Sequence-specific interactions of proteins with DNA are central to all aspects of the utilization of genetic information in any organism. The lactose repressor of E. coli served as a paradigm for such interactions even before the chemical structure of the interacting partners was elucidated. The lac repressor protein recognizes the lac operator, a particular region of base pairs in the chromosome of E. coli and binds to it tightly with a dissociation constant of 10 À11 ± 10 À13 m. Sitespecific recognition of DNA by the lac repressor is interrupted by an inducer, such as lactose, to allow the production of the enzymes necessary for the utilization of this carbon source. [3] A major conformational change in the lac repressor structure takes place as the result of inducer binding. [4, The majority of inducers that bind to the lac repressor are galactose derivatives, such as isopropyl-d-thiogalactoside (IPTG), o-nitrophenyl-d-galactoside (ONPG), and 1,6-allolactose. [6] Other sugars like o-nitrophenylfucoside (ONPF) also bind strongly to the lac repressor. Capacitance measurements have been successfully used as a basis for the construction of biosensors for sensitive detection of the human chorionic gonadotropin (HCG) hormone, by immobilization of antibodies on the electrode surface. Heavy metals can also be detected, with heavy metal binding proteins as recognition elements. The capacitive transduction principle has now been used for the development of a biosensor to monitor inducer molecules or DNA, through the use of a repressor protein as the biological recognition element.
Biosensors prepared by immobilizing the lac repressor protein on a gold surface modified with thioctic acid have been used in the experimental set-up presented schematically, along with the detection principle in Figure . The specificity of lac repressor based biosensors for operator DNA was tested by injections of plasmid p310 DNA, two linearized plasmid DNAs, and genomic DNA. Plasmid p310 DNA (2455 base pairs (bp) in length) was constructed by cloning a 24 bp fragment that contains the lac ideal operator into the NheI site of plasmid pEE4. One linearized plasmid DNA was obtained by digestion of the plasmid p310 DNA with EcoRI, and the target lac operator (the second linearized plasmid DNA) was excised as an 84 bp fragment by cutting with EcoRI and HindIII.