| 1. |
- Wassing, Gabriela M., et al.
(författare)
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DNA Blocks the Lethal Effect of Human Beta-Defensin 2 Against Neisseria meningitidis
- 2021
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Ingår i: Frontiers in Microbiology. - : Frontiers Media SA. - 1664-302X. ; 12
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Tidskriftsartikel (refereegranskat)abstract
- Neisseria meningitidis is a gram-negative bacterium that often asymptomatically colonizes the human nasopharyngeal tract. These bacteria cross the epithelial barrier can cause life-threatening sepsis and/or meningitis. Antimicrobial peptides are one of the first lines of defense against invading bacterial pathogens. Human beta-defensin 2 (hBD2) is an antimicrobial peptide with broad antibacterial activity, although its mechanism of action is poorly understood. Here, we investigated the effect of hBD2 on N. meningitidis. We showed that hBD2 binds to and kills actively growing meningococcal cells. The lethal effect was evident after 2 h incubation with the peptide, which suggests a slow killing mechanism. Further, the membrane integrity was not changed during hBD2 treatment. Incubation with lethal doses of hBD2 decreased the presence of diplococci; the number and size of bacterial microcolonies/aggregates remained constant, indicating that planktonic bacteria may be more susceptible to the peptide. Meningococcal DNA bound hBD2 in mobility shift assays and inhibited the lethal effect of hBD2 in a dose-dependent manner both in suspension and biofilms, supporting the interaction between hBD2 and DNA. Taken together, the ability of meningococcal DNA to bind hBD2 opens the possibility that extracellular DNA due to bacterial lysis may be a means of N. meningitidis to evade immune defenses.
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| 2. |
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| 3. |
- Aronsson, Göran, et al.
(författare)
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Remarkably slow folding of a small protein.
- 1997
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Ingår i: FEBS Letters. - 0014-5793 .- 1873-3468. ; 411:2-3, s. 359-364
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Tidskriftsartikel (refereegranskat)abstract
- Equilibrium denaturation of the 72 amino acid alpha/beta-protein MerP, by acid, guanidine hydrochloride, or temperature, is fully reversible and follows a two-state model in which only the native and unfolded states are populated. A cis-trans equilibrium around a proline peptide bond causes a heterogeneity of the unfolded state and gives rise to a slow- and a fast folding population. With a rate constant of 1.2 s(-1) for the major fast folding population, which has none of the common intrinsically slow steps, MerP is the slowest folding protein of this small size yet reported.
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| 4. |
- Berglund, Anders, et al.
(författare)
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The equilibrium unfolding of MerP characterized by multivariate analysis of 2D NMR data
- 2005
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Ingår i: Journal of magnetic resonance. - San Diego : Academic Press. - 1090-7807 .- 1096-0856. ; 172:1, s. 24-30
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Tidskriftsartikel (refereegranskat)abstract
- A general problem when analysing NMR spectra that reflect variations in the environment of target molecules is that different resonances are affected to various extents. Often a few resonances that display the largest frequency changes are selected as probes to reflect the examined variation, especially in the case, where the NMR spectra contain numerous resonances. Such a selection is dependent on more or less intuitive judgements and relying on the observed spectral variation being primarily caused by changes in the NMR sample. Second, recording changes observed for a few (albeit significant) resonances is inevitably accompanied by not using all available information in the analysis. Likewise, the commonly used chemical shift mapping (CSM) [Biochemistry 39 (2000) 26, Biochemistry 39 (2000) 12595] constitutes a loss of information since the total variation in the data is not retained in the projection into this single variable. Here, we describe a method for subjecting 2D NMR time-domain data to multivariate analysis and illustrate it with an analysis of multiple NNIR experiments recorded at various folding conditions for the protein MerP. The calculated principal components provide an unbiased model of variations in the NNIR spectra and they can consequently be processed as NMR data, and all the changes as reflected in the principal components are thereby made available for visual inspection in one single NMR spectrum. This approach is much less laborious than consideration of large numbers of individual spectra, and it greatly increases the interpretative power of the analysis.
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| 5. |
- Brorsson, Ann-Christin, et al.
(författare)
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GuHCl and NaCl-dependent hydrogen exchange in MerP reveals a well-defined core with an unusual exchange pattern
- 2006
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Ingår i: Journal of Molecular Biology. - : Elsevier BV. - 0022-2836 .- 1089-8638. ; 357:5, s. 1634-1646
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Tidskriftsartikel (refereegranskat)abstract
- We have analysed hydrogen exchange at amide groups to characterise the energy landscape of the 72 amino acid residue protein MerP. From the guanidine hydrochloride (GuHCl) dependence of exchange in the pre-transitional region we have determined free energy values of exchange (ΔGHX) and corresponding m-values for individual amide protons. Detailed analysis of the exchange patterns indicates that for one set of amide protons there is a weak dependence on denaturant, indicating that the exchange is dominated by local fluctuations. For another set of amide protons a linear, but much stronger, denaturant dependence is observed. Notably, the plots of free energy of exchange versus [GuHCl] for 16 amide protons show pronounced upward curvature, and a close inspection of the structure shows that these residues form a well-defined core in the protein. The hydrogen exchange that was measured at various concentrations of NaCl shows an apparent selective stabilisation of this core. Detailed analysis of this exchange pattern indicates that it may originate from selective destabilisation of the unfolded state by guanidinium ions and/or selective stabilisation of the core in the native state by chloride ions. © 2006 Elsevier Ltd. All rights reserved.
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| 6. |
- Brorsson, Ann-Christin, 1969-
(författare)
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The Folding Energy Landscape of MerP
- 2004
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- This thesis is based on studies, described in four papers, in which the folding energy landscape of MerP was investigated by various techniques. MerP is a water-soluble 72 amino acid protein with a secondary structure consisting of four anti-parallel β-strands and two α-helices on one side of the sheet in the order β1α1β2β3α2β4.The first paper describes the use of CD and fluorescence analysis to examine the folding/unfolding process of MerP. From these experiments it was found that the protein folds according to a two-state model in which only the native and unfolded forms are populated without any visible intermediates. With a rate constant of 1.2 s-1, the folding rate was found to be unusually slow for a protein of this size.The studies presented in the second and third papers were based on measurements of native-state amide proton exchange at different temperatures (Paper II) and GuHCl concentrations (Paper III) in the pre-transitional region. In these studies partially unfolded forms were found for MerP which are essentially unrelated to each other. Thus, in the folding energy landscape of MerP, several intermediates seem to occur on different folding trajectories that are parallel to each other. The slow folding rate of MerP might be coupled to extensive visitation of these conformations. Hydrogen exchange in MerP did also reveal structure-dependent differences in compactness between the denatured states in GuHCl and H2O.In the last paper multivariate data analysis was applied to 2-dimensional NMR data to detect conformational changes in the structure of MerP induced by GuHCl. From this analysis it was suggested that regions involved in the most flexible part of the protein structure are disrupted at rather low denaturant concentrations (< 2.1 M GuHCl) while the native structures of the most stable parts are still not completely ruptured at 2.9 M GuHCl.Finally, the stability, kinetics, contact order and folding nuclei of six proteins with similar topology (MerP, U1A, S6, ADA2h, AcP and HPr) were compared. In this analysis it was found that their folding properties are quite diverse, despite their topological similarities, and no general rules that have been formulated yet can adequately predict their folding behaviour.
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| 7. |
- Brorsson, Ann-Christin, et al.
(författare)
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The “Two-State folder” MerP forms partially unfolded structures that show temperature dependent hydrogen exchange
- 2004
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Ingår i: Journal of Molecular Biology. - London : Academic Press. - 0022-2836 .- 1089-8638. ; 340:2, s. 333-344
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Tidskriftsartikel (refereegranskat)abstract
- We have analysed the folding energy landscape of the 72 amino acid protein MerP by monitoring native state hydrogen exchange as a function of temperature in the range of 7-55 degrees C. The temperature dependence of the hydrogen exchange has allowed us to determine DeltaG, DeltaH and DeltaC(p) values for the conformational processes that permit hydrogen exchange. When studied with the traditional probes, fluorescence and CD, MerP appears to behave as a typical two-state protein, but the results from the hydrogen exchange analysis reveal a much more complex energy landscape. Analysis at the individual amino acid level show that exchange is allowed from an ensemble of partially unfolded structures (i.e. intermediates) in which the stabilities at the amino acid level form a broad distribution throughout the protein. The formation of partially unfolded structures might contribute to the unusually slow folding of MerP.
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| 8. |
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| 9. |
- Helmfors, Linda, 1983-
(författare)
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Understanding the dual nature of lysozyme: part villain – part hero : A Drosophila melanogaster model of lysozyme amyloidosis
- 2014
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Amyloid proteins are a distinct class of proteins that can misfold into β-sheet rich structures that later mature to form the characteristic species known as amyloid fibrils, and accumulate in tissues in the human body. The misfolding event is often caused by mutations (or outer factors such as changes in pH) that destabilize the native protein structure. The mature amyloid fibrils were initially believed to be associated with diseases connected to protein misfolding such as Alzheimer’s disease (AD), Parkinson’s disease, transthyretin amyloidosis and lysozyme amyloidosis. However, now it is known that many different factors are involved in these diseases such as failure in protein clearance, lysosomal dysfunction and formation of intermediate misfolded protein species, which possess cytotoxic properties, preceding the formation of mature fibrils.In this thesis the amyloidogenic protein lysozyme has been examined in vivo by using Drosophila melanogaster (fruit fly) as a model organism. The effects of over-expressing human lysozyme and amyloidogenic variants in Drosophila have been investigated both in the absence and presence of the serum amyloid P component (SAP), a protein known to interact with amyloid species. In addition, the role of lysozyme in AD has been investigated by co-expressing human lysozyme and amyloid β in Drosophila.The lysozyme protein is an enzyme naturally found in bodily fluids such as tears, breast milk and saliva. It is engaged in the body’s defense and acts by hydrolyzing the cell wall of invading bacteria. Certain disease-associated point mutations in the gene encoding lysozyme destabilize the protein and cause it to misfold which results in systemic amyloidosis. To investigate the in vivo misfolding behavior of lysozyme we developed and established a Drosophila model of lysozyme amyloidosis. SAP is commonly found attached to amyloid deposits in the body; however, the role of SAP in amyloid diseases is unknown. To investigate the effect of SAP in lysozyme misfolding, these two proteins were co-expressed in Drosophila.The amyloid β peptide is involved in AD, building up the plaques found in AD patient brains. These plaques trigger neuroinflammation and since lysozyme is upregulated during various inflammation conditions, a possible role of lysozyme in AD was investigated by overexpressing lysozyme in a Drosophila model of AD. Interaction between lysozyme and the amyloid β protein was also studied by biophysical measurements.During my work with this thesis, the dual nature of lysozyme emerged; on the one hand a villain, twisted by mutations, causing the lysozyme amyloidosis disease. On the other hand a hero, delaying the toxicity and maybe the neurological damage caused by the amyloid β peptide.
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| 10. |
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