Anne Botev

Freie Universität Berlin, Berlín, Berlin, Germany

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Publications (4)25.27 Total impact

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    ABSTRACT: The amyloid-β (Aβ) peptide is contained within the C-terminal fragment (β-CTF) of the amyloid precursor protein (APP) and is intimately linked to Alzheimer's disease. In vivo, Aβ is generated by sequential cleavage of β-CTF within the γ-secretase module. To investigate γ-secretase function, in vitro assays are in widespread use which require a recombinant β-CTF substrate expressed in bacteria and purified from inclusion bodies, termed C100. So far, little is known about the conformation of C100 under different conditions of purification and refolding. Since C100 dimerization influences the efficiency and specificity of γ-secretase cleavage, it is also of great interest to determine the secondary structure and the oligomeric state of the synthetic substrate as well as the binding properties of small molecules named γ-secretase modulators (GSMs) which we could previously show to modulate APP transmembrane sequence interactions [Richter et al. (2010) Proc. Natl. Acad. Sci. U.S.A. 107, 14597-14602]. Here, we use circular dichroism and continuous-wave electron spin resonance measurements to show that C100 purified in a buffer containing SDS at micelle-forming concentrations adopts a highly stable α-helical conformation, in which it shows little tendency to aggregate or to form higher oligomers than dimers. By surface plasmon resonance analysis and molecular modeling we show that the GSM sulindac sulfide binds to C100 and has a preference for C100 dimers.
    Biochemistry 02/2011; 50(5):828-35. DOI:10.1021/bi1014002 · 3.19 Impact Factor
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    ABSTRACT: The identification of hereditary familial Alzheimer disease (FAD) mutations in the amyloid precursor protein (APP) and presenilin-1 (PS1) corroborated the causative role of amyloid-beta peptides with 42 amino acid residues (Abeta42) in the pathogenesis of AD. Although most FAD mutations are known to increase Abeta42 levels, mutations within the APP GxxxG motif are known to lower Abeta42 levels by attenuating transmembrane sequence dimerization. Here, we show that aberrant Abeta42 levels of FAD mutations can be rescued by GxxxG mutations. The combination of the APP-GxxxG mutation G33A with APP-FAD mutations yielded a constant 60% decrease of Abeta42 levels and a concomitant 3-fold increase of Abeta38 levels compared with the Gly(33) wild-type as determined by ELISA. In the presence of PS1-FAD mutations, the effects of G33A were attenuated, apparently attributable to a different mechanism of PS1-FAD mutants compared with APP-FAD mutants. Our results contribute to a general understanding of the mechanism how APP is processed by the gamma-secretase module and strongly emphasize the potential of the GxxxG motif in the prevention of sporadic AD as well as FAD.
    Journal of Biological Chemistry 05/2010; 285(28):21636-43. DOI:10.1074/jbc.M109.088005 · 4.60 Impact Factor
  • Alzheimer's and Dementia 07/2009; 5(4). DOI:10.1016/j.jalz.2009.05.368 · 17.47 Impact Factor
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    [Show abstract] [Hide abstract]
    ABSTRACT: The identification of hereditary familial Alzheimer disease (FAD) mutations in the amyloid precursor protein (APP) and presenilin-1 (PS1) corroborated the causative role of amyloid-β peptides with 42 amino-acid residues (Aβ42) in the pathogenesis of AD. While most FAD mutations are known to increase Aβ42 levels, mutations within the APP GxxxG motif are known to lower Aβ42 levels by attenuating transmembrane sequence dimerization. Here we show that aberrant Aβ42 levels of FAD mutations can be rescued by GxxxG mutations. The combination of the APP-GxxxG mutation G33A with APP-FAD mutations yielded a constant 60% decrease of Aβ42 levels and a concomitant 3-fold increase of Aβ38 levels compared to G33-wild-type as determined by ELISA. In the presence of PS1-FAD mutations the effects of G33A were attenuated, apparently attributable to a different mechanism of PS1-FAD mutants compared to APP-FAD mutants. Our results contribute to a general understanding of the mechanism how APP is processed by the γ-secretase module and strongly emphasize the potential of the GxxxG motif in the prevention of sporadic AD as well as FAD. APP and APLPs were conventionally thought to exist and to act as monomers. However, biochemical and structural data have accumulated over the last years indicating that APP and APLPs exist as functional dimers or are even present in higher oligomeric units (1-6). Interactions of APP and APLPs were reported to promote cell adhesion in a homo-and heterotypic manner (7, 8). Among other mechanisms the varying strength of APP dimerization mediated through N-terminal sites (5) or by the transmembrane sequence (TMS) (9) has been reported to influence APP processing.