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1.
Biochem Soc Trans ; 33(Pt 5): 1111-2, 2005 Nov.
Article in English | MEDLINE | ID: mdl-16246057

ABSTRACT

A number of human neurodegenerative diseases involve aggregated amyloid proteins in the brain, e.g. Alzheimer's disease (beta-amyloid) and Parkinson's disease (alpha-synuclein). Other examples are rare familial dementias which involve the BRI gene. In a British family, mutation of the termination codon extends the reading frame of BRI to yield a furin-processed 34-residue peptide (Abri; British dementia peptide), 11 residues longer than the wild-type (WT). In a Danish family, a ten-base insertion also yields a 34-residue peptide (Adan; Danish dementia peptide). To explore the roles of Abri and Adan in neurodegeneration, we synthesized Abri and Adan in oxidized and reduced forms and generated transgenic mice colonies expressing the WT and mutated forms of BRI. We have generated transgenic mice colonies bearing the genes coding for WT-BRI, Adan and Abri under the control of the Thy1 promoter. Whereas WT-BRI transgenic mice express full-length WT-BRI protein in their brains, Adan protein is fully processed to small peptides.


Subject(s)
Amyloid/genetics , Neurotoxins , Adaptor Proteins, Signal Transducing , Alzheimer Disease/genetics , Alzheimer Disease/metabolism , Amino Acid Sequence , Amyloid/chemistry , Brain/metabolism , Humans , Membrane Glycoproteins , Membrane Proteins , Molecular Sequence Data , Parkinson Disease/genetics , Parkinson Disease/metabolism
2.
Biochem Soc Trans ; 32(Pt 1): 33-6, 2004 Feb.
Article in English | MEDLINE | ID: mdl-14748707

ABSTRACT

beta-Amyloid, a neurotoxic peptide deposited in the brains of Alzheimer's disease patients, is released by a series of membrane-limited proteolytic events. beta-Secretase activity is enhanced by cellular targeting into intracellular cholesterol-rich microdomains, which are dispersed by statins.


Subject(s)
Alzheimer Disease/metabolism , Amyloid beta-Peptides/metabolism , Endopeptidases/metabolism , Amyloid Precursor Protein Secretases , Androstenes/pharmacology , Aspartic Acid Endopeptidases , Cell Line , Cholesterol/metabolism , Gene Expression Regulation , Glycosylation , Humans , Hydrogen-Ion Concentration , Protein Processing, Post-Translational/drug effects , Signal Transduction
3.
J Nutr Health Aging ; 7(1): 31-6, 2003.
Article in English | MEDLINE | ID: mdl-12679838

ABSTRACT

The objectives of this article are to summarise evidence that amyloidogenesis is a causal factor in Alzheimer's disease, to outline the main pathways of amyloidogenesis in Alzheimer's disease, describe contemporary evidence showing that the processing of the amyloid precursor protein and amyloidogenesis is strongly influenced by the levels of intracellular cholesterol. Moreover, we shall suggest a mechanistic hypothesis that could explain the observed epidemiological links between the use of inhibitors of cholesterol biosynthesis in patients and the observed reduced risk of Alzheimer's disease.


Subject(s)
Alzheimer Disease/genetics , Alzheimer Disease/metabolism , Amyloid beta-Protein Precursor/genetics , Amyloid beta-Protein Precursor/metabolism , Cholesterol/genetics , Cholesterol/metabolism , Amyloid Precursor Protein Secretases , Aspartic Acid Endopeptidases/metabolism , Cells, Cultured , Electrophoresis, Polyacrylamide Gel , Endopeptidases , Humans , Hydroxymethylglutaryl-CoA Reductase Inhibitors/administration & dosage , Lovastatin/administration & dosage
4.
Biochemistry ; 40(12): 3449-57, 2001 Mar 27.
Article in English | MEDLINE | ID: mdl-11297410

ABSTRACT

Familial British dementia (FBD) is a rare neurodegenerative disorder and shares features with Alzheimer's disease, including amyloid plaque deposits, neurofibrillary tangles, neuronal loss, and progressive dementia. Immunohistochemical and biochemical analysis of plaques and vascular amyloid of FBD brains revealed that a 4 kDa peptide named ABri is the main component of the highly insoluble amyloid deposits. In FBD patients, the ABri peptide is produced as a result of a point mutation in the usual stop codon of the BRI gene. This mutation produces a BRI precursor protein 11 amino acids longer than the wild-type protein. Mutant and wild-type precursor proteins both undergo furin cleavage between residues 243 and 244, producing a peptide of 34 amino acids in the case of ABri and 23 amino acids in the case of the wild-type (WT) peptide. Here we demonstrate that the intramolecular disulfide bond in ABri and the C-terminal extension are required to elongate initially formed dimers to oligomers and fibrils. In contrast, the shorter WT peptide did not aggregate under the same conditions. Conformational analyses indicate that the disulfide bond and the C-terminal extension of ABri are required for the formation of beta-sheet structure. Soluble nonfibrillar ABri oligomers were observed prior to the appearance of mature fibrils. A molecular model of ABri containing three beta-strands, and two beta-hairpins annealed by a disulfide bond, has been constructed, and predicts a hydrophobic surface which is instrumental in promoting oligomerization.


Subject(s)
Amyloid/metabolism , Dementia/metabolism , Disulfides/chemistry , Peptide Fragments/metabolism , Adaptor Proteins, Signal Transducing , Amino Acid Sequence , Amyloid/chemistry , Amyloid/genetics , Amyloid/ultrastructure , Binding Sites/genetics , Circular Dichroism , Coloring Agents , Congo Red/metabolism , Dementia/genetics , Dementia/pathology , Dimerization , Humans , Kinetics , Membrane Glycoproteins , Membrane Proteins , Microscopy, Electron , Models, Molecular , Molecular Sequence Data , Peptide Fragments/chemistry , Peptide Fragments/genetics , Peptide Fragments/ultrastructure , Protein Structure, Secondary/genetics , Solutions , Spectroscopy, Fourier Transform Infrared , United Kingdom
5.
Biochemistry ; 38(51): 16766-76, 1999 Dec 21.
Article in English | MEDLINE | ID: mdl-10606508

ABSTRACT

The multiple roles of voltage-sensitive K(+) channels (Kv1 subfamily) in brain are served by subtypes containing pore-forming alpha (1.1-1.6) and auxiliary beta subunits, usually in an (alpha)(4)(beta)(4) stoichiometry. To facilitate structure/activity analysis, combinations that are prevalent in neurones and susceptible to alpha-dendrotoxin (alphaDTX) were reproduced in mammalian cells, using Semliki Forest virus. Infected Chinese hamster ovary cells expressed N-glycosylated Kv1.1 and 1.2 alpha subunits (M(r) approximately 60 and 62 K) that assembled and bound [(125)I]-alphaDTX with high affinity; an appreciable proportion appeared on the cell surface, with Kv1.2 showing a 5-fold enrichment in a plasma membrane fraction. To obtain 'native-like' alpha/beta complexes, beta1.1 or 2.1 (M(r) approximately 42 and 39 K, respectively) was co-expressed with Kv1.1 or 1.2. This slightly enhanced N-glycosylation and toxin binding, most notable with beta2. 1 and Kv1.2. Solubilization of membranes from cells infected with Kv. 1.2 and beta2.1, followed by Ni(2+) chromatography, gave a purified alpha1.2/beta2.1 complex with a size of approximately 405 K and S(20, W) = 15.8 S. Importantly, these values indicate that four alpha and beta subunits co-assembled as in neurones, a conclusion supported by the size ( approximately 260 K) of the homo-tetramer formed by Kv1.2 alone. Thus, an authentic K(+) channel octomer has been reconstructed; oligomeric species were also found in plasma membranes. To create 'authentic-like' hetero-oligomeric channels, Kv1.1 and 1.2 were co-expressed and shown to have assembled by the precipitation of both with IgGs specific for either. Consistently, confocal microscopy of cells labeled with these antibodies showed that the relatively low surface content of Kv1.1 was increased by Kv1.2. [(125)I]-alphaDTX binding to these complexes was antagonized by DTX(k), a probe selective for Kv1.1, in a manner that mimicks the pattern observed for the Kv1.1/1.2-containing channels in neuronal membranes.


Subject(s)
Neurons/metabolism , Potassium Channels, Voltage-Gated , Potassium Channels/biosynthesis , Potassium Channels/genetics , Semliki forest virus/genetics , Animals , CHO Cells , Cell Membrane/genetics , Cell Membrane/metabolism , Cricetinae , Elapid Venoms/metabolism , Iodine Radioisotopes , Kv1.1 Potassium Channel , Kv1.2 Potassium Channel , Molecular Mimicry , Neurons/chemistry , Plasmids/genetics , Plasmids/metabolism , Potassium Channel Blockers , Potassium Channels/isolation & purification , Potassium Channels/metabolism , Protein Binding/genetics , Recombinant Proteins/antagonists & inhibitors , Recombinant Proteins/biosynthesis , Recombinant Proteins/isolation & purification , Recombinant Proteins/metabolism , Subcellular Fractions/metabolism , Synaptic Membranes/genetics , Synaptic Membranes/metabolism
6.
Pflugers Arch ; 435(1): 43-54, 1997 Dec.
Article in English | MEDLINE | ID: mdl-9359902

ABSTRACT

We have examined the effects of co-expression of Kvbeta1.1 and Kvbeta2.1 subunits on the gating of rat brain Kv1.4 channels, expressed in Xenopus oocytes. Expression of Kv1.4 subunits alone produced a rapidly inactivating "A" type current, which activated at potentials beyond -60 mV in a solution containing high levels of rubidium. Current activation curves obtained from tail current measurements were fitted with a Boltzmann function, with V1/2 = -47 mV and k = 10 mV. Neither the Kvbeta1.1 nor Kvbeta2.1 subunits altered the voltage dependence of activation. Both subunits accelerated the activation time constant of Kv1.4, without affecting its voltage dependence. Surprisingly, the Kvbeta2.1 subunit, which lacks an N-terminal inactivation domain, was almost as effective as the Kvbeta1.1 subunit in speeding up Kv1.4. Steady-state inactivation of Kv1.4 was unchanged upon co-expression with either Kvbeta1.1 or Kvbeta2.1 subunits. Kv1.4 recovered from inactivation with two time constants; apart from an approximately 50% lengthening of the slow time constant with a high Kvbeta2.1 injection ratio, neither time constant was altered by either the Kvbeta1.1 or Kvbeta2.1 subunits, suggesting little interaction with recovery from C-type inactivation. Clearly, beta subunits have the potential to modify the gating of Kv1.4 channels in the brain more subtly than has been suggested previously.


Subject(s)
Ion Channel Gating , Potassium Channels, Voltage-Gated , Potassium Channels/chemistry , Potassium Channels/physiology , Animals , Electric Conductivity , Female , Gene Expression , Kv1.4 Potassium Channel , Membrane Potentials , Oocytes/metabolism , Oocytes/physiology , Potassium Channels/genetics , RNA, Complementary/genetics , Rats , Recombinant Proteins , Xenopus laevis
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