RESUMO
Apolipoprotein E (apoE), a protein genetically linked to the incidence of Alzheimer's disease, forms SDS-stable complexes in vitro with beta-amyloid peptide (Abeta), the primary component of senile plaques. In the present study, we investigated whether apoE was able to bind full-length Abeta precursor protein (APP). Using a maltose-binding-protein-APP fusion protein and human very-low-density lipoprotein (VLDL), we detected an interaction of apoE with APP that was inhibited by Abeta or anti-apoE antibody. Saturation-binding experiments indicated a single binding equilibrium with an apparent 1:1 stoichiometry and a dissociation constant of 15 nM. An interaction was also observed using apoE from cerebrospinal fluid or delipidated VLDL, as well as recombinant apoE. APP.apoE complexes were SDS-stable, and their formation was not inhibited by reducing conditions; however, they were dissociated by SDS under reducing conditions. ApoE.APP complexes formed high-molecular-mass aggregates, and competition experiments suggested that amino acids 14-23 of Abeta are responsible for complex-formation. Finally, no differences were found when studying the interaction of APP with apoE3 or apoE4. Taken together, our results demonstrate that apoE may form stable complexes with the Abeta moiety of APP with characteristics similar to those of complexes formed with isolated Abeta, and suggest the intriguing possibility that apoE-APP interactions may be pathologically relevant in vivo.
Assuntos
Precursor de Proteína beta-Amiloide/química , Precursor de Proteína beta-Amiloide/metabolismo , Apolipoproteínas E/química , Apolipoproteínas E/metabolismo , Lipoproteínas VLDL/metabolismo , Sequência de Aminoácidos , Precursor de Proteína beta-Amiloide/isolamento & purificação , Apolipoproteínas E/isolamento & purificação , Sítios de Ligação , Proteínas de Transporte , Centrifugação com Gradiente de Concentração , Eletroforese em Gel de Poliacrilamida , Humanos , Cinética , Lipoproteínas VLDL/química , Lipoproteínas VLDL/isolamento & purificação , Proteínas Ligantes de Maltose , Dados de Sequência Molecular , Fragmentos de Peptídeos/química , Ligação Proteica , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/isolamento & purificação , Proteínas Recombinantes de Fusão/metabolismoRESUMO
Resultado de 129 punçöes da vilosidade coriônica para diagnóstico pré-natal no primeiro trimestre de gestaçäo. Os métodos de rotina par preparaçäo direta, preparaçäo de 24 horas e cultura de longa duraçäo, bem como as indicaçöes säo descritos detalhadamente. Tendo em vista a precocidade desse exame e alto sucesso de técnica (97,4%), pode-se concluir pela sua aceitabilidade como método complementar de diagnóstico genético pré-natal