UNIVERSAL BIOSENSORS

THE PRIMARY BENEFIT OF BIOSENSORS IS THE CAPACITY TO GAIN RAPID INFORMATION ON THE PRESENCE OF DIFFERENT ANALYTES. A MAJOR ADVANTAGE OF BIOSENSORS OVER MORE SOPHISTICATED APPROACHES IS THAT IT DOES NOT REQUIRE SOPHISTICATED INSTRUMENTATION. BIOSENSOR DEVELOPMENT IS OF ENORMOUS BIOTECHNOLOGICAL INTEREST AS WITNESSED BY MORE THAN 8000 PATENTS DEPOSITED IN THE WORLD-WIDE PATENT DATABASE THAT HAVE THE TERM ¿BIOSENSOR¿ IN THEIR TITLE. HOWEVER, MOST BIOSENSORS ARE WHOLE CELL BIOSENSORS THAT RELY ON LIFE BACTERIA AND THAT ARE BASED ON CELLULAR ANALYTE RECOGNITION THAT IN TURN TRIGGER CHANGES IN REPORTER GENE TRANSCRIPTION. SUCH BIOSENSORS, ALTHOUGH USEFUL, PRESENT A NUMBER OF DISADVANTAGES RELATED TO THE NEED OF ACTIVE BACTERIA FOR MEASUREMENTS, EFFECTS OF ANALYTES ON BIOSENSOR VIABILITY AND PROBLEMS OF ANALYTE BIOAVAILABILITY. THIS PROJECTS AIMS AT EXPLOITING MAJOR ADVANCES IN BACTERIAL SENSOR RESEARCH SUCH AS THE IDENTIFICATION OF THE LIGAND PROFILE OF AN ENORMOUS NUMBER OF SENSOR DOMAINS AND RESEARCH ON GGDEF BASED SIGNAL TRANSDUCTION SYSTEMS. THIS PROJECT IS BASED ON THE ABUNDANT FAMILY OF PROTEINS CONTAINING A GGDEF DOMAIN AND A SENSOR DOMAIN AT ITS N-TERMINAL EXTENSION. IT HAS BEEN SHOWN THAT LIGAND BINDING TO THE SENSOR DOMAIN ALTERS THE CATALYTIC ACTIVITY OF THE GGDEF DOMAINS. WE ENVISAGE HERE THE CONSTRUCTION OF A NOVEL, ENZYME BASED BIOSENSOR TYPE IN WHICH WELL CHARACTERIZED SENSOR DOMAINS (SUCH AS THOSE FOR AROMATIC CONTAMINANTS OR UREA) ARE FUSED TO THE N-TERMINUS OF GGDEF DOMAINS, SUBSTITUTING THE NATIVE SENSOR DOMAIN. GGDEF DOMAINS CHOSEN HAVE PREVIOUSLY BEEN SHOWN TO BE ACTIVE UNDER IN VITRO CONDITIONS. OUR HYPOTHESIS IS BASED ON THE ASSUMPTION THAT LIGAND BINDING TO THE CHIMERA WILL ALTER GGDEF ACTIVITY THAT CAN BE MONITORED BY THE MALACHITE GREEN BASED ASSAYS THAT PERMITS THE DETECTION OF PICOMOLAR AMOUNTS OF INORGANIC PHOSPHATE. NO INSTRUMENTATION IS REQUIRED TO OBSERVE THE BIOSENSOR OUTPUT SINCE THE PRESENCE OF LIGAND WILL RESULT IN A GREEN COLOR.