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Protein oligomerization induced by oleic acid at the solidliquid interface: equine lysozyme cytotoxic complexes
Umeå University, Faculty of Medicine, Department of Medical Biochemistry and Biophysics.
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2009 (English)In: The FEBS Journal, ISSN 1742-464X, E-ISSN 1742-4658, Vol. 276, no 15, 3975-3989 p.Article in journal (Refereed) Published
Abstract [en]

Protein oligomeric complexes have emerged as a major target of current research because of their key role in aggregation processes in living systems and in vitro. Hydrophobic and charged surfaces may favour the self-assembly process by recruiting proteins and modifying their interactions. We found that equine lysozyme assembles into multimeric complexes with oleic acid (ELOA) at the solid–liquid interface within an ion-exchange chromatography column preconditioned with oleic acid. The properties of ELOA were characterized using NMR, spectroscopic methods and atomic force microscopy, and showed similarity with both amyloid oligomers and the complexes with oleic acid and its structural homologous protein α-lactalbumin, known as humanα-lactalbumin made lethal for tumour cells (HAMLET). As determined by NMR diffusion measurements, ELOA may consist of 4–30 lysozyme molecules. Each lysozyme molecule is able to bind 11–48 oleic acids in various preparations. Equine lysozyme acquired a partially unfolded conformation in ELOA, as evident from its ability to bind hydrophobic dye 8-anilinonaphthalene-1-sulfonate. CD and NMR spectra. Similar to amyloid oligomers, ELOA also interacts with thioflavin-T dye, shows a spherical morphology, assembles into ring-shaped structures, as monitored by atomic force microscopy, and exerts a toxic effect in cells. Studies of well-populated ELOA shed light on the nature of the amyloid oligomers and HAMLET complexes, suggesting that they constitute one large family of cytotoxic proteinaceous species. The hydrophobic surfaces can be used profitably to produce complexes with very distinct properties compared to their precursor proteins.

Place, publisher, year, edition, pages
2009. Vol. 276, no 15, 3975-3989 p.
Keyword [en]
amyloid, HAMLET, lysozyme, oleic acid, oligomers
URN: urn:nbn:se:umu:diva-29842DOI: 10.1111/j.1742-4658.2009.07107.xOAI: diva2:278282
Available from: 2009-11-25 Created: 2009-11-25 Last updated: 2011-03-31Bibliographically approved
In thesis
1. Protein complexes: assembly, structure and function
Open this publication in new window or tab >>Protein complexes: assembly, structure and function
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

 Most proteins must fold into their native conformations to fulfil their biological functions. Failure of proteins to fold leads to cell pathology and a broad range of human diseases referred to as protein misfolding disease, e.g., Alzheimer’s disease, Parkinson’s disease, and type II diabetes. More than 40 proteins are known to be connected with misfolding diseases. These proteins share no sequence homology but all assemble into cross-b sheet containing insoluble fibrillar aggregates. Despite the pathological conditions that these proteins can induce, living organisms can take advantage of the inherent ability of these proteins to form such structures and to generate novel and diverse biological function, the functional amyloid.

 This thesis examines different aspects of cross-b sheet containing aggregates. The first paper describes the humoral response to aggregated structures of insulin and the astrocytical biomarker S100B in patients suffering from Parkinson’s disease. We show that the patients have an increased immunreactivity towards insulin and S100B in Parkinson’s disease patients compared to a control group.

 The second part of this work focuses on a functional amyloid. HAMLET (human a-lactalbumin made lethal for tumour cells) is a complex of a-lactalbumin and oleic acid, which kills tumour cells but not healthy differentiated cells. We wish to expand the concept of HAMLET to a structurally related protein and therefore create and characterize a complex of equine lysozyme and oleic acid (Paper II). We chose equine lysozyme because both proteins (equine lysozyme and a-lactalbumin) share common ancestors and are spatially related. The newly designed complex was named ELOA, for equine lysozyme with oleic acid. ELOA represents a functional oligomer due to its multimeric state and its ability to bind amyloid specific dyes. In the third paper, we investigate the interaction of the cytotoxic ELOA with live cells in real time to find a mechanistic model (Paper III).

 It is known that HAMLET is not only tumouricidal but is also toxic towards many bacteria. Therefore in the last part of the thesis, we investigated the effects of ELOA on different bacterial strains and focused on its interplay Streptococcus pneumoniae (Paper IV).

 These studies have added significantly to many aspects of protein folding and misfolding from its involvement in Parkinson’s disease to the newly gained functions and structural aspects of de novo produced ELOA.

Place, publisher, year, edition, pages
Umeå: Umeå university, 2009. 40 p.
Umeå University medical dissertations, ISSN 0346-6612 ; 1318
HAMLET, ELOA, lysozyme, amyloid
Research subject
Medical Biochemistry
urn:nbn:se:umu:diva-29792 (URN)978-91-7264-913-2 (ISBN)
Public defence
2009-12-15, 3A9, KBC Huset, Umeå, 00:00 (English)
Available from: 2009-11-25 Created: 2009-11-23 Last updated: 2009-11-25Bibliographically approved

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Wilhelm, KristinaSchleucher, JürgenMorozova-Roche, Ludmilla
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