Km Shams Ud Doha

Understanding protein secretion has considerable importance in the biotechnology industry and important implications in a broad range of normal and pathological conditions including development, immunology, and tissue function. While great progress has been made in studying individual proteins in the secretory pathway, measuring and quantifying mechanistic changes in the pathway's activity remains challenging due to the complexity of the biomolecular systems involved. Systems biology has begun… Show more

Understanding protein secretion has considerable importance in the biotechnology industry and important implications in a broad range of normal and pathological conditions including development, immunology, and tissue function. While great progress has been made in studying individual proteins in the secretory pathway, measuring and quantifying mechanistic changes in the pathway's activity remains challenging due to the complexity of the biomolecular systems involved. Systems biology has begun to address this issue with the development of algorithmic tools for analyzing biological pathways; however most of these tools remain accessible only to experts in systems biology with extensive computational experience. Here, we expand upon the user-friendly CellFie tool which quantifies metabolic activity from omic data to include secretory pathway functions, allowing any scientist to infer protein secretion capabilities from omic data. We demonstrate how the secretory expansion of CellFie (secCellFie) can be used to predict metabolic and secretory functions across diverse immune cells, hepatokine secretion in a cell model of NAFLD, and antibody production in Chinese Hamster Ovary cells. Show less

As systems biology approaches to virology have become more tractable, highly studied viruses such as HIV can now be analyzed in new, unbiased ways, including spatial proteomics. We employed here a differential centrifugation protocol to fractionate Jurkat T cells for proteomic analysis by mass spectrometry; these cells contain inducible HIV-1 genomes, enabling us to look for changes in the spatial proteome induced by viral gene expression. Using these proteomics data, we evaluated the merits of… Show more

As systems biology approaches to virology have become more tractable, highly studied viruses such as HIV can now be analyzed in new, unbiased ways, including spatial proteomics. We employed here a differential centrifugation protocol to fractionate Jurkat T cells for proteomic analysis by mass spectrometry; these cells contain inducible HIV-1 genomes, enabling us to look for changes in the spatial proteome induced by viral gene expression. Using these proteomics data, we evaluated the merits of several reported machine learning pipelines for classification of the spatial proteome and identification of protein translocations. From these analyses we found that classifier performance in this system was organelle-dependent, with Bayesian t-augmented Gaussian mixture modeling outperforming support vector machine (SVM) learning for mitochondrial and ER proteins, but underperforming on cytosolic, nuclear, and plasma membrane proteins by QSep analysis. We also observed a generally higher performance for protein translocation identification using a Bayesian model, BANDLE, on row-normalized data. Comparative BANDLE analysis of cells induced to express the wild-type viral genome vs. cells induced to express a genome unable to express the accessory protein Nef identified known Nef-dependent interactors such as TCR signaling components and coatomer complex. Lastly, we found that SVM classification showed higher consistency and was less sensitive to HIV-dependent noise. These findings illustrate important considerations for studies of the spatial proteome following viral infection or viral gene expression and provide a reference for future studies of HIV-gene-dropout viruses. Show less

Despite their therapeutic potential, many protein drugs remain inaccessible to patients since they are difficult to secrete. Each recombinant protein has unique physicochemical properties and requires different machinery for proper folding, assembly, and post-translational modifications (PTMs). Here we aimed to identify the machinery supporting recombinant protein secretion by measuring the protein-protein interaction (PPI) networks of four different recombinant proteins (SERPINA1, SERPINC1… Show more

Despite their therapeutic potential, many protein drugs remain inaccessible to patients since they are difficult to secrete. Each recombinant protein has unique physicochemical properties and requires different machinery for proper folding, assembly, and post-translational modifications (PTMs). Here we aimed to identify the machinery supporting recombinant protein secretion by measuring the protein-protein interaction (PPI) networks of four different recombinant proteins (SERPINA1, SERPINC1, SERPING1 and SeAP) with various PTMs and structural motifs using the proximity-dependent biotin identification (BioID) method. We identified PPIs associated with specific features of the secreted proteins using a Bayesian statistical model, and found proteins involved in protein folding, disulfide bond formation and N-glycosylation were positively correlated with the corresponding features of the four model proteins. Among others, oxidative folding enzymes showed the strongest association with disulfide bond formation, supporting their critical roles in proper folding and maintaining the ER stability. Knock down of ERP44, a measured interactor with the highest fold change, led to the decreased secretion of SERPINC1, which relies on its extensive disulfide bonds. Proximity-dependent labeling successfully identified the transient interactions supporting synthesis of secreted recombinant proteins and refined our understanding of key molecular mechanisms of the secretory pathway during recombinant protein production. Show less

Human milk oligosaccharides (HMOs) afford many health benefits to breast-fed infants, such as protection against infection and regulation of the immune system, through the formation of non-covalent interactions with protein receptors. However, the molecular details of these interactions are poorly understood. Here, we describe the application of catch-and-release electrospray ionization mass spectrometry (CaR-ESI-MS) for screening natural libraries of HMOs against lectins. The HMOs in the… Show more

Human milk oligosaccharides (HMOs) afford many health benefits to breast-fed infants, such as protection against infection and regulation of the immune system, through the formation of non-covalent interactions with protein receptors. However, the molecular details of these interactions are poorly understood. Here, we describe the application of catch-and-release electrospray ionization mass spectrometry (CaR-ESI-MS) for screening natural libraries of HMOs against lectins. The HMOs in the libraries were first identified based on molecular weights (MWs), ion mobility separation arrival times (IMS-ATs) and collision-induced dissociation (CID) fingerprints of their deprotonated anions. The libraries were then screened against lectins and the ligands identified from the MWs, IMS-ATs and CID fingerprints of HMOs released from the lectin in the gas phase. To demonstrate the assay, four fractions, extracted from pooled human milk and containing ≥35 different HMOs, were screened against a C-terminal fragment of human galectin-3 (hGal-3C), for which the HMOs specificities have been previously investigated, and a fragment of the blood group antigen-binding adhesin (BabA) from Helicobacter pylori, for which the HMO specificities have not been previously established. The structures of twenty-one ligands, corresponding to both neutral and acidic HMOs, of hGal-3C were identified; all twenty-one were previously shown to be ligands for this lectin. The presence of HMO ligands at six other MWs was also ascertained. Show less

The intense interest in the mechanisms responsible for the beneficial effects of breast-feeding on infant health has created a significant need for analytical methods capable of rapidly identifying interactions between human milk oligosaccharides (HMOs) and their protein receptors. Currently, there are no established, high-throughput assays for the screening libraries of free (unmodified) HMOs against lectins. The present work describes a rapid and label- and immobilization-free assay, based on… Show more

The intense interest in the mechanisms responsible for the beneficial effects of breast-feeding on infant health has created a significant need for analytical methods capable of rapidly identifying interactions between human milk oligosaccharides (HMOs) and their protein receptors. Currently, there are no established, high-throughput assays for the screening libraries of free (unmodified) HMOs against lectins. The present work describes a rapid and label- and immobilization-free assay, based on catch-and-release electrospray ionization mass spectrometry (CaR-ESI-MS), capable of simultaneously screening mixtures of free HMOs of known concentration for binding to lectins in vitro. Ligand identification relies on the molecular weights (MWs), ion mobility separation arrival times and collision-induced dissociation fingerprints of HMO anions released from the target protein in the gas phase. To establish the reliability of the assay, a library of thirty-one free HMOs, ranging in size from tri- to octasaccharide, was screened against three human galectin (hGal) proteins (a stable mutant of hGal1 (hGal-1), a C-terminal fragment of hGal-3 (hGal-3C) and hGal-7), with known HMO affinities. When implemented using an equimolar concentration library, the CaR-ESI-MS assay identified 100% of ligands with affinities >500 M-1 and ≥93% of all HMO ligands (hGal-1 – 31 of 31 ligands; hGal-3C – 25 of 25; hGal-7 – 28 of 30); no false positives were detected. The assay also successfully identified the majority of the highest affinity HMO ligands (or isomer sets that contain the highest affinity ligands) in the library for each of the three hGal. Notably, for each lectin, CaR-ESI-MS screening required <1 h to complete and consumed <5 ng of each HMO and <0.5 μg of protein. Show less

The affinities of thirty-two free human milk oligosaccharides (HMOs) for four human galectin proteins - a stable mutant of hGal1 (hGal-1), a C-terminal fragment of hGal-3 (hGal-3C), hGal-7 and an N-terminal fragment of hGal-9 (hGal-9N) – were measured using electrospray ionization mass spectrometry (ESI-MS). The binding data show that each of the four galectins recognize the majority of the HMOs tested (hGal-1 binds thirty-two HMOs, hGal-3C binds twenty-six, hGal-7 binds thirty-one and hGal-9N… Show more

The affinities of thirty-two free human milk oligosaccharides (HMOs) for four human galectin proteins - a stable mutant of hGal1 (hGal-1), a C-terminal fragment of hGal-3 (hGal-3C), hGal-7 and an N-terminal fragment of hGal-9 (hGal-9N) – were measured using electrospray ionization mass spectrometry (ESI-MS). The binding data show that each of the four galectins recognize the majority of the HMOs tested (hGal-1 binds thirty-two HMOs, hGal-3C binds twenty-six, hGal-7 binds thirty-one and hGal-9N binds twenty-six). Twenty-five of the HMOs tested bind all four galectins, with affinities ranging from 103 M-1 to 105 M-1. The reliability of the ESI-MS assay for quantifying the affinities of HMOs for lectins was established from the agreement found between the ESI-MS data and affinities of a small number of HMOs for hGal-1, hGal-3C and hGal-7 measured by isothermal titration calorimetry (ITC). Comparison of the relative affinities (of fourteen HMOs) measured by ESI-MS with the reported specificities of hGal-1, hGal-3, hGal-7 and hGal-9 for these same HMOs established from the shotgun human milk glycan microarray (HM-SGM-v2) showed fair-to-poor correlation, with evidence of false positives and false negatives in the microarray data. The results of this study suggest that HMO specificities of lectins established using microarrays may not accurately reflect their true HMO-binding properties and that the use of “in solution” assays such as ESI-MS and ITC is to be preferred. Show less

An electrospray ionization mass spectrometry (ESI-MS) assay for screening carbohydrate libraries against lectins is described. The assay is based on the proxy protein ESI-MS method, which combines direct ESI-MS protein–ligand binding measurements and competitive protein binding, to simultaneously detect and quantify protein–carbohydrate interactions. Specific interactions between components of the library and the target protein (PT) are identified from changes in the relative abundances (as… Show more

An electrospray ionization mass spectrometry (ESI-MS) assay for screening carbohydrate libraries against lectins is described. The assay is based on the proxy protein ESI-MS method, which combines direct ESI-MS protein–ligand binding measurements and competitive protein binding, to simultaneously detect and quantify protein–carbohydrate interactions. Specific interactions between components of the library and the target protein (PT) are identified from changes in the relative abundances (as measured by ESI-MS) of the carbohydrate complexes of a proxyprotein (Pproxy), which binds to all components of the library with known affinity, upon addition of PT to the solution. The magnitude of the change in relative abundance of a given Pproxy–ligand complex provides a quantitative measure of the affinity of the corresponding PT–ligand interaction. A mathematical framework for the implementation of the method in the case of monovalent (single binding site) Pproxy and monovalent and multivalent (multiple equivalent and independent binding sites) PT is described. The application of the method to screen small libraries of oligosaccharides, on the basis of human histo-blood group antigens and milk oligosaccharides, against an N-terminal fragment of the family 51 carbohydrate-binding module, a fucose-binding lectin fromRalstonia solanacearum, and human norovirus VA387 P particle (24-mer of the protruding domain of the capsid protein), serves to demonstrate the reliability and versatility of the assay. Show less

The application of liquid sample desorption electrospray ionization mass spectrometry (liquid
sample DESI-MS) for quantifying protein-carbohydrate interactions in vitro is described.
Association constants for the interactions between lysozyme and β-D-GlcNAc-(1→4)-β-D-
GlcNAc-(1→4)-D-GlcNAc and β-D-GlcNAc-(1→4)-β-D-GlcNAc-(1→4)-β-D-GlcNAc-(1→4)
-D-GlcNAc, and between a single chain antibody and α-D-Galp-(1→2)-[α-D-Abep-(1→3)]-
α-D-Manp-OCH3 and… Show more

The application of liquid sample desorption electrospray ionization mass spectrometry (liquid
sample DESI-MS) for quantifying protein-carbohydrate interactions in vitro is described.
Association constants for the interactions between lysozyme and β-D-GlcNAc-(1→4)-β-D-
GlcNAc-(1→4)-D-GlcNAc and β-D-GlcNAc-(1→4)-β-D-GlcNAc-(1→4)-β-D-GlcNAc-(1→4)
-D-GlcNAc, and between a single chain antibody and α-D-Galp-(1→2)-[α-D-Abep-(1→3)]-
α-D-Manp-OCH3 and β-D-Glcp-(1→2)-[α-D-Abep-(1→3)]-α-D-Manp-OCH3 measured using
liquid sample DESI-MS were found to be in good agreement with values measured by isothermal
titration calorimetry and the direct ESI-MS assay. The reference protein method, which was
originally developed to correct ESI mass spectra for the occurrence of nonspecific ligand-protein
binding, was shown to reliably correct liquid sample DESI mass spectra for nonspecific binding.
The suitability of liquid sample DESI-MS for quantitative binding measurements carried out
using solutions containing high concentrations of the non-volatile biological buffer phosphate
buffered saline (PBS) was also explored. Binding of lysozyme to β-D-GlcNAc-(1→4)-β-D-
GlcNAc-(1→4)-D-GlcNAc in aqueous solutions containing up to 1X PBS was successfully
monitored using liquid sample DESI-MS; with ESI-MS the binding measurements were limited
to concentrations less than ~0.02X PBS. Show less

Gomphostemma parviflorum (Lamiaceae) is a medicinal plant of Bangladesh which has been used traditionally in the treatment of painful and inflammatory conditions such as asthma, headache, fever, etc.

OBJECTIVE: To investigate the anti-nociceptive, anti-inflammatory, CNS depressant and antimicrobial activity of ethanolic extracts of leaves (GPLE) and roots (GPRE) of the plant.

MATERIALS AND METHODS: The anti-nociceptive potentials of the extracts were studied using acetic… Show more

Gomphostemma parviflorum (Lamiaceae) is a medicinal plant of Bangladesh which has been used traditionally in the treatment of painful and inflammatory conditions such as asthma, headache, fever, etc.

OBJECTIVE: To investigate the anti-nociceptive, anti-inflammatory, CNS depressant and antimicrobial activity of ethanolic extracts of leaves (GPLE) and roots (GPRE) of the plant.

MATERIALS AND METHODS: The anti-nociceptive potentials of the extracts were studied using acetic acid-induced writhing test in mice, anti-inflammatory activity was investigated using carrageenan-induced paw edema in rats, CNS depressant activities were evaluated using pentobarbitone-induced sleeping time, Hole cross and Open field tests in mice while the anti-microbial activity was studied by in vitro disc diffusion method.

RESULTS: The extracts GPLE and GPRE significantly (P<0.001) and dose dependently inhibited the acetic acid-induced writhing in mice with 73.15% and 53.69% inhibition, respectively at the dose of 200 mg/kg. At the same dose GPLE and GPRE significantly inhibited carrageenan-induced rats paw edema at the end of 4 hour with 35.54% and 28.17% inhibition, respectively. The extracts significantly prolonged the pentobarbitone-induced sleeping time and decreased the locomotory activities in open field and Hole cross tests in mice. The GPLE showed strong antimicrobial activity against gram Gram-positive and gram Gram-negative bacteria with zones of inhibition ranging from 8- to 20 mm at a concentration of 400 μg/disc.

CONCLUSION: The findings of the study indicate that the leaves and roots of G. parviflorum possess anti-nociceptive, anti-inflammatory and CNS depressant activity and revealed the antimicrobial activities of leaves extract of the plant. The results justify the traditional use of the plant in the treatment of painful and inflammatory disorders. Show less

The present study was designed to evaluate analgesic potential of the methanolic extracts of the leaves of the both Glycosmis pentaphylla (Rutaceae) and Eugenia fruticosa (Myrtaceae). The analgesic activity was evaluated using the acetic acid (0.7% i.p.)-induced writhing inhibition method in swiss albino mice. The methanolic extract of leaves of Glycosmis pentaphylla, at the dose of 200 and 400 mg/kg body weight significantly (p< 0.001) reduced the number of writhes with 31.97% and 44.21% of… Show more

The present study was designed to evaluate analgesic potential of the methanolic extracts of the leaves of the both Glycosmis pentaphylla (Rutaceae) and Eugenia fruticosa (Myrtaceae). The analgesic activity was evaluated using the acetic acid (0.7% i.p.)-induced writhing inhibition method in swiss albino mice. The methanolic extract of leaves of Glycosmis pentaphylla, at the dose of 200 and 400 mg/kg body weight significantly (p< 0.001) reduced the number of writhes with 31.97% and 44.21% of inhibition, respectively compared to control group. The methanolic extracts of leaves of Eugenia fruticosa significantly and dose dependently reduced the pain threshold (p< 0.001) with 51.02 and 72.1% of writhing inhibition when compared to the control group at the dose of 200 and 400 mg/kg body weight, respectively which were comparable to that of the standard drug Diclofenac Na (65.31% inhibition of writhing, p< 0.001). The results of the study demonstrated the potential antinociceptive activity of the methanolic extracts of the leaves of Glycosmis pentaphylla and Eugenia fruticosa which validated the traditional uses of the both plants in painful diseases and further investigations to elucidate the mechanism of action are required.

Keywords: Glycosmis pentaphylla; Eugenia fruticosa, Methanolic extract, Writhing, Acetic acid, Antinociceptive activity. Show less

Successive petroleum ether, ethyl acetate and methanol extracts of the whole plant of Amaranthus spinosus Linn. were investigated for the analgesic activity. Experiments were carried out with these extracts for their peripheral and central antinociceptive potentials on acetic acid induced writhing and radiant heat tail-flick models in mice, respectively. In both the models, methanolic extract showed significant writhing inhibition as well as the elongation of tail-flick time at a dose of 500… Show more

Successive petroleum ether, ethyl acetate and methanol extracts of the whole plant of Amaranthus spinosus Linn. were investigated for the analgesic activity. Experiments were carried out with these extracts for their peripheral and central antinociceptive potentials on acetic acid induced writhing and radiant heat tail-flick models in mice, respectively. In both the models, methanolic extract showed significant writhing inhibition as well as the elongation of tail-flick time at a dose of 500 mg/kg body weight. A linear dose response relationship was also observed. Show less

Plants are playing an important role in therapy of many diseases. Here we have studied on a plant which is Dillenia indica. In this study, ethanolic extract of the leaf and fruit of Dillenia indica was the experimental sample where antidiarrhoeal activity was analyzed. The plant parts showed antidiarrhoeal activity which was measured by castor oil induced method and charcoal plug method. In case of castor oil induced method the percent of inhibition of wet feces was measured and significantly… Show more

Plants are playing an important role in therapy of many diseases. Here we have studied on a plant which is Dillenia indica. In this study, ethanolic extract of the leaf and fruit of Dillenia indica was the experimental sample where antidiarrhoeal activity was analyzed. The plant parts showed antidiarrhoeal activity which was measured by castor oil induced method and charcoal plug method. In case of castor oil induced method the percent of inhibition of wet feces was measured and significantly decreased the total number of feces produced by the administration of castor oil; in 2nd hour (for leaf extract, 6 for 200 mg/kg dose and 4 at dose of 400 mg/kg and for fruit extract, 5.33 for 200 mg/kg dose and 4 at dose of 400 mg/kg) and 3rd hour (for leaf extract,3 for 200 mg/kg dose and 2 at dose of 400 mg/kg and for fruit extract 4 for 200 mg/kg dose and 3 at dose of 400 mg/kg). In charcoal plug method the activity was measured by distance traveled by the charcoal plug. The result was slightly lower than the control and the standard loperamide showed the maximum antidiarrhoeal effect. The average distance traveled by leaf was 28.67 cm and fruit was 22.67 cm which was lower than the control 41 cm where as the standard loperamide showed the average traveled length of 16 cm.
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The aim of the present study was to evaluate and compare the antioxidant, antimicrobial and cytotoxic effects of the methanolic extracts of Glycosmis pentaphylla stems and leaves. Efforts have been given to identify the activities by using highest grade solvent for preparing the extracts. A survey of the antioxidant effect was performed using DPPH method. The methanolic extract of Glycosmis pentaphylla stems showed moderate antioxidant property (103.35 µg/ml) where the leaves showed very little… Show more

The aim of the present study was to evaluate and compare the antioxidant, antimicrobial and cytotoxic effects of the methanolic extracts of Glycosmis pentaphylla stems and leaves. Efforts have been given to identify the activities by using highest grade solvent for preparing the extracts. A survey of the antioxidant effect was performed using DPPH method. The methanolic extract of Glycosmis pentaphylla stems showed moderate antioxidant property (103.35 µg/ml) where the leaves showed very little (337.62 µg/ml). This antioxidant activity may be due to some polyphenolic compounds identified from this plant. Both the extracts showed moderate antimicrobial activity with the highest zone of inhibition for E. coli (23.67±0.76 mm) and Salmonella paratyphi (15.33±0.76 mm) among the tested micro-organisms. This inhibition may legit the traditional implication of the stems of G. pentaphylla as toothbrush. The cytotoxic activity of stems extract of G. pentaphylla was found to be highly potent (5.53 µg/ml, 95% CI, 7.27-4.21) where as the leaves also showed significant activity (47.34 µg/ml, 95% CI, 50.77- 44.15). Several antitumor alkaloids were identified from this plant in the laboratory. The study results also indicate that Glycosmis pentaphylla is a good source of medicinally important compounds.

Keywords: Glycosmis pentaphylla, Antioxidant activity, Antimicrobial activity, Cytotoxic activity, DPPH. Show less

Present study reports CNS depressant and analgesic activities of methanol extract obtained from the leaves of Piper longum L (MEPL). CNS depressant activity was evaluated by using open field and hole cross tests at doses of 250 and 500 mg/kg body weight while peripheral analgesic activity was evaluated by using acetic acid induced writhing method and formalin test respectively in rat model at 100 and 200 mg/kg body weight. The results of the statistical analysis showed that the plant extract… Show more

Present study reports CNS depressant and analgesic activities of methanol extract obtained from the leaves of Piper longum L (MEPL). CNS depressant activity was evaluated by using open field and hole cross tests at doses of 250 and 500 mg/kg body weight while peripheral analgesic activity was evaluated by using acetic acid induced writhing method and formalin test respectively in rat model at 100 and 200 mg/kg body weight. The results of the statistical analysis showed that the plant extract had significant (p<0.01) dose dependent CNS depressant and analgesic activities. Locomotor activity and exploratory behavior of rats in hole cross and open field test were decreased in the test group comparing the control group indicating CNS depressant effect of the extract which was comparable with the standard drug diazepam. The extract also showed better analgesic effects at both doses characterized by reduction in the number of writhes in the acetic acidinduced writhing model and reduction of licking time in the formalin test when compared to the control group. The extract, at the dose of 200 mg/kg, exerted a maximum of 57.58% inhibition of writhing response and 58.8% inhibition was observed for reference drug Indomethacin. So, the present results suggest that the methanol extract of P. longum leaves possesses remarkable CNS depressant and analgesic activities. Show less