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The β-strand D of transthyretin trapped in two discrete conformations
Umeå University, Faculty of Science and Technology, Umeå Centre for Molecular Pathogenesis (UCMP) (Faculty of Science and Technology). (Sauer-Eriksson)
Umeå University, Faculty of Medicine, Umeå Centre for Molecular Pathogenesis (UCMP) (Faculty of Medicine). (Sauer-Eriksson)
Umeå University, Faculty of Science and Technology, Umeå Centre for Molecular Pathogenesis (UCMP) (Faculty of Science and Technology). (Sauer-Eriksson)
Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). (Lundgren)
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2004 (English)In: Biochimica et Biophysica Acta, ISSN 0006-3002, E-ISSN 1878-2434, Vol. 1700, no 1, 93-104 p.Article in journal (Refereed) Published
Abstract [en]

Conformational changes in native and variant forms of the human plasma protein transthyretin (TTR) induce several types of amyloid diseases. Biochemical and structural studies have mapped the initiation site of amyloid formation onto residues at the outer C and D beta-strands and their connecting loop. In this study, we characterise an engineered variant of transthyretin, Ala108Tyr/Leu110Glu, which is kinetically and thermodynamically more stable than wild-type transthyretin, and as a consequence less amyloidogenic. Crystal structures of the mutant were determined in two space groups, P2(1)2(1)2 and C2, from crystals grown in the same crystallisation set-up. The structures are identical with the exception for residues Leu55-Leu58, situated at beta-strand D and the following DE loop. In particular, residues Leu55-His56 display large shifts in the C2 structure. There the direct hydrogen bonding between beta-strands D and A has been disrupted and is absent, whereas the beta-strand D is present in the P2(1)2(1)2 structure. This difference shows that from a mixture of metastable TTR molecules, only the molecules with an intact beta-strand D are selected for crystal growth in space group P2(1)2(1)2. The packing of TTR molecules in the C2 crystal form and in the previously determined amyloid TTR (ATTR) Leu55Pro crystal structure is close-to-identical. This packing arrangement is therefore not unique in amyloidogenic mutants of TTR.

Place, publisher, year, edition, pages
Amsterdam: Elsevier, 2004. Vol. 1700, no 1, 93-104 p.
Keyword [en]
Crystallography, X-Ray, Dimerization, Humans, Hydrogen Bonding, Hydrogen-Ion Concentration, Models, Molecular, Mutation, Prealbumin, Protein Conformation, Protein Denaturation
National Category
Biochemistry and Molecular Biology Biophysics
Identifiers
URN: urn:nbn:se:umu:diva-13614DOI: 10.1016/j.bbapap.2004.04.004ISI: 000222367400012PubMedID: 15210129OAI: oai:DiVA.org:umu-13614DiVA: diva2:153285
Available from: 2007-10-12 Created: 2007-10-12 Last updated: 2017-01-24Bibliographically approved
In thesis
1. Transthyretin from a structural perspective
Open this publication in new window or tab >>Transthyretin from a structural perspective
2004 (English)Doctoral thesis, comprehensive summary (Other academic)
Alternative title[sv]
Transthyretin ur ett strukturellt perspektiv
Abstract [en]

Conformational changes in human proteins can induce several types of diseases. The nature of the conformational changes is largely unknown, but some lead to amyloid fibril formation. Amyloid fibrils accumulate in the extra-cellular space of tissues resulting in disruption of organ function. Transthyretin (TTR) is a plasma protein involved in three amyloid diseases, familial amyloidotic polyneuropathy, familial amyloidotic cardiomyopathy, and senile systemic amyloidosis. The latter disease involves conformational changes in the wild-type structure of the protein, whereas the others are caused by a gene mutation.

Our goal is to increase the knowledge of why and how some proteins aggregate into amyloid fibrils by solving and analyzing structures of different TTR variants of which some can form amyloid fibrils, whereas others cannot. The crystal structures of wild-type TTR and many of its disease-causing mutants have previously been determined, and observed structural discrepancies between mutant and wild type were claimed to be of importance for amyloid formation. We performed a comparative analysis of all, at that point, known structures of TTR. As a reference for our study, we determined a 1.5 Å resolution structure of human wild-type TTR. We found that the previously reported structural differences between wild type and mutant TTR were insignificant and did not provide clues to the mechanism for amyloid formation.

We showed the double mutant TTR-Ala108Tyr/Leu110Glu to be less amyloidogenic than wild-type transthyretin. Since the structure of few non-amyloidogenic mutants are known, we solved its structure in two space groups, C2 and P21212, where the latter was consistent with most of the structures of transthyretin. Only the highly amyloidogenic mutant ATTR-Leu55Pro has previously been solved in C2. The packing of molecules in our C2 crystal was close-to-identical to the ATTR-Leu55Pro crystal structure, ruling out the described ATTR-Leu55Pro packing interactions as significant for amyloidosis. The C2 structure displayed a large shift in residues Leu55-Leu58, a structural change previously found only in amyloidogenic TTR variants. Combined with previous data, this suggests that transthyretin in solution contains a mixture of molecules with different conformations. This metastability of transthyretin provides insight to why some proteins aggregate into amyloid fibrils.

The natural ligand thyroxine has been shown to stabilize TTR. Small molecules, based on thyroxine, with the potential to serve as inhibitors for amyloid fibril formation are under development. Iodine is a component of thyroxine and we found that TTR also bound free iodide ions. Taking advantage of the anomalous scattering of iodide, we solved the iodide-bound TTR structure using the single-wavelength anomalous dispersion method. In addition, we determined the TTR-chloride structure. Both chloride and iodide stabilized transthyretin where iodide stabilized better. From the thyroxine-TTR structure, three halogen-binding pockets have been identified in each TTR monomer. We found three bound iodides per TTR monomer, two of which were in the thyroxine-binding channel. This indicates that only two of the three halogen-binding pockets in the thyroid-hormone binding channel are optimal for halogen binding. Our results might be useful for the continuing design of small molecule ligands, which in the end can lead to inhibitors for amyloid diseases.

Place, publisher, year, edition, pages
Umeå: Umeå centrum för molekylär patogenes (UCMP) (Teknisk-naturvetenskaplig fakultet), 2004. 50 p.
Keyword
Cell and molecular biology, X-ray crystallography, amyloidosis, structural comparison, anomalous diffraction, Cell- och molekylärbiologi
National Category
Biochemistry and Molecular Biology
Research subject
Molecular Biology
Identifiers
urn:nbn:se:umu:diva-190 (URN)91-7305-593-X (ISBN)
Public defence
2004-02-27, Major Groove, 6L Sjukhusområdet, Umeå Universitet 901 87, Umeå, 10:00
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Available from: 2004-02-05 Created: 2004-02-05 Last updated: 2017-01-24Bibliographically approved

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Olofsson, AndersLundgren, ErikSauer-Eriksson, Elisabeth

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Hörnberg, AndreasOlofsson, AndersEneqvist, ThereseLundgren, ErikSauer-Eriksson, Elisabeth
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Umeå Centre for Molecular Pathogenesis (UCMP) (Faculty of Science and Technology)Umeå Centre for Molecular Pathogenesis (UCMP) (Faculty of Medicine)Department of Molecular Biology (Faculty of Medicine)
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