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1.
Biochim Biophys Acta Biomembr ; 1860(5): 1105-1113, 2018 May.
Article in English | MEDLINE | ID: mdl-29391167

ABSTRACT

The ß-secretase (BACE1) features a unique sulfur rich motif (M462xxxC466xxxM470xxxC474xxxC478) in its transmembrane helix (BACE1-TM) which is characteristic for proteins involved in copper ion storage and transport. While this motif has been shown to promote BACE1-TM trimerization and binding of copper ions in vitro, the structural basis for the interaction of copper ions with the BACE1-TM is still not well understood. Using molecular dynamics (MD) simulations, we show that membrane embedded BACE1-TMs adopt a flexible trimeric structure that binds and conducts copper ions through variable coordination. In coarse-grained (CG) MD simulations, the spontaneous assembly of BACE1-TMs trimers results in a right-handed helix packing arrangement. In subsequent atomistic MD simulations the sulfur rich motif defines characteristic copper ion coordination sites along a constricted partially solvated axial pore. Sliding and tilting of BACE1-TMs along smooth A459xxxA463/464xxA467 surfaces, facilitated by a central P472 induced kink, enables copper ions to alternate between different coordination sites, including the prominent C466 and M470. We shed light into the structural arrangement of BACE1-TM trimers and propose a mechanism for copper ion conduction that might also apply to other proteins involved in metal ion transport.


Subject(s)
Amyloid Precursor Protein Secretases/chemistry , Amyloid Precursor Protein Secretases/metabolism , Aspartic Acid Endopeptidases/chemistry , Aspartic Acid Endopeptidases/metabolism , Copper/metabolism , Amyloid Precursor Protein Secretases/genetics , Aspartic Acid Endopeptidases/genetics , Helix-Loop-Helix Motifs/genetics , Humans , Ion Transport/genetics , Ions/metabolism , Models, Molecular , Molecular Dynamics Simulation , Protein Binding/genetics , Protein Interaction Domains and Motifs/genetics , Protein Multimerization/genetics
2.
Sci Rep ; 5: 15410, 2015 Oct 29.
Article in English | MEDLINE | ID: mdl-26510576

ABSTRACT

The amyloid-ß42 (Aß42) peptide is believed to be the main culprit in the pathogenesis of Alzheimer disease (AD), impairing synaptic function and initiating neuronal degeneration. Soluble Aß42 oligomers are highly toxic and contribute to progressive neuronal dysfunction, loss of synaptic spine density, and affect long-term potentiation (LTP). We have characterized a short, L-amino acid Aß-oligomer Interacting Peptide (AIP) that targets a relatively well-defined population of low-n Aß42 oligomers, rather than simply inhibiting the aggregation of Aß monomers into oligomers. Our data show that AIP diminishes the loss of Aß42-induced synaptic spine density and rescues LTP in organotypic hippocampal slice cultures. Notably, the AIP enantiomer (comprised of D-amino acids) attenuated the rough-eye phenotype in a transgenic Aß42 fly model and significantly improved the function of photoreceptors of these flies in electroretinography tests. Overall, our results indicate that specifically "trapping" low-n oligomers provides a novel strategy for toxic Aß42-oligomer recognition and removal.


Subject(s)
Alzheimer Disease/drug therapy , Amyloid beta-Peptides/antagonists & inhibitors , Oligopeptides/pharmacology , Peptide Fragments/antagonists & inhibitors , Protein Aggregation, Pathological/drug therapy , Synapses/metabolism , Synaptic Transmission/drug effects , Alzheimer Disease/metabolism , Alzheimer Disease/pathology , Amyloid beta-Peptides/metabolism , Animals , Peptide Fragments/metabolism , Protein Aggregation, Pathological/metabolism , Protein Aggregation, Pathological/pathology , Rats , Rats, Wistar , Synapses/pathology
3.
J Biol Chem ; 290(48): 28737-45, 2015 Nov 27.
Article in English | MEDLINE | ID: mdl-26416887

ABSTRACT

Alzheimer disease is the most severe neurodegenerative disease worldwide. In the past years, a plethora of small molecules interfering with amyloid-ß (Aß) aggregation has been reported. However, their mode of interaction with amyloid fibers is not understood. Non-steroidal anti-inflammatory drugs (NSAIDs) are known γ-secretase modulators; they influence Aß populations. It has been suggested that NSAIDs are pleiotrophic and can interact with more than one pathomechanism. Here we present a magic angle spinning solid-state NMR study demonstrating that the NSAID sulindac sulfide interacts specifically with Alzheimer disease Aß fibrils. We find that sulindac sulfide does not induce drastic architectural changes in the fibrillar structure but intercalates between the two ß-strands of the amyloid fibril and binds to hydrophobic cavities, which are found consistently in all analyzed structures. The characteristic Asp(23)-Lys(28) salt bridge is not affected upon interacting with sulindac sulfide. The primary binding site is located in the vicinity of residue Gly(33), a residue involved in Met(35) oxidation. The results presented here will assist the search for pharmacologically active molecules that can potentially be employed as lead structures to guide the design of small molecules for the treatment of Alzheimer disease.


Subject(s)
Alzheimer Disease , Amyloid beta-Peptides/chemistry , Sulindac/analogs & derivatives , Amyloid beta-Peptides/metabolism , Anti-Inflammatory Agents, Non-Steroidal , Humans , Hydrophobic and Hydrophilic Interactions , Magnetic Resonance Spectroscopy , Protein Binding , Protein Structure, Secondary , Sulindac/chemistry , Sulindac/therapeutic use
4.
Biochemistry ; 54(17): 2777-84, 2015 May 05.
Article in English | MEDLINE | ID: mdl-25875527

ABSTRACT

Amyloid-ß (Aß) peptides are likely the molecular cause of neurodegeneration observed in Alzheimer's disease. In the brain, Aß42 and Aß40 are toxic and the most important proteolytic fragments generated through sequential processing of the amyloid precursor protein (APP) by ß- and γ-secretases. Impeding the generation of Aß42 and Aß40 is thus considered as a promising strategy to prevent Alzheimer's disease. We therefore wanted to determine key parameters of the APP transmembrane sequence enabling production of these Aß species. Here we show that the hydrophilicity of amino acid residues G33, T43, and T48 critically determines the generation of Aß42 and Aß40 peptides (amino acid numbering according to Aß nomenclature starting with aspartic acid 1). First, we performed a comprehensive mutational analysis of glycine residue G33 positioned within the N-terminal half of the APP transmembrane sequence by exchanging it against the 19 other amino acids. We found that hydrophilicity of the residue at position 33 positively correlated with Aß42 and Aß40 generation. Second, we analyzed two threonine residues at positions T43 and T48 in the C-terminal half of the APP-transmembrane sequence. Replacement of single threonine residues by hydrophobic valines inversely affected Aß42 and Aß40 generation. We observed that threonine mutants affected the initial γ-secretase cut, which is associated with levels of Aß42 or Aß40. Overall, hydrophilic residues of the APP transmembrane sequence decide on the exact initial γ-cut and the amounts of Aß42 and Aß40.


Subject(s)
Amyloid beta-Peptides/biosynthesis , Amyloid beta-Protein Precursor/metabolism , Amino Acid Sequence , Amyloid beta-Protein Precursor/chemistry , Blotting, Western , Cell Line , Enzyme-Linked Immunosorbent Assay , Humans , Hydrophobic and Hydrophilic Interactions , Molecular Sequence Data
5.
J Am Chem Soc ; 135(51): 19354-61, 2013 Dec 26.
Article in English | MEDLINE | ID: mdl-24304299

ABSTRACT

The ß-secretase or ß-site amyloid precursor protein cleaving enzyme 1 (BACE1) is the enzyme responsible for the formation of amyloid-ß peptides, which have a major role in Alzheimer pathogenesis. BACE1 has a transmembrane sequence (TMS), which makes it unique among related proteases. We noticed that the BACE1 TMS contains an uncommon sulfur-rich motif. The sequence MxxxCxxxMxxxCxMxC spans the entire TMS, resembles metal ion binding motifs, and is highly conserved among homologues. We used a synthetic 31-mer model peptide comprising the TMS to study metal ion binding and oligomerization. Applying diverse biochemical and biophysical techniques, we detected dimer and trimer formation of the TMS peptide with copper ions. Replacement of the central Cys466 by Ala essentially abolished these effects. We show that the peptide undergoes a redox reaction with copper ions resulting in a disulfide bridge involving Cys466. Further, we find peptide trimerization that depends on the presence of monovalent copper ions and the sulfhydryl group of Cys466. We identified Cys466 as a key residue for metal ion chelation and to be the core of an oligomerization motif of the BACE1-TMS peptide. Our results demonstrate a novel metal ion controlled oligomerization of the BACE1 TMS, which could have an enormous therapeutic importance against Alzheimer disease.


Subject(s)
Amyloid Precursor Protein Secretases/chemistry , Copper/analysis , Models, Biological , Sulfur/chemistry , Amino Acid Motifs , Animals , Circular Dichroism , Colorimetry , Humans , Mice , Rats , Sequence Alignment , Spectroscopy, Fourier Transform Infrared
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