title: L-Lactate-selective microbial sensor based on flavocytochrome b2-enriched yeast cells using recombinant and nanotechnology approaches.
abstract: In the recent years, nanotechnology is the most developing branch due to a wide variety of potential applications in biomedical, biotechnological and agriculture fields. The binding nanoparticles with various biological molecules makes them attractive candidates for using in sensor technologies. The particularly actual is obtaining the bionanomembranes based on biocatalytic elements with improved sensing characteristics. The aim of this investigation is to study the properties of microbial L-lactate-selective sensor based on using the recombinant Hansenula polymorpha yeast cells overproducing flavocytochrome b2 (FC b2), as well as additionally enriched by the enzyme bound with gold nanoparticles (FC b2-nAu). Although, the high permeability of the living cells to nanoparticles is being intensively studied (mostly for delivery of drugs), the idea of using both recombinant technology and nanotechnology to increase the amount of the target enzyme in the biosensing layer is really novel. The FC b2-nAu-enriched living and permeabilized yeast cells were used for construction of a bioselective membrane of microbial L-lactate-selective amperometric biosensor. Phenazine methosulphate was served as a free defusing electron transfer mediator which provides effective electron transfer from the reduced enzyme to the electrode surface. It was shown that the output to L-lactate of FC b2-nAu-enriched permeabilized yeast cells is 2.5-fold higher when compared to the control cells. The obtained results confirm that additional enrichment of the recombinant yeast cell by the enzyme bound with nanoparticles improves the analytical parameters of microbial sensor.
authors.affiliation: Department of Analytical Biotechnology, Institute of Cell Biology, NAS of Ukraine, Drahomanov Street 14/16, Lviv 79005, Ukraine.
authors.forename: Maria
authors.initials: M
authors.lastname: Karkovska
authors.affiliation: Department of Analytical Biotechnology, Institute of Cell Biology, NAS of Ukraine, Drahomanov Street 14/16, Lviv 79005, Ukraine.
authors.forename: Oleh
authors.initials: O
authors.lastname: Smutok
authors.affiliation: Department of Analytical Biotechnology, Institute of Cell Biology, NAS of Ukraine, Drahomanov Street 14/16, Lviv 79005, Ukraine.
authors.forename: Nataliya
authors.initials: N
authors.lastname: Stasyuk
authors.affiliation: Department of Analytical Biotechnology, Institute of Cell Biology, NAS of Ukraine, Drahomanov Street 14/16, Lviv 79005, Ukraine; Institute of Applied Biotechnology and Basic Sciences, Rzeszow University, Sokolowska Street 26, Kolbuszowa, Poland. Electronic address: gonchar@cellbiol.lviv.ua.
authors.forename: Mykhailo
authors.initials: M
authors.lastname: Gonchar
mesh_terms: D001692:Biological Transport; D015374:Biosensing Techniques / Q000379:methods*; D004274:DNA, Recombinant / Q000235:genetics*; D006046:Gold / Q000737:chemistry / Q000378:metabolism; D042844:L-Lactate Dehydrogenase (Cytochrome) / Q000235:genetics*; D019344:Lactic Acid / Q000032:analysis*; D053768:Metal Nanoparticles; D036103:Nanotechnology / Q000379:methods*; D010843:Pichia / Q000166:cytology* / Q000235:genetics* / Q000378:metabolism
chemical_list: D004274:DNA, Recombinant; D019344:Lactic Acid; D006046:Gold; D042844:L-Lactate Dehydrogenase (Cytochrome)
keywords: Amperometric biosensor; Enzyme-enriched cells; Flavocytochrome b(2); Gold nanoparticles; Microbial biosensor; Recombinant yeast cells; l-Lactate