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- Ahl, Ing-Marie, 1974-, et al.
(author)
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Coexpression of yeast copper chaperone (yCCS) and CuZn-superoxide dismutases in Escherichia coli yields protein with high copper contents
- 2004
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In: Protein Expression and Purification. - : Elsevier BV. - 1046-5928 .- 1096-0279. ; 37:2, s. 311-319
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Journal article (peer-reviewed)abstract
- To fully understand the function of the Cu- and Zn-containing superoxide dismutases in normal and disordered cells, it is essential to study protein variants with full metal contents. We describe the use of an Escherichia coli-based expression system for the overproduction of human intracellular wild type CuZn-superoxide dismutase (SOD), the CuZnSOD variant F50E/G51E (monomeric), two amyotrophic lateral sclerosis-related mutant CuZnSOD variants (D90A and G93A), and PseudoEC-SOD, all with high Cu contents. This system is based on coexpression of the SOD variants with the yeast copper chaperone yCCS during growth in a medium supplemented with Cu2+ and Zn2+. The recombinant SOD enzymes were all found in the cytosol and represented 30-50% of the total bacterial protein. The enzymes were purified to homogeneity and active enzymes were obtained in high yield. The resulting proteins were characterized through immunochemical reactivity and specific activity analyses, in conjunction with mass-, photo-, and atomic absorption-spectroscopy. © 2004 Elsevier Inc. All rights reserved.
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| 5. |
- Ahl, Ing-Marie, 1974-
(author)
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Protein Engineering of Extracellular Superoxide Dismutase : Characterization of Binding to Heparin and Cellular Surfaces
- 2010
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Doctoral thesis (other academic/artistic)abstract
- Accumulating evidence indicates that oxygen free radicals are involved in many diseases and pathological conditions, such as aging, inflammation, reperfusion damage of ischemic tissue and various cardiovascular diseases. Extracellular superoxide dismutase (ECSOD) thus plays a major role in the maintenance of cells by providing protection against these toxic substances in the extracellular space. Various animal studies have shown that ECSOD has the ability to protect against many of these disorders, and interest has therefore evolved in the potential therapeutic use of the enzyme.However, despite strenuous efforts, large-scale production of the enzyme has not been achieved. To overcome this problem, a mimic of the enzyme, PseudoECSOD, has previouslybeen constructed. This chimera is easy to produce in large amounts and has all the structural, enzymatic and heparin-binding characteristics of ECSOD, making it a potential substitute for ECSOD in therapeutic situations. However, the copper content of PseudoECSOD has been shown to be rather low, and since the copper ion is very important for the catalytic function of the enzyme, a production system that utilizes a copper chaperone for proper insertion of copper into the active site of the enzyme was constructed. The results show that the copper content of PseudoECSOD produced by this system is close to 100 %.In order to use PseudoECSOD therapeutically, further investigations of its binding capability and protective properties are needed. Therefore, the binding of ECSOD and PseudoECSOD to heparin was investigated using isothermal titration calorimetry. The results show that although some purely ionic interactions are important for the binding between ECSOD and heparin, there is also a substantial contribution from non-ionic interactions. The investigation also showed that the C-terminal domain is the only part of ECSOD that contributes to productive binding, and that the binding of PseudoECSOD and ECSOD to heparin is similar.In addition, analysis of mutant proteins strongly indicated that the amino acids R210, K211 and R214 are important for optimal binding of ECSOD to heparin, accounting for about 30 % of the total binding energy. The structural placement of these amino acids in an α-helix also confirms the hypothesis postulated by Margalit et al., that a common structural motif for heparin-binding proteins may be two positively charged amino acids at a distance of approximately 20 Å in the 3D-structure, facing opposite directions of a α-helix. The importance of these residues was also confirmed by analysis of a phage display library of the C-terminal domain of ECSOD.The binding of PseudoECSOD to heparan sulfate on cell surfaces of two different cell types, HepG2 and endothelial cells, was also investigated. The results clearly show that PseudoECSOD binds to these cells in a very similar manner to ECSOD. To investigate the protective properties of PseudoECSOD against ischemia-reperfusion injuries, an isolated rabbit heart model was used. The results indicate that the enzyme has a protective effect. However, more experiments using the rabbit heart and other animal models are needed to identify the optimal dose for protective purposes. The protective properties of PseudoECSOD in human tissue should also be thoroughly investigated.In summary, the findings in these studies, together with earlier results showing the close resemblance of PseudoECSOD to ECSOD in structural, enzymatic and heparin-binding properties, further support the proposition that PseudoECSOD may be a good substitute for ECSOD to use in therapeutic interventions.
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- Ahl, Ing-Marie, et al.
(author)
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Thermodynamic Characterization of the Interaction between the C-Terminal Domain of Extracellular Superoxide Dismutase and Heparin by Isothermal Titration Calorimetry
- 2009
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In: BIOCHEMISTRY. - : American Chemical Society (ACS). - 0006-2960 .- 1520-4995. ; 48:41, s. 9932-9940
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Journal article (peer-reviewed)abstract
- Extracellular superoxide dismutase (ECSOD) interacts with heparin through its C-terminal domain. In this study we used isothermal titration calorimetry (ITC) to get detailed thermodynamic information about the interaction. We have shown that the interaction between ECSOD and intestinal mucosal heparin (M-w 6000-30000 Da) is exothermic and driven by enthalpy at physiological salt concentration. However, the contribution from entropy is favorable for binding or small isolated heparin fragments. By studying different size-defined heparin fragments, we also concluded that it hexasaccharide moiety is sufficient for strong binding to ECSOD. The binding involves proton transfer from the buffer to the ECSOD-heparin complex, and the results indicate that the number of ionic interactions made between ECSOD and heparin upon binding varies from three to five for heparin and an octasaccharide fragment, respectively. Surprisingly and despite the many charges found oil both the protein and the polysaccharide, our results indicate that the nonionic contribution to the binding is large. From the temperature dependence we have calculated the constant pressure heat capacity change (Delta C-p) of the interaction to -644 J K-1 mol(-1) and -306 J K-1 mol(-1) for heparin and all octasaccharide, respectively
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| 7. |
- Akner-Kohler, Cheryl, et al.
(author)
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Aesthetics and nanostructure
- 2011
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In: International Innovation. - : Research Media. - 2041-4552. ; , s. 97-99
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Journal article (pop. science, debate, etc.)
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| 8. |
- Akner-Koler, Cheryl, et al.
(author)
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Aesthetics and nanostructure
- 2011
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In: International Innovation. - : Research Media. - 2041-4552. ; :6, s. 97-97
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Journal article (other academic/artistic)abstract
- Professors Cheryl Akner-Kohler and Lena Tibell of the Nanoform project describe the artistic methods and benefits that it might afford to both science and society.
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| 9. |
- Andersson, Johanna, 1972-
(author)
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Barns teckningar som utgångspunkt i det naturvetenskapliga samtalet
- 2019
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Doctoral thesis (other academic/artistic)abstract
- Barns föreställningar utgör en viktig del i deras begreppsbildning inom naturvetenskap vilket betonas inom konstruktivismen. I denna avhandling utgör en socialkonstruktivistisk syn på lärande det teoretiska ramverket. Det övergripande syftet med avhandlingen är att i samtal med barn i åldrarna 4–13 år och med utgångspunkt i deras teckningar, utveckla kunskap om de föreställningar de ger uttryck för inom fyra naturvetenskapliga områden: värme, blandning, människokroppen och vad som är levande/inte levande. Två forskningsfrågor behandlas i avhandlingen: Vilka föreställningar ger barn uttryck för i teckningar och i samtal om naturvetenskapliga fenomen? Vilka metodologiska förutsättningar och utmaningar finns det när det gäller att använda teckningar som utgångspunkt och som meningsskapande redskap för att fånga barns föreställningar? En multimodal metod bestående av teckningar, samtal och barnens aktivitet användes vid datainsamlingen, vilket är i linje med ett socialsemiotiskt perspektiv.Avhandlingen består av fyra studier. Resultatet i de två första studierna visar att barns föreställningar om blandningar var något mer utvecklade än vad som visats i tidigare studier, medan deras föreställningar gällande värme överensstämde med vad som tidigare rapporterats.Den tredje studien visar att barnen känner till fler organ i människokroppen och visar, till skillnad från vad som framkommit i tidigare forskning, förmåga att rita kopplingar mellan organen. I den fjärde studien talar en majoritet av barnen om död som en motsats till liv och några ritade att det som inte lever tidigare har levt. Barn som är medvetna om mikroskopiska objekt klassificerar dem som levande. Förklaringarna visar på en inkonsekvens i barnens resonemang om växter lever eller inte.Metodologiskt framkom att barns föreställningar med fördel fångas genom deras egna teckningar tillsammans med deras förklaringar av dessa. Teckningarna fungerar även som hjälpmedel för att föra fram och delge olika resonemang. I avhandlingen diskuteras barns lösningar på rittekninska problem som de mötte vid sidan av den naturvetenskapliga uppgiften. I studien om människokroppen handlar dessa problem om svårigheten i att överföra den tredimensionella människokroppen till två dimensioner. En annan svårighet var att kroppens organ, skelett, muskler och vävnader ligger ovanpå varandra. Där använde barnen strategin att rita röntgenbilder för att visa det som ligger dolt. Barn i olika åldrar löste ofta dessa rittekniska problem på ett mycket kreativt sätt i kombination med deras muntliga/skriftliga kommentarer. Förutom teckningarnas betydelse diskuteras skillnaden mellan kontextualiserade och dekontextualiserade uppgifter. De olika områdena som studerats i avhandlingens artiklar befinner sig på olika abstraktionsnivåer vilket påverkar barnens föreställningar och hur detta uttrycks in bilder. Naturvetenskap innefattar olika dimensioner där det handlar om att både lära sig strukturell och processuell kunskap. Den multimodala metoden gav barnen hjälp med att samla, strukturera och uttrycka sina tankar
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| 10. |
- Andersson, Johanna, et al.
(author)
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Children's reasoning and representations about living and non-living things
- 2013
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Conference paper (peer-reviewed)abstract
- Understanding of the concept ‘life’ and what characterise ‘living things’ is important as a foundation for learning in biology. In a more general view, this understanding can make children develop awareness, respect and responsibility for life as members of a society and in decision making for sustainable development. The present pilot study aim to investigate 5-6 year old pre-school children’s reasoning and representations about living and nonliving things. In cognitive developmental research, the concept of life is well investigated but, questions still remain regarding how children reason around and represent these concepts. Previous research has found that children have difficulties in including plants as living things. Moreover, it is found that young children include e.g. the sun, clouds and rocks as living things. The methods that have been used are often quantitative and use picture-cards with different objects for the children to categorize. In the present pilot study a modified methodology was applied. Children’s drawings of what they consider as living and non-living were collected and picture-cards were used as point of departure for reasoning. In interviews the children were encouraged to explain and express their ideas. The drawings and the cards mainly worked as a meaning making tool for the children. Results from the study will be presented and discussed.
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