RESUMO
Brain acetylcholinesterase (AChE) forms stable complexes with amyloid-beta peptide (Abeta) during its assembly into filaments, in agreement with its colocalization with the Abeta deposits of Alzheimer's brain. The association of the enzyme with nascent Abeta aggregates occurs as early as after 30 min of incubation. Analysis of the catalytic activity of the AChE incorporated into these complexes shows an anomalous behavior reminiscent of the AChE associated with senile plaques, which includes a resistance to low pH, high substrate concentrations, and lower sensitivity to AChE inhibitors. Furthermore, the toxicity of the AChE-amyloid complexes is higher than that of the Abeta aggregates alone. Thus, in addition to its possible role as a heterogeneous nucleator during amyloid formation, AChE, by forming such stable complexes, may increase the neurotoxicity of Abeta fibrils and thus may determine the selective neuronal loss observed in Alzheimer's brain.
Assuntos
Acetilcolinesterase/química , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/química , Proteínas do Tecido Nervoso/química , Neurônios/patologia , Doença de Alzheimer/metabolismo , Animais , Morte Celular , Células Cultivadas , Embrião de Galinha , Estabilidade Enzimática , Modelos Logísticos , Células PC12 , Ratos , SolubilidadeRESUMO
A monoclonal antibody (mAb) 25B1 directed against fetal bovine-serum acetylcholinesterase (FBS AChE) was used to examine the ability of the cholinergic enzyme to promote the assembly of amyloid-beta peptides (A beta) into Alzheimers fibrils. This mAb binds to the peripheral anionic site of the enzyme and allosterically inhibits catalytic activity of FBS AChE. Several techniques, including thioflavine-T fluorescence, turbidity, and negative-staining at the electron microscopy level, were used to assess amyloid formation. Inhibition of amyloid formation was dependent on the molar ratio AChE:mAb 25B1, and at least 50% of the inhibition of the AChE promoting effect occurs at a molar ratio similar to that required for inhibition of the esterase activity. Our results suggest that mAb 25B1 inhibits the promotion of the amyloid fibril formation triggered by AChE by affecting the lag period of the A beta aggregation process.
Assuntos
Acetilcolinesterase/imunologia , Acetilcolinesterase/metabolismo , Peptídeos beta-Amiloides/biossíntese , Anticorpos Monoclonais/farmacologia , Inibidores da Colinesterase/farmacologia , Peptídeos beta-Amiloides/química , Peptídeos beta-Amiloides/genética , Animais , Sítios de Ligação , Bovinos , Humanos , Cinética , Camundongos , Microscopia Eletrônica , Estrutura Molecular , Fragmentos de Peptídeos/biossíntese , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genética , Mutação PuntualRESUMO
Dengue-2 virus strains from different locations were compared by T1-RNAse-resistant oligonucleotide fingerprinting and antigen signature analysis. The latter technique involved construction of radioimmunoassays using monoclonal antibodies that recognize nine distinct dengue-2 type-specific and flavivirus cross-reactive epitopes over a range of antigen concentrations. A statistical method was used to align unknown dengue antigen concentrations in different strain preparations, allowing comparison of binding profiles. Twenty-six dengue-2 virus strains were separated into five distinct groups (topotypes) on the basis of unique RNA fingerprints. Two of these were represented by New Guinea C, the prototype virus isolated in 1944, and a Philippine strain; others were segregated on the basis of greater than or equal to 80% shared oligonucleotides into similarity groups representing Burma/Thailand (8 strains), Puerto Rico (12 strains), and Jamaica (4 strains). Signature analysis of the prototype and four geographic topotype strains revealed striking antigenic differences. In contrast, a high degree of antigenic similarity was found among strains from the same geographic region. Variation between antigenically distinct strains occurred at both type-specific and group-reactive epitopes, but the widest differences appeared at group-reactive determinants. Signature analysis provides a more rapid and simpler means than RNA fingerprinting of monitoring changes or new introductions of dengue virus populations in a geographic region.