigated. The level of both gonad index (GI) and body weight in the sea urchins showed seasonal variations and were higher during spring and summer. The main saturated fatty acids (SFA) of sea urchin gonad were C14:0 and C16:0, and both varied with season. Arachidonic acid and eicosapentaenoic acid were the dominant poly-unsaturated fatty acids (PUFA), while C16:1, C18:1 and C22:1 were the predominant monoenoic fatty acids (MFA). The levels of total fatty acids were higher in summer, and decreased to the lowest in December (13.2 ± 2.3 mg/g). The SFA level was higher in summer, while the PUFA level was higher in winter. The most dominant free amino acid (FAA) of sea urchin gonad was glycine, where alanine, proline, arginine and glutamic acid were the predominant FAAs. Arginine and valine were in increasing from September to February. Meanwhile glycine, alanine and glutamic acid also presented seasonal change. Overall, the gonad of Taiwan's sea urchin T. gratilla offered higher gonad content, better color and higher level of lipid in summer. It suggested the better period for consuming T. gratilla in Taiwan is from May to July.

RBC deformability and amino acid concentrations after hypo-osmotic challenge may reflect chronic cell hydration status in healthy young men

Biomarkers of chronic cell hydration status are needed to determine whether chronic hyperosmotic stress increases chronic disease risk in population-representative samples. In vitro, cells adapt to chronic hyperosmotic stress by upregulating protein breakdown to counter the osmotic gradient with higher intracellular amino acid concentrations. If cells are subsequently exposed to hypo-osmotic conditions, the adaptation results in excess cell swelling and/or efflux of free amino acids. This study explored whether increased red blood cell (RBC) swelling and/or plasma or urine amino acid concentrations after hypo-osmotic challenge might be informative about relative chronic hyperosmotic stress in free-living men. Five healthy men (20–25 years) with baseline total water intake below 2 L/day participated in an 8-week clinical study: four 2-week periods in a U-shaped A-B-C-A design. Intake of drinking water was increased by +0.8 ± 0.3 L/day in period 2, and +1.5 ± 0.3 L/day in period 3, and returned to baseline intake (0.4 ± 0.2 L/day) in period 4. Each week, fasting blood and urine were collected after a 750 mL bolus of drinking water, following overnight water restriction. The periods of higher water intake were associated with significant decreases in RBC deformability (index of cell swelling), plasma histidine, urine arginine, and urine glutamic acid. After 4 weeks of higher water intake, four out of five participants had ½ maximal RBC deformability below 400 mmol/kg; plasma histidine below 100 μmol/L; and/or undetectable urine arginine and urine glutamic acid concentrations. Work is warranted to pursue RBC deformability and amino acid concentrations after hypo-osmotic challenge as possible biomarkers of chronic cell hydration.

Precursors for ornithine and citrulline synthesis in neonatal pigs

Citrulline (CIT) is an amino acid synthesized by gut and utilized for the synthesis of the conditionally essential amino acid arginine (ARG). In turn, the immediate precursor for CIT synthesis, ornithine (ORN), can originate from proline (PRO) and glutamine (GLN) via ornithine aminotransferase (OAT, i.e., de novo synthesis) or from ARG via arginase. To test the hypothesis that during the neonatal period de novo synthesis is the main source of ORN for CIT synthesis, neonatal pigs were infused iv or ig with [U‐13C6]ARG, [U‐13C5]GLN or [U‐13C5]PRO during the fasted and fed periods. (ureido)[15N]CIT and [D2]ORN were infused iv for the entire infusion protocol. During fasting plasma PRO (13%) and ORN (19%) were the main precursors for CIT synthesis, whereas plasma ARG (62%) was the main precursor for plasma ORN. During feeding enteral (27%) and plasma (12%) PRO were the main precursors for the ORN utilized in the synthesis of CIT, together with plasma ORN (27%). Enteral PRO and GLN were utilized directly by the gut to produce ORN utilized for CIT synthesis. ARG was not utilized by the gut, which is consistent with the lack of arginase activity in neonates. ARG, however, was the main source (47%) of plasma ORN and in this way contributed to CIT synthesis. During the neonatal period the directionality of OAT, together with the lack of arginase activity in the gut, determine that the de novo pathway is the main source of ORN for CIT production.

Determination of plasma dibasic amino acids following trimethylsilyl–trifluoroacyl derivatization using gas chromatography–mass spectrometry

A rapid analytical method was developed to quantify dibasic amino acids (ornithine, lysine and arginine) after two-step derivatization procedure with good sensitivity and specificity on human plasma. If early diagnosis has not been made, patients with inborn metabolic disorders such as HHH syndrome, Hyperornithinemia and dibasic aminoaciduria rapidly progress to sudden death, physical defect or mental retardation resulting in storage of the toxic material into the brain. Therefore, it is necessary to develop the analytical method for rapid screening and/or correct confirmation diagnosis. The formation of trimethylsilyl derivative of the carboxylic (COO-) functional group was performed by adding MSTFA. Five μL of methyl orange was added to the residue until the color changed into yellow. Consecutively, the trifluoroacyl derivative of the amino (-NH2) functional group was produced by adding MBTFA. Specific ions was chosen for quantification with following ions; m/z 166 and m/z 212 for ornithine, m/z 180 and m/z 395 for lysine, and m/z 292 and, m/z 519 for arginine. A calibration curve showed a linear relationship for the dibasic amino acids spiked to pooled normal plasma showing R(2) of 0.9955-0.9979 in the range of 0.1-600 ng investigated. The utility of the method for screening and diagnosis was demonstrated by recovery 80-125 % and reproducibility with RSD (9-17 %) at low, medium and high concentration fortified to pooled plasma. Collectively, the present study suggest that this method could be useful for diagnosis, screening, therapeutic monitoring of metabolic disorders on dietary therapy with excellent sensitivity and rapidity.

IMPROVED APPROACH FOR THE LABELING OF ARGININE, GLUTAMIC, AND ASPARTIC ACID SIDE CHAINS IN PROTEINS USING CHROMATOGRAPHIC TECHNIQUES

Specific chemical modification is one of the basic techniques of protein chemistry. Inter alia can be used for detection of surface accessible amino acid residues; this information is of particular importance for studies of the participation of residues in intermolecular interactions of a protein. We achieved an improvement of the technique for arginine, aspartic, and glutamic acid modification using a simple combination of gel permeation and reversed-phase chromatography prior to mass spectrometry analysis. The improved protocol was tested on cytochrome c and M-PMV matrix protein. In both proteins, all accessible arginines and a high number of acidic amino acids were modified. These results indicate that the new protocol can be useful in protein structure analysis, generally.

Biochemical Composition during the Embryonic Development and Freshly Hatched Zoea of Macrobrachium idae (Heller, 1862)

In the present study an attempt has been made to know the biochemical composition of freshly hatched zoea of Macrobrachium idea. The content of protein in egg was gradually decreased from egg stage-I (63.98%) to egg stage-IV (51.85%), carbohydrate increased from egg stage-I (3.06%) to freshly hatched 1st zoea (4.06%), lipid values showed a decreasing trend from egg stage-I (14.97%) to freshly hatched 1st zoea (4.86%) and moisture increased from egg stage-I (61.68%) to freshly hatched 1st zoea stage (78.80%). The fatty acid composition of egg stage and zoea was in the range of 32.56-41.60 % for saturated, 26.09-36.22 % for monounsaturated and 9.07-11.92 % for polyunsaturated respectively and they were high in egg stages - I, III and IV. The unsaturated fatty acid contents were higher than the saturated fatty acid content. The amino acids showed increasing trends from egg stage-I to IV. Among the amino acids leucine, lysine, valine, methionine and arginine increased gradually as development progressed in egg stages and isoleucine, phenylalanine, histidine, tryptophan and threonine had a change of high-low-high.

Response Chamomile Plants (Matricaria chamomilla L.) to Plant Distance and Spraying with Two Amino Acid Proline and Arginine and their Affectivity on Growth, Flower Yield and Volatile Oil Content and its Quality

Response Chamomile Plants (Matricaria chamomilla L.) to Plant Distance and Spraying with Two Amino Acid Proline and Arginine and their Affectivity on Growth, Flower Yield and Volatile Oil Content and its Quality

The influence of flushes on some constituents of mushrooms (Agaricus bisporus) cultivated on different composts

The effect of two different composts: horse manure and broiler chicken manure and the influence of flushes during the growing cycle on some chemical components contained in the mushroom (<i>Agaricus bisporus</i>) was studied. The strain Somycel 11 was analyzed. The following determinations were made: dry matter, total nitrogen and crude protein, nitrates, nitrites and amino acids composition. It has been noted that the crude protein from. combined flushes III to VI of mushrooms cultivated on horse manure was about 24% higher than in those grown on broiler chicken manure. Some changes in amino acids composition, especially lysine, histidine, arginine, aspartic acid, serine, methionine, glutamic acid and alanine were observed, depending on the size of the fruit-body, flushes and type of compost.

Identification of amino acid residues essential to the activity of lyase CpcT1 from Nostoc sp. PCC7120

The phycocyanin lyase CpcT1 (encoded by gene all5339) and lyase CpcS1 (encoded by gene alr0617) are capable of catalyzing the phycocyanobilin (PCB) covalently bound to the different sites of phycocyanin's and phycoerythrocyanin's β subunits, respectively. Lyase CpcS1, whose catalytic mechanism had been researched clearly, participates in the covalent coupling of phycobilin and apoprotein in the form of chaperone, and its important amino acids have been confirmed. In order to identify the functional amino acid residues of CpcT1, chemical modification was conducted to arginine, histidine, tryptophan, lysine and amino acid carboxyl of CpcT1. The results indicated that the catalytic activity of the CpcT1 was changed. After the modification of arginine, tryptophan and histidine, site-directed mutations were performed to those highly conserved amino acids which were selected by means of homologous comparison. The mutated lyase, apoprotein and the enzymes that synthesize the phycobilins were recombined in Escherichia coli (E. coli) and in vitro, yielding chromoproteins, which were detected by fluorescence and UV absorption spectrometry. The spectra were compared with that of the chromoprotein catalyzed by wild type lyase CpcT1, achieving relative specific activities of the various mutants. Meanwhile, the mutants were expressed in E. coli, and then circular dichroism structure of near-UV region was determined. The results demonstrated that H33F, W175S, R97A, C137S and C116S influence the catalytic activity of CpcT1. Being different from wild CpcT1, a great deal of α helix was involved in the structure of circular dichroism of R97A and W13S. CpcT1 or its mutants and the enzymes that synthesize the phycobilins, were reconstituted in E. coli and detected by spectra to check the bounding of lyases and PCB. The results of spectra and SDS-PAGE confirm that CpcT1 and its mutants cannot bind phycobilin, differing from the catalytic mechanism of CpcS1.

Estimation of Protein in Anhydrous Milk Fat

A method for the estimation of protein in anhydrous milk fat is described. The protein concentration is estimated by quantifying the arginine and aspartic acid (ASX) released upon acid hydrolysis of the anhydrous milk fat. Arginine and aspartic acid are derivatized with the fluorescent tag 9-fluorenylmethoxycarbonyl (FMOC), the derivatives are quantified by reversed phase HPLC, and the protein concentration in the anhydrous milk fat is estimated as 8.77 × (Arg + ASX). Method suitability was defined by experimental assessments of FMOC-arginine and FMOC-aspartic acid linearity (R 2 averages > 0.999), protein estimate precision [day-to-day RSD values (n = 8 days) ranged from 7.3 to 14 % for protein concentrations of 9.42 to 40.0 mg/kg of anhydrous milk fat], protein estimate accuracy [spike recovery average = 95.6 % (9.3 %), n = 8, spiking level = 20 mg/kg and agreement of the experimental protein estimate for butter, 0.611 (0.044) g/100 g, n = 3 lots, with a published value 0.6/100 g], and analyte selectivity (baseline resolution of FMOC-arginine and FMOC-aspartic acid from the FMOC derivatives of other common amino acids). The method provides for a reliable estimation of the protein content of anhydrous milk fat, when present at concentrations >2 mg/kg.

Mutation analysis of keratin 9 gene in a family with epidermolytic palmoplantar keratoderma

To analyze potential mutation of keration 9 gene (KRT9) in a Chinese family affected with epidermolytic palmoplantar keratoderma (EPPK) and to correlate genotype with the phenotype. Genomic DNA was extracted from peripheral blood samples of 12 patients and 13 healthy individuals from the family and 100 unrelated individuals. Polymerase chain reaction (PCR) was used to amplify exons 1 and 6 of KRT9 gene. PCR products were sequenced bidirectionally in order to identify potential mutations. A heterozygous transversional mutation, 488G→A, was identified in exon 1 of KRT9 gene in all patients, which has resulted in substitution of a glutamine residue for arginine acid at position 163 (R163Q) of the KRT9 protein. The same mutation was not found in the 13 healthy members from the family and 100 unrelated individuals. The 488G→A mutation of KRT9 gene is probably the cause of EPPK in this Chinese family.

Improvement of clavulanic acid production by Streptomyces clavuligerus with peanut derivatives

Clavulanic acid (CA) production by Streptomyces clavuligerus was compared in fermentation media containing various nitrogen sources, including soybean meal, peptone, corn steep powder, cotton seed meal, ammonium sulfate, and ammonium nitrate, respectively. The basal fermentation medium was also supplemented with various concentrations of peanut derivatives. The highest CA concentration was obtained in medium containing soybean meal supplemented with peanut protein. The production of N2-(2-carboxyethyl)-L-arginine (CEA) by a wild-type and a mutant strain of Streptomyces clavuligerus [in which the gene encoding β-lactam synthetase (orf3 ) was inactivated] in fermentation broth was compared. The assay of CA and CEA in the peanut protein-containing fermentation broth of the orf3 blocked mutant of S. clavuligerus and the analysis of amino acids and fatty acids of peanut seed flour showed that peanut protein can provide the aspartate amino acids family, arginine, and glycerol for the production of CA. Based on these results, we conclude that peanut seed flour and its derivatives can be used for CA production on an industrial scale as a supplementary nitrogen and carbon source.

Identification and characterization of Lactobacillus florum strains isolated from South African grape and wine samples

A total of 213 strains of lactic acid bacteria were examined in this study. Among these, 30 strains previously isolated from South African grape and wine samples remained unidentified. The identification of these isolates was performed by BLAST and phylogenetic analyses of 16S rDNA gene sequences, which indicated that the isolates belonged to Lactobacillus florum. In this work, we also designed a discriminative species-specific primer FLOR targeting the 16S rDNA gene of Lb. florum. The validity and specificity of this primer was confirmed. Of particular interest in this study was to further evaluate the identified strains for the presence of genes encoding enzymes of oenological relevance. Reference strains included three flower-associated Lb. florum (F9-1(T), F9-2 and F17) and two Lactobacillus lindneri (AWRI B530 and DSM 20691) strains. Lb. lindneri strains were incorporated as being the closest relatives of Lb. florum. PCR detection results revealed that all Lb. florum strains and Lb. lindneri AWRI B530 (grape isolate) possessed the majority of the tested genes relative to DSM 20691 (beer isolate); these enzyme-encoding genes included malolactic enzyme, peptidases (PepC, PepI, PepN), citrate lyase (α- and β-subunits), phenolic acid decarboxylase and arginine deiminase pathway enzymes (arginine deiminase and ornithine transcarbamylase). Sequence verification of PCR-generated fragments was performed by sequencing. The sequence data were used to construct the phylogenetic trees, which indicated that our Lb. florum isolates cluster with other Lb. florum strains of flower origin but rather distinct from other LAB species, with Lb. lindneri being the next closest species.

Highly Selective Extraction and Transport through a Bulk Liquid Membrane of L-Cysteine Using [K • DC18C6]+ Complexes

It is shown that dicyclohexyl-18-crown-6 (DC18C6) dissolved in chloroform can extract efficiently and selectively the anionic form of L-cysteine from aqueous solutions containing potassium ions. This ability depends on the aqueous phase pH and type of the salt used in this phase. Under optimized conditions L-cysteine can be quantitatively extracted into the organic phase. In such conditions there was no detectable extraction of amino acids tryptophan, tyrosine, leucine, methionine, arginine, asparagine, aspartic acid, glycine, serine, and threonine. The transport of L-cysteine from potassium chloride/potassium hydroxide solution across a bulk liquid membrane containing DC18C6 dissolved in chloroform has been investigated. The parameters influencing the transport efficiency such as the pH of the feed phase, the kind and concentration of the receiving phase, carrier concentration, the kind of organic diluents, and time were examined and optimized. An efficient stripping at the membrane/stripping phase interface was found using hydrochloric acid as strippant. Selectivity of the proposed system was evaluated by performing a series of competitive transport experiments on the binary mixtures containing L-cysteine with the competitor amino acids noted earlier. This investigation exhibits excellent transport selectivity of the process towards L-cysteine with respect to the examined amino acids.

Dynamical behaviour of the human β1-adrenoceptor under agonist binding

The human β1-adrenoceptor (hβ1AR) is a transmembrane (TM) protein responsible for the signal transduction pathway via agonist interaction. Despite its importance, hβ1AR activation mechanism is still unclear. The most studied and widely accepted mechanism is the disruption of a salt bridge between TM3 arginine and TM6 glutamic acid, called ionic lock. In this work, we constructed a functional hβ1AR-model equilibrated in a membrane environment to study the influence of agonist binding on the dynamical behaviour of hβ1AR and on the opening of the ionic lock. The results indicate that the agonist (R-noradrenaline) disturbs the hβ1AR, causing a TM helices rotation, disrupting the ionic lock. This rotational motion occurs in opposite directions in the intercellular and extracellular domains of hβ1AR, opening the ionic lock.