Chlamydomonas is a model organism to study photosynthesis. Thylakoid membranes comprise several p... more Chlamydomonas is a model organism to study photosynthesis. Thylakoid membranes comprise several proteins belonging to photosystems I and II. In this chapter, we show the accurate proteomic measurements in thylakoid membranes. The chlorophyll-containing membrane protein complexes were precipitated using chloroform/methanol solution. These complexes were separated using two-dimensional gel electrophoresis, and the resolved spots were exercised from the gel matrix and digested with trypsin. These peptide fragments were separated by MALDI-TOF, and the isotopic masses were blasted to a MASCOT server to obtain the protein sequence. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF). The method discussed here would be a useful method for the separation and identification of thylakoid membrane proteins.
Objective: Assessment of anticancer activity of the plant cell suspension cultures of Ocimum sanc... more Objective: Assessment of anticancer activity of the plant cell suspension cultures of Ocimum sanctum by 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and Trypan blue dye exclusion assay against A549 (human lung cancer cell line). Materials and Methods: In vitro anticancer activity of ethanol, acetone, and aqueous leaf extracts of O. sanctum was evaluated on A549 cancerous cell line by MTT assay and Trypan blue dye exclusion assay. MTT assay is based on the capacity of mitochondrial enzymes of viable cells to reduce the yellow soluble salt MTT to purple-blue insoluble formazan precipitate which is then quantified spectrophotometrically at 570 nm. Trypan blue assay is based on staining of cells. Cells are then counted using hemocytometer under the microscope, non-viable cells were stained blue and viable cells remain unstained. Results: The aqueous leaf extract of O. sanctum has not shown any anticancer activity. However, potent anticancer activity was show...
Journal of Photochemistry and Photobiology B-biology, Jan 1, 2009
Human serum albumin (HSA) is a predominant protein in the blood. Most drugs can bind to HSA and b... more Human serum albumin (HSA) is a predominant protein in the blood. Most drugs can bind to HSA and be transported to target locations of the body. For this study, we have extracted 3-trans-feruloyl maslinic acid (FMA) from the medicinal plant Tetracera asiatica, its a non-fluorescent derivative have potent anti-cancer, anti-HIV, anti-diabetic, and anti-inflammatory activities. The binding constant of the compound with HSA, calculated from fluorescence data, was found as KFMA = 1.42 ± 0.01 × 108 M−1, which corresponds to 10.9 kcal M−1 of free energy. Furthermore, microTOF-Q mass spectrometry data showed binding of FMA at nanomolar concentrations of FMA to free HSA. The study detected a mass increase from 66,560 Da (free HSA) to 67,919 Da (HSA + drug). This indicated a strong binding of FMA to HSA, resulting in an increase of the protein’s absorbance and fluorescence. The secondary structure of HSA + FMA (0.1 mM) complexes showed the protein secondary structure became partially unfolded upon interaction of FMA with HSA, as well as indicating that HSA–FMA complexes were formed. Docking experiments uncovered the binding mode of FMA in HSA molecule. It was found that FMA binds strongly in different places with hydrogen bonding at IB domain of Arg 114, Leu 115 and Asp 173.
Chlamydomonas is a model organism to study photosynthesis. Thylakoid membranes comprise several p... more Chlamydomonas is a model organism to study photosynthesis. Thylakoid membranes comprise several proteins belonging to photosystems I and II. In this chapter, we show the accurate proteomic measurements in thylakoid membranes. The chlorophyll-containing membrane protein complexes were precipitated using chloroform/methanol solution. These complexes were separated using two-dimensional gel electrophoresis, and the resolved spots were exercised from the gel matrix and digested with trypsin. These peptide fragments were separated by MALDI-TOF, and the isotopic masses were blasted to a MASCOT server to obtain the protein sequence. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF). The method discussed here would be a useful method for the separation and identification of thylakoid membrane proteins.
Objective: Assessment of anticancer activity of the plant cell suspension cultures of Ocimum sanc... more Objective: Assessment of anticancer activity of the plant cell suspension cultures of Ocimum sanctum by 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and Trypan blue dye exclusion assay against A549 (human lung cancer cell line). Materials and Methods: In vitro anticancer activity of ethanol, acetone, and aqueous leaf extracts of O. sanctum was evaluated on A549 cancerous cell line by MTT assay and Trypan blue dye exclusion assay. MTT assay is based on the capacity of mitochondrial enzymes of viable cells to reduce the yellow soluble salt MTT to purple-blue insoluble formazan precipitate which is then quantified spectrophotometrically at 570 nm. Trypan blue assay is based on staining of cells. Cells are then counted using hemocytometer under the microscope, non-viable cells were stained blue and viable cells remain unstained. Results: The aqueous leaf extract of O. sanctum has not shown any anticancer activity. However, potent anticancer activity was show...
Journal of Photochemistry and Photobiology B-biology, Jan 1, 2009
Human serum albumin (HSA) is a predominant protein in the blood. Most drugs can bind to HSA and b... more Human serum albumin (HSA) is a predominant protein in the blood. Most drugs can bind to HSA and be transported to target locations of the body. For this study, we have extracted 3-trans-feruloyl maslinic acid (FMA) from the medicinal plant Tetracera asiatica, its a non-fluorescent derivative have potent anti-cancer, anti-HIV, anti-diabetic, and anti-inflammatory activities. The binding constant of the compound with HSA, calculated from fluorescence data, was found as KFMA = 1.42 ± 0.01 × 108 M−1, which corresponds to 10.9 kcal M−1 of free energy. Furthermore, microTOF-Q mass spectrometry data showed binding of FMA at nanomolar concentrations of FMA to free HSA. The study detected a mass increase from 66,560 Da (free HSA) to 67,919 Da (HSA + drug). This indicated a strong binding of FMA to HSA, resulting in an increase of the protein’s absorbance and fluorescence. The secondary structure of HSA + FMA (0.1 mM) complexes showed the protein secondary structure became partially unfolded upon interaction of FMA with HSA, as well as indicating that HSA–FMA complexes were formed. Docking experiments uncovered the binding mode of FMA in HSA molecule. It was found that FMA binds strongly in different places with hydrogen bonding at IB domain of Arg 114, Leu 115 and Asp 173.
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