RESUMO
Protein nanoarrays containing integrin alphavbeta3 or BSA were fabricated on ProLinker-coated Au surface by dip-pen nanolithography (DPN). An atomic force microscope (AFM) tip coated with ProLinker was modified by vitronectin. We measured the interaction force between nanoarrayed integrin alphavbeta3 or BSA and immobilized vitronectin on the cantilever tip by employing tethering-unbinding method. The unbinding force between integrin alphavbeta3 and vitronectin (1087+/-62 pN) was much higher than that of between BSA and vitronectin (643+/-74 pN). These results demonstrate that one can distinguish a specific protein interaction from non-specific interactions by means of force measurement on the molecular interactions between the nanoarrayed protein and its interacting protein on the AFM tip.
Assuntos
Integrina alfaVbeta3/química , Microscopia de Força Atômica/instrumentação , Nanotecnologia/instrumentação , Análise Serial de Proteínas/instrumentação , Mapeamento de Interação de Proteínas/instrumentação , Transdutores , Vitronectina/química , Adsorção , Materiais Revestidos Biocompatíveis/química , Desenho de Equipamento , Análise de Falha de Equipamento , Integrina alfaVbeta3/ultraestrutura , Microscopia de Força Atômica/métodos , Nanotecnologia/métodos , Análise Serial de Proteínas/métodos , Ligação Proteica , Mapeamento de Interação de Proteínas/métodos , Estresse Mecânico , Propriedades de Superfície , Vitronectina/ultraestruturaRESUMO
Protein nanoarrays are addressable ensembles of nano-scale protein domain on solid surfaces. This method can serve as a useful platform for ultraminiaturized bioanalysis. In this study, we investigated single molecular nanopatterning and molecular interaction of proteins that were immobilized on Prolinker surface of gold-coated silicon wafer by using dip-pen nanolithography (DPN) method. Contact force and humidity were optimized at 0.01 nN and 80%, respectively. The domain features of protein nanoarrays were developed at the contact time of 5 s. The optimized conditions for the nanoarray process were applied to create protein nanoarray using integrin alpha(v)beta3 and angiogenin. Constructed protein nanoarrays using integrin alpha(v)beta3 have single molecular monolayer with regular domain shape (height 15 +/- 5 nm). The changed height value due to the single molecular interaction between integrin alpha(v)beta3 and vitronectin was approximately 30 +/- 5 nm on Prolinker surface as measured with atomic force microscopy tip. Taken together, these results suggest that protein nanoarray on Prolinker surface fabricated by well-controlled DPN process can be used to analyze single molecular interaction of protein.
