| 1. |
- Arnell, Robert, et al.
(författare)
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Biotechnological Approach to the Synthesis of 9α-Hydroxylated Steroids
- 2007
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Ingår i: Preparative Biochemistry & Biotechnology. - : Informa UK Limited. - 1082-6068 .- 1532-2297. ; 37:4, s. 309-321
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Tidskriftsartikel (refereegranskat)abstract
- The steroid 9α-hydroxylase gene has been cloned from Mycobacterium smegmatis into Escherichia coli BL21. Progesterone added to bioreactors was subjected to in vivo transformation into 9α-hydroxyprogesterone. In 7 days, 43.6 mg9α-hydroxyprogesterone was formed from 53.8 mg/L progesterone. The enzyme also has shown evidence of processing 4-androstene-3,17-dione in vivo. An extensive analytical method development, including LLE, HPLC-DAD, MS, andNMR was performed to verify the product and to enable a quantitative analysis. Protocols for analytical and preparative separation have been developed, using binaphtol as internal standard. Both the growth pattern and the bioconversion ratewere unaffected by the presence of binaphtol in the bioreactor. The enzyme was purified by immobilised metal affinity and ion exchange chromatography, resulting in low in vitro activity.
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| 2. |
- Atmeh, Ragheb F., et al.
(författare)
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Isolation of High Density Lipoprotein Subclasses by Electrofiltration and Their Chemical Components
- 2009
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Ingår i: Preparative Biochemistry & Biotechnology. - : Informa UK Limited. - 1082-6068 .- 1532-2297. ; 39:3, s. 248-265
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Tidskriftsartikel (refereegranskat)abstract
- The exact role of high density lipoprotein in atheroprotection is not well understood yet due to its complex nature; it comprises more than ten subclasses that vary in size, composition, and function. Isolation and characterization of these subclasses is an important step for further studies addressing their functions in health and disease. In this work, we present a novel method that is relatively simple and efficient for isolation of high density lipoprotein subclasses. The method depends on fractional filtration of the subclasses through a preformed gel membrane system under the effect of an electric field, where the stepwise isolation of the subclasses depends on differences in their rates of migration in polyacrylamide gel. Using this design, we were able to isolate seven high density lipoprotein subclasses with relative molecular masses of 42,000-50,000; 71,000; 103,000; 124,000; 150,000; 182,000; and 219,000. All the subclasses contained apolipoprotein A-I, phosphatidylcholine, sphingomyelin, free cholesterol, esterified cholesterol, and triacylglycerols. Some fractions of some samples contained the apolipoproteins A-II, C-I, C-II, C-III, and E. A subclass of molecular mass of 106,000 was identified and isolated from a healthy young subject that contained albumin and apoA-I with some free and esterified cholesterol, but with no triacylglycerols. This electrofiltration technique offers a novel tool for isolating pure native high density lipoprotein subclasses in a concentrated form that can be used directly for detailed studies of their physicochemical and physiological properties.
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| 3. |
- Danielsson, Ravi, et al.
(författare)
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Aqueous polymer two-phase systems and their use in fragmentation and separation of biological membranes for the purpose of mapping the membrane structure.
- 2013
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Ingår i: Preparative Biochemistry & Biotechnology. - : Informa UK Limited. - 1532-2297 .- 1082-6068. ; 43:5, s. 512-525
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Tidskriftsartikel (refereegranskat)abstract
- When solutions of two different polymers are mixed, phase separation often occurs even at low concentrations of polymers. One polymer usually collects in one phase and the other polymer in the other phase. When water is used as solvent, two aqueous, immiscible, phases are obtained. The same holds for aqueous mixtures of a salt and a polymer. Such aqueous two-phase systems (ATPS) are very useful for separation of high-molecular-weight biomolecules such as proteins and nucleic acids and also for cells, cell organelles, and membrane vesicles. The phase systems can be made highly selective and they are also mild toward biomolecules and cell particles. In this review we describe how ATPS can be used for fragmentation and separation analyses of biological membranes and how this can be used for mapping of the photosynthetic membrane, the thylakoid, of green leaves.
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| 4. |
- Emek, Sinan C, et al.
(författare)
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A large scale method for preparation of plant thylakoids for use in body weight regulation.
- 2010
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Ingår i: Preparative biochemistry & biotechnology. - : Informa UK Limited. - 1532-2297 .- 1082-6068. ; 40:1, s. 13-27
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Tidskriftsartikel (refereegranskat)abstract
- A method for preparation of thylakoids from plant leaves on a large scale is described. The method involves: 1) disruption of the cells with a blender followed by filtration to remove large cell debris and non disrupted cells. 2) precipitation of the thylakoids by adjusting the pH to the isoelectric point, pH 4.7. 3) a washing step by dilution of the precipitate in water followed by precipitation at the same pH. 4) concentration of the precipitate by freeze- thawing or freeze -drying to get the final product. The product is characterized, with respect to protein composition, by SDS-PAGE and mass-spectroscopy, the content of carotenoids, particularly the xanthophylls violaxanthin, antheraxanthin, and zeaxanthin. The thylakoid preparation has about the same capacity to inhibit pancreatic lipase/colipase activity as thylakoids prepared by standard laboratory methods using sucrose in the medium and centrifugation. In a study with mice, it was found that, when the thylakoids were added to the food over 32 days, they significantly reduced the body weight gain and the percentage body fat. The large scale method described here allows studies on the effect of thylakoids in appetite regulation on experimental animals in a longer lasting time and also on humans.
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| 5. |
- Gronlund, Hugo, et al.
(författare)
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Direct Detection Of Single-Nucleotide Polymorphisms In Bacterial DNA By SNPtrap
- 2011
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Ingår i: Preparative Biochemistry & Biotechnology. - : Informa UK Limited. - 1532-2297 .- 1082-6068. ; 41:2, s. 166-174
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Tidskriftsartikel (refereegranskat)abstract
- A major challenge with single-nucleotide polymorphism (SNP) fingerprinting of bacteria and higher organisms is the combination of genome-wide screenings with the potential of multiplexing and accurate SNP detection. Single-nucleotide extension by the minisequencing principle represents a technology that both is highly accurate and enables multiplexing. A current bottleneck for direct genome analyses by minisequencing, however, is the sensitivity, since minisequencing relies on linear signal amplification. Here, we present SNPtrap, which is a novel approach that combines the specificity and possibility of multiplexing by minisequencing with the sensitivity obtained by logarithmic signal amplification by polymerase chain reaction (PCR). We show a SNPtrap proof of principle in a model system for two polymorphic SNP sites in the Salmonella tetrathionate reductase gene (ttrC).
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| 6. |
- Karim, Zoheb, et al.
(författare)
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Immobilization of horseradish peroxidase on β-cyclodextrin-capped silver nanoparticles : Its future aspects in biosensor application
- 2016
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Ingår i: Preparative Biochemistry & Biotechnology. - : Informa UK Limited. - 1082-6068 .- 1532-2297. ; 46:4, s. 321-327
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Tidskriftsartikel (refereegranskat)abstract
- This study aimed to work out a simple and high-yield procedure for the immobilization of horseradish peroxidase on silver nanoparticle. Ultraviolet–visible (UV-vis) and Fourier-transform infrared spectroscopy and transmission electron microscopy were used to characterize silver nanoparticles. Horseradish peroxidase was immobilized on β-cyclodextrin-capped silver nanoparticles via glutaraldehyde cross-linking. Single-cell gel electrophoresis (Comet assay) was also performed to confirm the genotoxicity of silver nanoparticles. To decrease toxicity, silver nanoparticles were capped with β-cyclodextrin. A comparative stability study of soluble and immobilized enzyme preparations was investigated against pH, temperature, and chaotropic agent, urea. The results showed that the cross-linked peroxidase was significantly more stable as compared to the soluble counterpart. The immobilized enzyme exhibited stable enzyme activities after repeated uses.
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| 7. |
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| 8. |
- Karlsson, K, et al.
(författare)
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Purification and characterization of substance P endopeptidase activities in the rat spinal cord
- 1997
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Ingår i: PREPARATIVE BIOCHEMISTRY & BIOTECHNOLOGY. - 1082-6068. ; 27:1, s. 59-78
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Tidskriftsartikel (övrigt vetenskapligt/konstnärligt)abstract
- Two enzymes with substance P degrading activity were purified from the membrane bound fraction of the rat spinal cord. The purified enzymes were characterized with regard to biochemical and kinetic properties. One of the enzymes exhibited close Similarity
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| 9. |
- Persson, R, et al.
(författare)
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dUTPase from Escherichia coli; high-level expression and one-step purification
- 2002
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Ingår i: Preparative Biochemistry & Biotechnology. - 1532-2297. ; 32:2, s. 157-172
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Tidskriftsartikel (refereegranskat)abstract
- The dut gene, which encodes Eseherichia coli deoxyuridine 5'-triphosphate nucleotidohydrolase (dUTPase), has been recloned to increase overexpression of the enzyme and to enable simplification of the purification protocol into a one-step procedure. The gene was cloned into the vector pET-3a and expressed in E. coli BL21(DE3) pLysS under the control of a bacteriophage T7 promotor. Induction results in production of dUTPase corresponding to 60% of the extracted protein. Phosphocellulose chromatography at low pH was utilised for one-step purification, resulting in a homogenous preparation of the recombinant protein with a highly specific activity. The yield of purified enzyme is 500mg per litre of bacterial culture, a significant increase compared to previously employed methods.
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| 10. |
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| 11. |
- Ekholm, Elin, 1981-, et al.
(författare)
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Coping with provoked vestibulodynia in a relational context : A cluster analysis of coping patterns and their associations with relational cognitions and goals
- 2024
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Ingår i: European Journal of Pain. - : John Wiley & Sons. - 1090-3801 .- 1532-2149.
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Provoked vestibulodynia (PVD) is a common pain condition, negatively impacting the relationships and sexual lives of sufferers. Women's coping behaviour has been associated with psychosexual outcomes, yet coping patterns in clinical PVD samples are unexplored, and it is not known how women's coping relates to their relational context.METHOD: Women (N = 128) with PVD answered questionnaires about psychosexual function, pain coping (avoidance and endurance), relational- and pain catastrophizing, sexual goals and perceived partner responses. Cluster analysis was used to explore coping patterns; clusters were validated on measures of pain and psychosexual function and compared on catastrophizing, sexual goals and partner responses using multivariate analyses of variance.RESULTS: The analysis yielded four clusters: endurance; combined high avoidance and endurance; avoidance; and combined low. The group with high levels of both avoidance and endurance coping displayed the worst psychosexual outcomes and high levels of pain- and relational catastrophizing, approach and avoidance goals and perceived negative partner responses.CONCLUSION: There are distinct patterns of coping among women with PVD, and these coping patterns are associated with psychosexual outcomes and relational cognitions and goals, and perceived partner responses. Women who alternate between avoidance and endurance are more distressed and report worse psychosexual functioning. This pattern needs to be identified and addressed in the treatment of PVD.SIGNIFICANCE STATEMENT: This study extends previous findings on vulvar pain coping patterns to a clinical population of women with PVD. It is further the first study to address the relationship between relational variables, such as partner responses and relational catastrophizing and different coping patterns. Thus, the contribution of this study is the contextualizing of coping patterns among women with PVD. The results showed that a combined pattern of avoidance and endurance coping is associated with high distress, poor psychosexual outcomes, and indications of insufficient relational coping, highlighting the need for clinical assessment and intervention to target both women's individual coping patterns and their relational context.
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| 12. |
- Ekholm, Elin, 1981-, et al.
(författare)
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Coping with provoked vestibulodynia in a relational context-A cluster analysis of coping patterns and their associations with relational cognitions and goals
- 2024
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Ingår i: European Journal of Pain. - : John Wiley & Sons. - 1090-3801 .- 1532-2149.
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Tidskriftsartikel (refereegranskat)abstract
- BackgroundProvoked vestibulodynia (PVD) is a common pain condition, negatively impacting the relationships and sexual lives of sufferers. Women's coping behaviour has been associated with psychosexual outcomes, yet coping patterns in clinical PVD samples are unexplored, and it is not known how women's coping relates to their relational context.MethodWomen (N = 128) with PVD answered questionnaires about psychosexual function, pain coping (avoidance and endurance), relational- and pain catastrophizing, sexual goals and perceived partner responses. Cluster analysis was used to explore coping patterns; clusters were validated on measures of pain and psychosexual function and compared on catastrophizing, sexual goals and partner responses using multivariate analyses of variance.ResultsThe analysis yielded four clusters: endurance; combined high avoidance and endurance; avoidance; and combined low. The group with high levels of both avoidance and endurance coping displayed the worst psychosexual outcomes and high levels of pain- and relational catastrophizing, approach and avoidance goals and perceived negative partner responses.ConclusionThere are distinct patterns of coping among women with PVD, and these coping patterns are associated with psychosexual outcomes and relational cognitions and goals, and perceived partner responses. Women who alternate between avoidance and endurance are more distressed and report worse psychosexual functioning. This pattern needs to be identified and addressed in the treatment of PVD.Significance StatementThis study extends previous findings on vulvar pain coping patterns to a clinical population of women with PVD. It is further the first study to address the relationship between relational variables, such as partner responses and relational catastrophizing and different coping patterns. Thus, the contribution of this study is the contextualizing of coping patterns among women with PVD. The results showed that a combined pattern of avoidance and endurance coping is associated with high distress, poor psychosexual outcomes, and indications of insufficient relational coping, highlighting the need for clinical assessment and intervention to target both women's individual coping patterns and their relational context.
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