| 1. |
- Lundberg, Peter, et al.
(författare)
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Permeabilization of Plant Cells: 31P NMR Studies of the Permeability of the Tonoplast
- 1986
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Ingår i: Plant Cell Reports. - : Springer. - 0721-7714 .- 1432-203X. ; 5, s. 13-16
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Tidskriftsartikel (refereegranskat)abstract
- A suspension culture of Catharanthus roseus has been used to study the permeability of cell membranes after treatment with various concentrations of a permeabilizing agent (DMSO). The uptake and release (after permeabilization) of inorganic phosphate (Pi) by cells have been investigated by 32P radiotracer and non-invasive phosphorus-31 NMR experiments. These studies have demonstrated that measurements of the Pi-efflux from plant cells provide a reliable measure of the permeability of the tonoplast.
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| 2. |
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| 3. |
- Nilsson, Kjell, et al.
(författare)
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A General Method for the Immobilization of Cells with Preserved Viability
- 1983
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Ingår i: Applied Microbiology and Biotechnology. ; 17, s. 319-326
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Tidskriftsartikel (refereegranskat)abstract
- Microbial, algal, plant and animal cells have been immobilized, with preserved viability, by entrapment in various matrices according to a new bead polymerization technique. The cell polymer/monomer mixture is kept suspended in a hydrophobic phase such as soy, paraffin, or silicon oil, tri-n-butylphosphate, or dibutyphtalate, which is compatible with the cells. The various monomers or polymers tested include agarose, agar, carrageenan, alginate, fibrin, and polyacrylamide. Furthermore, by adjustment of the stirring speed of the suspension, beads of desired diameter can easily be obtained. The entrapped cells are fully viable and biosynthetically active.
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| 4. |
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| 5. |
- Guo, Ming, et al.
(författare)
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The Difference of Serum Protein Transport between Echinosides and Verbascoside
- 2020
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Ingår i: Journal of the Chemical Society of Pakistan. - : The chemical Society of Pakistan. - 0253-5106. ; 42:3, s. 369-382
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Tidskriftsartikel (refereegranskat)abstract
- Verbascoside (VER) is the enzymatic hydrolysis product of echinacoside (ECH). The molecular structures of ECH and VER have different glucosyl groups so they bind to serum albumin in different ways, resulting in different pharmacological actions. In this report, we have examined the binding characteristics between human serum albumin (HSA) and ECH/VER by molecular modeling and spectroscopic approaches. Molecular modeling revealed that VER bound to HSA mainly through hydrogen bonds, van der Waals forces and hydrophobic forces. The spectroscopic results showed that the interactions between HSA and VER/ECH involved a static binding process, and the bonding strength of the VER-HSA complex was stronger than that of the ECH-HSA complex. The value of the binding distances (r) was low, which indicated the occurrence of energy transfer. The reaction conformational pattern of HSA-VER and HSA-ECH gave a "two-state model" based on fluorescent phase diagram analysis. According to the thermodynamic model, the main forces between interaction of VER and HSA were hydrogen bonds and van der Waals forces, whereas the interaction between ECH and HSA was hydrophobic force. The fluorescence polarization analysis demonstrated that the interaction between HSA and VER or ECH generated a non-covalent complex. Compared with ECH, VER was more likely to bind with HSA because of its smaller molecular size and low polarity. The results of the spectral analysis concurred with the molecular modeling data, which provides a helpful reference for the study of the molecular reaction mechanism of VER/ECH binding to HSA.
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| 6. |
- Muthusamy, Saraladevi, et al.
(författare)
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Heterologous Production of Cyprosin B inNicotiana benthamiana: Unveiling the Role of the Plant-Specific Insert Domain in Protein Function and Subcellular Localization
- 2024
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Ingår i: bioRxiv. - Cold Spring Harbor : Cold Spring Harbor Laboratory Press (CSHL). - 2692-8205.
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- The aqueous extract of Cynara cardunculus flowers is traditionally used in cheese production across Mediterranean countries. To meet the growing industrial demand for plant-based milk-clotting enzymes and to explore potential biotechnological applications, we initiated a study to heterologously produce cyprosin B (CYPB), a key milk-clotting enzyme from C. cardunculus, in Nicotiana benthamiana. We also investigated the role of its plant-specific insert (PSI) domain in the CYPB’s activity and its localization. In this study, full-length CYPB and a PSI domain deleted CYPB (CYPBΔPSI) were transiently expressed in N. benthamiana leaves using Agrobacterium-mediated infiltration. The leaves were harvested nine days post-infiltration, and proteins were purified, yielding approximately 81 mg/kg (CYPB) and 60 mg/kg (CYPBΔPSI) fresh weight. CYPBΔPSI showed significantly higher proteolytic activity (156.72 IU/mg) than CYPB (57.2 IU/mg), indicating that the PSI domain is not essential for enzymatic activity and that its removal results in enhanced enzymatic efficiency. In the milk-clotting activity assay, CYPBΔPSI demonstrated a significantly faster clotting time than full-length CYPB, indicating enhanced milk-clotting efficiency for CYPBΔPSI. Subcellular localization studies revealed that CYPB and PSI were localized in the vacuole and endocytic vesicles. In contrast, CYPBΔPSI was primarily localized in the endoplasmic reticulum (ER) and the tonoplast, suggesting that the PSI domain is critical for vacuolar targeting and membrane permeabilization that affects overall protein yield. This study demonstrates the feasibility of using N. benthamiana as a platform for the scalable production of more efficient recombinant CYPB. It highlights the multifunctional role of the PSI domain in vacuolar sorting without impairing its functionality. These results underscore the potential of plant-based expression systems as a viable alternative for the industrial production of plant milk-clotting enzymes, with significant implications for sustainable cheese production.
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| 7. |
- Guo, Ming, et al.
(författare)
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Comparison of the interaction between lactoferrin and isomeric drugs
- 2017
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Ingår i: Spectrochimica Acta Part A - Molecular and Biomolecular Spectroscopy. - : Elsevier. - 1386-1425 .- 1873-3557. ; 173, s. 593-607
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Tidskriftsartikel (refereegranskat)abstract
- The binding properties of pentacyclic triterpenoid isomeric drugs, i.e. ursolic acid (UA) and oleanolic acid (OA), to bovine lactoferrin (BLF) have been studied by molecule modeling, fluorescence spectroscopy, UV-visible absorbance spectroscopy and infrared spectroscopy (IR). Molecular docking, performed to reveal the possible binding mode or mechanism, suggested that hydrophobic interaction and hydrogen bonding play important roles to stabilize the complex. The results of spectroscopic measurements showed that the two isomeric drugs both strongly quenched the intrinsic fluorescence of BLF through a static quenching procedure although some differences between UM and OA binding strength and non-radiation energy transfer occurred within the molecules. The number of binding sites was 3.44 and 3.10 for UA and OA, respectively, and the efficiency of Forster energy transfer provided a distance of 0.77 and 1.21 nm for UA and OA, respectively. The conformation transformation of BLF affected by the drugs conformed to the "all-or-none" pattern. In addition, the changes of the ratios of alpha-helices, beta-sheets and beta-turns of BLF during the process of the interaction were obtained. The results of the experiments in combination with the calculations showed that there are two modes of pentacyclic triterpenoid binding to BLF instead of one binding mode only governed by the principle of the lowest bonding energy.
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| 8. |
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| 9. |
- Bramble, J L, et al.
(författare)
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Calcium and Phosphate Effects on Growth and Alkaloid Production in Coffea arabica: Experimental Results and Mathematical Model.
- 1991
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Ingår i: Biotechnology and Bioengineering. - : Wiley. - 0006-3592 .- 1097-0290. ; 37:9, s. 859-868
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Tidskriftsartikel (refereegranskat)abstract
- Plant, mammalian, and microbial cells are commonly immobilized in calcium alginate gels for the production of valuable secondary metabolites. However, calcium ions are known to inhibit growth in various type of cells, and calcium is an integral part of such gels. Therefore, an investigation was conducted to evaluate the effect of calcium on the growth and alkaloid production of a model cell-line, Coffea arabica, in suspension culture before attempting to immobilize such cells in alginate. A kinetic model was then developed from the results to describe cell growth and alkaloid production and the mechanism by which calcium influences these variables. In addition, it was observed that there was a characteristic relationship between the concentration of calcium in the external medium and the concentration of extracellular and intracellular phosphate. The intracellular phosphate level was, in turn, related to the production of alkaloids. Using these results, a dynamic mathematical model of cell growth and alkaloid production was developed based on the proposed roles of calcium and phosphate. The model showed satisfactory agreement with three sets of experiments at different calcium concentrations. A possible linkage between the calcium and phosphate results is postulated based on the limited solubility of calcium phosphate.
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| 10. |
- Bramble, J L, et al.
(författare)
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Plant Cell Culture using a Novel Bioreactor: The Magnetically Stabilized Fluidized Bed
- 1990
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Ingår i: Biotechnology progress (Print). - : Wiley. - 8756-7938 .- 1520-6033. ; 6:6, s. 452-457
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Tidskriftsartikel (refereegranskat)abstract
- A novel bioreactor using magnetically stabilized fluidized bed (MSFB) technology has been developed that has certain advantages for cultivating cells continuously. In this system, the cells are protected from shear and are constrained to move through the fermenter in lock-step fashion by being immobilized in calcium alginate beads. The MSFB permits good mass transfer, minimizes particle collisions, and allows for the production of cells while maintaining a controlled cell residence time. Details of the experimental system are described. In addition, the experimental performance of an MSFB used to grow plant cells in batch mode is compared to the results obtained in shake flask culture.
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