MRS Fall Meeting 2012, Massachusetts, Amerika Birleşik Devletleri, 25 - 30 Kasım 2012, ss.21
Inorganic Binding Peptide Based Patterning of Multifunctional Fusion Protein on Metal Surface
Inorganic binding peptides are of importance in nanobiotechnology since they can be exploited to tailor peptide based hybrid systems and novel materials for practical applications in a wide range of areas including materials science, engineering, environment and medicine. In our group, we have identified and characterized peptides that can bind a variety of inorganic materials specifically and with high affinity using biocombinatorial techniques. As previously reported, we have selected two gold binding peptide (AuBP1 and AuBP2) sequences using a FliTrx random peptide display library. Here, we employed a single step peptide based controlled patterning where gold binding peptide, AuBP2, was used for the directed immobilization of a multifunctional protein on gold substrates. Multifunctional protein which includes green fluorescent protein (GFPuv) and Maltose Binding Protein (MBP) was successfully produced through genetic conjugation of combinatorially selected AuBP2 to GFPuv. Fusion DNA fragment was cloned into pMALc4x expression vector encoding N-terminal MBP tag. Following expression and purification, the excitation, emission wavelengths and fluorescence intensity of the multifunctional protein MBP-GFPuv-AuBP2 were compared with a negative control MBP-GFPuv. It has been shown that MBP-GFPuv and MBP-GFPuv-AuBP2 proteins have equivalent excitation and emission wavelengths with similar intensities. Protein patterns were prepared by directed stamping of MBP-GFPuv-AuBP and control MBP-GFPuv fusion proteins on flat gold surface. Fluorescence microscopy images of the protein-arrayed gold surfaces showed that the multifunctional MBP-GFPuv-AuBP protein immobilized on gold surface displays higher affinity compared to the negative control. Site-specific protein immobilization and patterning is important for the fabrication of efficient biotechnological tools such as biosensors and microarrays. Using AuBP2 as a specific molecular linker, we demonstrated for the controlled immobilization of a fluorescent heterofunctional protein on metal surface.