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DNA-heme conjugates for the reconstitution and DNA directed assembly of redox enzymes


Currently there is a great interest in using DNA as scaffold for the assembly of nanoscale arrays. The DNA directed assembly of proteins at solid substrates (glass or metal) or at the surface of nanoparticles is an exciting area of DNA nanotechnology, which will open novel ways of utilising the large variety of highly functional proteins. Nowadays the scientific and technological interest is rapidly moving towards the production of enzyme arrays. The proposed project is aimed to explore a new concept for generating micro and nanostructured redox enzyme -DNA conjugates by means of DNA directed assembly. This will be achieved by covalently linking heme with a single stranded DNA and use of the resulting DNA-heme conjugate for reconstitution of heme dependent enzymes. Subsequently, the DNA tagged enzymes will be immobilised on gold nanoparticles, which contain complementary capture DNA.

The redox enzymes adsorbed on the gold surface will be characterised by electrochemical and spectroscopie means, including novel sensitive technique; surface enhanced resonance Raman scattering (SERRS). Additionally, the DNA-enzyme-gold nanoparticle conjugates will be immobilised at macroscopic gold electrode byDNA hybridisation, taking the advantage of the second coding DNA sequence attached to the gold nanoparticles. These experiments should provide the basis for continuing research devoted to the DNA directed assembly of micro and nano-structured enzyme arrays, which have many possible applications in the area of biosensing and material's research. Moreover, the results of this work will also provide information useful for the production of micro-arrays that can be utilised for screening of enzymes in the unknown sample as well as for the investigation of the activity of enzyme libraries.

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