5th International Symposium on Sensor Science
Prof. Dr. Evgeny Katz
Department of Chemistry & Biomolecular Science, Clarkson University, USA
Prof. Dr. Wolfgang Schuhmann
Center for Electrochemical Sciences (CES), Ruhr-University Bochum, Germany
Prof. Dr. Jose M. Pingarrón
Complutense University of Madrid, Faculty of Chemistry, Spain
Prof. Dr. Lital Alfonta
Department of Life Sciences, Ilse Katz Institute for Nanoscale Science and Technology, Ben-Gurion University of the Negev
* Genetically Engineering of Enzymes and Electrode Modifications for Biosensing Applications
Biosensing efficiency, selectivity and sensitivity relies first and foremost on a successful interfacing between enzymes and sensing surfaces. An Interface that allows from one hand a specific analyte recognition and on the other hand an efficient signal transduction. Some of the challenges in biosensing stem from wrong orientation of the enzyme towards the sensing interface and from the need to use mediated electron transfer with a diffusional redox mediator due to a difficulty in relaying a signal from a redox center that is deeply buried inside the protein matrix. Using genetic code expansion tools, and genetic engineering approaches we were able to modify redox enzymes and surfaces for biosensing and biofuel cell applications so they could have superior properties over native enzymes. In my talk, I will demonstrate how does site specific wiring of redox enzymes which is genetically encoded, can improve electron transfer due to controlled and short electron transfer distances and due to proper enzyme orientation. I will also demonstrate how a rational genetic engineering of an enzyme gives it superior properties for biosensing purposes compared to those of the native enzyme.
Prof. Dr. Michael J. Schöning
Institute of Nano- and Biotechnologies (INB), FH Aachen University of Applied Sciences, Germany
Prof. Dr. Dmitry Kolpashchikov
Chemistry Department, University of Central Florida, Orlando, FL, USA
*Nucleic Acid Analysis Using Multifunctional Hybridization Sensors
Hybridization of nucleic acid probes remains one of the most common strategy for sensing of specific DNA and RNA sequences. Formats that use hybridization probes include qualitative PCR, microarrays, fluorescent in situ hybridization (FISH), to name a few. Moreover, specific recognition of RNA sequences is on demand by gene silencing approaches, e.g. antisense and siRNA. Hybridization probes are nucleic acid oligomers of 15−25 nucleotides (or longer) designed to be complementary to targeted analytes. The formation of a probe-analyte hybrid testifies that the analyte contains a nucleotide sequence complementary to the probe. This approach suffers from low selectivity (especially for temperatures <40oC), high cost for fluorescent probes, and poor target accessibility if folded natural RNA are analyzed. Part of the sensing problem arises from the affinity/selectivity dilemma: the higher the probe-analyte affinity, the lower the selectivity. To address these and other problems, we design multicomponent hybridization probes (MHP) that consist of several oligonucleotide components, which associate with RNA/DNA target and produce a detectable signal. Each MHP component serves specific function, thus enabling simultaneous improvement of multiple key characteristics. My presentation will cover the design of a probe that can differentiate single nucleotide substitutions in DNA in the entire temperature interval of 5-40oC; a molecular machine that tightly binds RNA analyte while remains highly selectivity; a strategy for recognition of highly variable viral genomes with high selectivity.
Dr. Marcos Pita
Instituto de Catálisis y Petroleoquímica, CSIC, Spain
*ATP Synthesis and Biosensing Coupled to the Electroenzymatic Activity of a Hydrogenase on an Electrode/Biomimetic Membrane Interface
Cells generate energy by coupling a proton gradient across a phospholipid bilayer membrane with the activity of a cross-membrane ATP synthase enzyme. In an effort to mimic this process in an artificial environment, we show that ATP can be efficiently produced starting from molecular hydrogen as a fuel.
The proton concentration in an electrode/phospholipid bilayer interface can be controlled and monitorised electrochemically by immobilizing the membrane-bound [NiFeSe]-hydrogenase from Desulfovibrio vulgaris Hildenborough. The electro-enzymatic oxidation of H2 generated a proton gradient across the supported biomimetic membrane that can be coupled to the in vitro synthesis of ATP by reconstituting ATP-synthase from E. coli on the biomimetic system. Such system is also suitable for developing an electrochemical biosensor of ATP.