ABSTRACT
A new biomimetic nanoreactor design, MaBiDz, is presented based on a copolymer brush in combination with superparamagnetic nanoparticles. This cellular nanoreactor features two species of magnetic particles, each functionalized with two components of a binary deoxyribozyme system. In the presence of a target mRNA analyte and a magnetic field, the nanoreactor is assembled to form a biocompartment enclosed by the polymeric brush that enables catalytic function of the binary deoxyribozyme with enhanced kinetics. MaBiDz was demonstrated here as a cellular sensor for rapid detection and imaging of a target mRNA biomarker for metastatic breast cancer, and its function shows potential to be expanded as a biomimetic organelle that can downregulate the activity of a target mRNA biomarker.
Subject(s)
DNA, Catalytic/chemistry , Magnetic Fields , Magnetite Nanoparticles/chemistry , Biomarkers, Tumor/analysis , Humans , MCF-7 Cells , Nuclear Proteins , Polymers , RNA, Messenger/analysis , Twist-Related Protein 1ABSTRACT
Detection of specific mRNA in living cells has attracted significant attention in the past decade. Probes that can be easily delivered into cells and activated at the desired time can contribute to understanding translation, trafficking and degradation of mRNA. Here we report a new strategy termed magnetic field-activated binary deoxyribozyme (MaBiDZ) sensor that enables both efficient delivery and temporal control of mRNA sensing by magnetic field. MaBiDZ uses two species of magnetic beads conjugated with different components of a multicomponent deoxyribozyme (DZ) sensor. The DZ sensor is activated only in the presence of a specific target mRNA and when a magnetic field is applied. Here we demonstrate that MaBiDZ sensor can be internalized in live MCF-7 breast cancer cells and activated by a magnetic field to fluorescently report the presence of specific mRNA, which are cancer biomarkers.
Subject(s)
Magnetics , RNA, Messenger/metabolism , Breast Neoplasms/metabolism , Breast Neoplasms/pathology , Cells, Cultured , DNA, Catalytic/metabolism , Female , Humans , MCF-7 CellsABSTRACT
Signal-controlled release of DNA from Fe3+ -cross-linked alginate hydrogel electrochemically deposited on an electrode surface was studied. The multiple input signals were logically processed with the help of the enzyme-biocatalyzed reactions. Boolean logic gates, OR, AND, INH, were realized with the biocatalytic reactions performed by the enzymes entrapped in the alginate film. Hydrogen peroxide produced by the enzymatic reactions resulted in the degradation of the alginate hydrogel and DNA release. The alginate degradation was facilitated by the formation of free radicals in the Fenton-type reaction catalyzed by iron cations cross-linking the alginate hydrogel. The studied approach is versatile and can be adapted to various chemical signals processed by various enzymes with differently implemented Boolean logic. This work illustrates a novel concept of functional integration of biomolecular computing and actuation.
Subject(s)
Alginates/chemistry , Computers, Molecular , Cross-Linking Reagents/chemistry , DNA/metabolism , Ferric Compounds/chemistry , Logic , Animals , Biocatalysis , DNA/chemistry , Esterases/chemistry , Esterases/metabolism , Glucose Oxidase/chemistry , Glucose Oxidase/metabolism , Glucuronic Acid/chemistry , Hexuronic Acids/chemistry , Horseradish Peroxidase/chemistry , Horseradish Peroxidase/metabolism , Lactate Dehydrogenases/chemistry , Lactate Dehydrogenases/metabolism , Mixed Function Oxygenases/chemistry , Mixed Function Oxygenases/metabolism , Nanoparticles/chemistry , Nanoparticles/metabolism , Silicon Dioxide/chemistry , Silicon Dioxide/metabolismABSTRACT
A new biocatalytic assay analyzing the simultaneous presence of creatine kinase (CK) and alanine transaminase (ALT) was developed aiming at the recognition of biofluids of different gender for forensic applications. Knowing the difference in the concentrations of CK and ALT enzymes in the blood of healthy adults of male and female groups we mimicked the samples of different gender with various CK-ALT concentrations. The analysis was performed using a multi-enzyme/multi-step biocatalytic cascade where the differences in both included enzymes resulted in an amplified difference in the final analytical response. The analysis performed in human serum solutions allowed discrimination of samples corresponding to male/female groups. The robustness of the developed analysis allowed determination of the gender for serum solutions after their drying and ageing at least for 1 hour. Importantly for forensic applications, reaction with a chromogenic reactant nitroblue tetrazolium allowed qualitative discrimination of the "male" and "female" samples by the naked eye.
Subject(s)
Biocatalysis , Forensic Sciences/methods , Sex Determination Analysis/methods , Adult , Alanine Transaminase/blood , Alanine Transaminase/metabolism , Biomarkers/blood , Biomarkers/metabolism , Creatine Kinase/blood , Creatine Kinase/metabolism , Female , Humans , MaleABSTRACT
We report the first systematic study of designed two-input biochemical systems as information processing gates with favorable noise transmission properties accomplished by modifying the gate's response from a convex shape to sigmoid in both inputs. This is realized by an added chemical "filter" process, which recycles some of the output back into one of the inputs. We study a system involving the biocatalytic function of the enzyme horseradish peroxidase, functioning as an AND gate. We consider modularity properties, such as the use of three different input chromogens that when oxidized yield signal detection outputs for various ranges of the primary input, hydrogen peroxide. We also examine possible uses of different filter effect chemicals (reducing agents) to induce the sigmoid response. A modeling approach is developed and applied to our data, allowing us to describe the enzymatic kinetics in the framework of a formulation suitable for evaluating the noise-handling properties of the studied systems as logic gates for information processing steps.
