RESUMO
Ataxin-3 (AT3) triggers spinocerebellar ataxia type 3 when it carries a polyglutamine stretch expanded beyond a critical threshold. By Fourier transform infrared spectroscopy and atomic force microscopy we previously showed that a normal (AT3Q24) and an expanded (AT3Q55) variant were capable of evolving into oligomers and protofibrils at 37 °C, whereas only the expanded form generated irreversibly aggregated fibrils that also were associated with a network of side-chain glutamine hydrogen bonding [Natalello et al. (2011) PLoS One. 6:e18789]. We report here that AT3Q24, when gradually heated up to 85 °C, undergoes aggregation similar to that observed at 37 °C; in contrast, AT3Q55 only generates large, amorphous aggregates. We propose a possible interpretation of the mechanism by which temperature affects the outcome of fibrillogenesis.
Assuntos
Proteínas Amiloidogênicas/química , Proteínas do Tecido Nervoso/química , Proteínas Nucleares/química , Multimerização Proteica , Proteínas Repressoras/química , Substituição de Aminoácidos , Proteínas Amiloidogênicas/biossíntese , Proteínas Amiloidogênicas/genética , Ataxina-3 , Humanos , Microscopia de Força Atômica , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/genética , Proteínas Nucleares/biossíntese , Proteínas Nucleares/genética , Ligação Proteica , Estrutura Quaternária de Proteína , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Repressoras/biossíntese , Proteínas Repressoras/genética , Espectroscopia de Infravermelho com Transformada de Fourier , TemperaturaRESUMO
We report on DNA arrays produced by dip pen nanolithography (DPN) on a novel Au-Ag micropatterned template stripped surface. DNA arrays have been investigated by atomic force microscopy (AFM) and scanning tunneling microscopy (STM) showing that the patterned template stripped substrate enables easy retrieval of the DPN-functionalized zone with a standard optical microscope permitting multi-instrument and multitechnique local detection and analysis. Moreover the smooth surface of the Au squares ( approximately 5-10 A roughness) allows AFM/STM to be sensitive to the hybridization of the oligonucleotide array with label-free target DNA. Our Au-Ag substrates, combining the retrieving capabilities of the patterned surface with the smoothness of the template stripped technique, are candidates for the investigation of DPN nanostructures and for the development of label-free detection methods for DNA nanoarrays based on the use of scanning probes.
