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1.
Melkamu B. Regasa Tesfaye R. Soreta Olu E. Femi Praveen C. Ramamurthy Saravana Kumar 《Journal of molecular recognition : JMR》2020,33(7)
Molecularly imprinted polymer‐modified glassy carbon electrode (GCE)‐based electrochemical sensor is prepared using the electropolymerization of aniline in the presence of melamine (MA) as a template. In this work, the advantages of molecularly imprinted conducting polymers (MICPs) and electroanalytical methods were combined to obtain an electronic device with better performances. The sensor performance was evaluated by cyclic voltammetry (CV) and square wave voltammetry (SWV) with the linear range of 0.6‐16 × 10?9M, quantification limit of 14.9 × 10?10M, and detection limit of 4.47 × 10?10M (S/N = 3). The selectivity of the sensor was tested in the presence of acetoguanamine (AGA), diaminomethylatrazine (DMT), casein, histidine, and glycine interfering molecules taken at the triple concentration with MA that demonstrated too small current response compared with that of the analyte indicating high specificity of the sensor towards the template. The sensor was successfully applied to determine MA in infant formula samples with significant recovery greater than 96% and relative standard deviation (RSD) less than 4.8%. Moreover, the good repeatability, recyclability, and stability make this sensor device promising for the real‐time monitoring of MA in different food stuffs. 相似文献
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Endaye Minbale Atlabachew Minaleshewa Mehari Bewketu Alemayehu Melkamu Mengistu Daniel Ayalew Kerisew Bizuayehu 《Biological trace element research》2020,195(2):669-678
Biological Trace Element Research - Characterization of coffee terroirs is important to determine authenticity and provide confidence for consumers to select the right product. In this regard,... 相似文献
3.
Melkamu G. Woldemariam Ian T. Baldwin Ivan Galis 《The Plant journal : for cell and molecular biology》2012,72(5):758-767
For most plant hormones, biological activity is suppressed by reversible conjugation to sugars, amino acids and other small molecules. In contrast, the conjugation of jasmonic acid (JA) to isoleucine (Ile) is known to enhance the activity of JA. Whereas hydroxylation and carboxylation of JA‐Ile permanently inactivates JA‐Ile‐mediated signaling in plants, the alternative deactivation pathway of JA‐Ile by its direct hydrolysis to JA remains unstudied. We show that Nicotiana attenuata jasmonoyl‐l ‐isoleucine hydrolase 1 (JIH1), a close homologue of previously characterized indoleacetic acid alanine resistant 3 (IAR3) gene in Arabidopsis, hydrolyzes both JA‐Ile and IAA‐Ala in vitro. When the herbivory‐inducible NaJIH1 gene was silenced by RNA interference, JA‐Ile levels increased dramatically after simulated herbivory in irJIH1, compared with wild‐type (WT) plants. When specialist (Manduca sexta) or generalist (Spodoptera littoralis) herbivores fed on irJIH1 plants they gained significantly less mass compared with those feeding on wild‐type (WT) plants. The poor larval performance was strongly correlated with the higher accumulation of several JA‐Ile‐dependent direct defense metabolites in irJIH1 plants. In the field, irJIH1 plants attracted substantially more Geocoris predators to the experimentally attached M. sexta eggs on their leaves, compared with empty vector plants, which correlated with higher herbivory‐elicited emissions of volatiles known to function as indirect defenses. We conclude that NaJIH1 encodes a new homeostatic step in JA metabolism that, together with JA and JA‐Ile‐hydroxylation and carboxylation of JA‐Ile, rapidly attenuates the JA‐Ile burst, allowing plants to tailor the expression of direct and indirect defenses against herbivore attack in nature. 相似文献
4.
Tamene Melkamu Diane Squillace Hirohito Kita Scott M. O’Grady 《The Journal of membrane biology》2009,229(2):101-113
Toll-like receptor (TLR1–6) mRNAs are expressed in normal human bronchial epithelial cells with higher basal levels of TLR3.
TLR2 mRNA and plasma membrane protein expression was enhanced by pretreatment with Poly IC, a synthetic double-stranded RNA
(dsRNA) known to activate TLR3. Poly IC also enhanced mRNA expression of adaptor molecules (MyD88 and TIRAP) and coreceptors
(Dectin-1 and CD14) involved in TLR2 signaling. Additionally, mRNA expression of TLR3 and dsRNA-sensing proteins MDA5 and
RIG-I increased following Poly IC treatment. In contrast, basal mRNA expression of TLR5 and TLR2 coreceptor CD36 was reduced
by 77% and 62%, respectively. ELISA of apical and basolateral solutions from Poly IC-stimulated monolayers revealed significantly
higher levels of IL-6 and GM-CSF compared with the TLR2 ligand PAM3CSK4. Pretreatment with anti-TLR2 blocking antibody inhibited the PAM3CSK4-induced increase in IL-6 secretion after Poly IC exposure. An increase in IL-6 secretion was also observed in cells stimulated
with Alternaria extract after pretreatment with Poly IC. However, IL-6 secretion was not stimulated by zymosan or lipothechoic acid (LTA).
These data demonstrated that upregulation of TLR2 following exposure to dsRNA enhances functional responses of the airway
epithelium to certain (PAM3CSK4), but not all (zymosan, LTA) TLR2 ligands and that this is likely due to differences in coreceptor expression. 相似文献
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6.
Tryptic digestion of proteins continues to be a workhorse of proteomics. Traditional tryptic digestion requires several hours to generate an adequate protein digest. A number of enhanced accelerated digestion protocols have been developed in recent years. Nonetheless, a need still exists for new digestion strategies that meet the demands of proteomics for high-throughput and rapid detection and identification of proteins. We performed an evaluation of direct tryptic digestion of proteins on a MALDI target plate and the potential for integrating RP HPLC separation of protein with on-target tryptic digestion in order to achieve a rapid and effective identification of proteins in complex biological samples. To this end, we used a Tempo HPLC/MALDI target plate deposition hybrid instrument (ABI). The technique was evaluated using a number of soluble and membrane proteins and an MRC5 cell lysate. We demonstrated that direct deposition of proteins on a MALDI target plate after reverse-phase HPLC separation and subsequent tryptic digestion of the proteins on the target followed by MALDI TOF/TOF analysis provided substantial data (intact protein mass, peptide mass and peptide fragment mass) that allowed a rapid and unambiguous identification of proteins. The rapid protein separation and direct deposition of fractions on a MALDI target plate provided by the RP HPLC combined with off-line interfacing with the MALDI MS is a unique platform for rapid protein identification with improved sequence coverage. This simple and robust approach significantly reduces the sample handling and potential loss in large-scale proteomics experiments. This approach allows combination of peptide mass fingerprinting (PMF), MS/MS peptide fragment fingerprinting (PPF) and whole protein MS for both protein identification and structural analysis of proteins. 相似文献
7.
In this work, the effect of Fenton reaction on two elastin cross-linked amino acids, desmosine (DES) and isodesmosine (IDE), in the absence or presence of different wavelength radiations generated from artificial sources has been evaluated using LC/ESI-MS. Irradiation as well as incubation of DES or IDE solutions in the presence of Fe(2+) and H(2)O(2) resulted in products with m/z 497.1 and 481.1 for [M+H](+). A strongly dose-dependent degradation of both amino acids was observed upon exposure to UVB at doses ranging from 0 to 3 J/cm(2) and a moderate dose-dependent degradation upon exposure to UVA at doses 10 times higher than that of UVB. A significant time-dependent degradation of DES and IDE was also observed upon exposure of these amino acids to a lamp emitting visible light similar to sunlight. Exposure of both amino acids to IR radiation (520 W) for 8 h did not cause significant degradation. 相似文献
8.
Tamene Melkamu Hirohito Kita Scott M. O’Grady 《Journal of cell communication and signaling》2013,7(2):109-118
Human bronchial epithelial cells exposed to synthetic double-stranded RNA (poly I:C) exhibited increased IL-6 and RANTES secretion and TLR2 expression that was inhibited following TLR3 silencing. Increased NF-κB and Stat3 phosphorylation were detected after poly I:C exposure and pretreatment with neutralizing antibody targeting IL-6 receptor α (IL-6Rα -nAb) or blocking Jak2 and Stat3 activity inhibited Stat3 phosphorylation. TLR2 up-regulation by poly I:C was also reduced by IL-6Rα-nAb and inhibitors of Jak2, Stat3 and NF-κB phosphorylation, whereas RANTES secretion was unaffected, but abolished following NF-κB inhibition. Treatment with exogenous IL-6 failed to increase TLR2. These findings demonstrate that TLR3 activation differentially regulates TLR expression through autocrine signaling involving IL-6 secretion, IL-6Rα activation and subsequent phosphorylation of Stat3. The results also indicate that NF-κB and Stat3 are required for TLR3-dependent up-regulation of TLR2 and that its delayed expression was due to a requirement for IL-6-dependent Stat3 activation. 相似文献
9.
Here we present a matrix-assisted laser desorption/ionization tandem time-of-flight (MALDI–TOF/TOF)-based label-free relative protein quantification strategy that involves sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE) separation of proteins followed by in-gel trypsin digestion. The main problem encountered in gel-based protein quantification is the difficulty in achieving complete and consistent proteolytic digestion. To solve this problem, we developed a high-pressure-assisted in-gel trypsin digestion method that is based on pressure cycling technology (PCT). The PCT approach performed at least as well as the conventional overnight in-gel trypsin digestion approach in parameters such as number of peaks detected, number of peptides identified, and sequence coverage, and the digestion time was reduced to 45 min. The gel/mass spectrometry (MS)-based label-free protein quantification method presented in this work proved the applicability of the signal response factor concept for relative protein quantification previously demonstrated by other groups using the liquid chromatography (LC)/MS platform. By normalizing the average signal intensities of the three most intense peptides of each protein with the average intensities of spiked synthetic catalase tryptic peptides, which we used as an internal standard, we observed spot-to-spot and lane-to-lane coefficients of variation of less than 10 and 20%, respectively. We also demonstrated that the method can be used for determining the relative quantities of proteins comigrating during electrophoretic separation. 相似文献
10.
The glucosinolate breakdown product indole‐3‐carbinol acts as an auxin antagonist in roots of Arabidopsis thaliana
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Ella Katz Sophia Nisani Brijesh S. Yadav Melkamu G. Woldemariam Ben Shai Uri Obolski Marcelo Ehrlich Eilon Shani Georg Jander Daniel A. Chamovitz 《The Plant journal : for cell and molecular biology》2015,82(4):547-555
The glucosinolate breakdown product indole‐3‐carbinol functions in cruciferous vegetables as a protective agent against foraging insects. While the toxic and deterrent effects of glucosinolate breakdown on herbivores and pathogens have been studied extensively, the secondary responses that are induced in the plant by indole‐3‐carbinol remain relatively uninvestigated. Here we examined the hypothesis that indole‐3‐carbinol plays a role in influencing plant growth and development by manipulating auxin signaling. We show that indole‐3‐carbinol rapidly and reversibly inhibits root elongation in a dose‐dependent manner, and that this inhibition is accompanied by a loss of auxin activity in the root meristem. A direct interaction between indole‐3‐carbinol and the auxin perception machinery was suggested, as application of indole‐3‐carbinol rescues auxin‐induced root phenotypes. In vitro and yeast‐based protein interaction studies showed that indole‐3‐carbinol perturbs the auxin‐dependent interaction of Transport Inhibitor Response (TIR1) with auxin/3‐indoleacetic acid (Aux/IAAs) proteins, further supporting the possibility that indole‐3‐carbinol acts as an auxin antagonist. The results indicate that chemicals whose production is induced by herbivory, such as indole‐3‐carbinol, function not only to repel herbivores, but also as signaling molecules that directly compete with auxin to fine tune plant growth and development. 相似文献