Local Environment Affects the Activity of Enzymes on a 3D Molecular Scaffold.

The ability to coordinate and confine enzymes presents an opportunity to affect their performance and to create chemically active materials. Recent studies show that polymers and biopolymers can be used to scaffold enzymes, and that can lead to the modulated biocatalytic efficiency. Here, we investigated the role of microenvironments on enzyme activity using a well-defined molecular scaffold. An enzyme, glucose oxidase (GOx), was positioned at different locations of a three-dimensional (3D) octahedral DNA scaffold (OS), allowing the enzyme's polyanionic environments to be altered. Using electrical sensing, based on a bipolar junction transistor, we measured directly and in real-time the enzyme's proton generation at these different microenvironments. We found a 200% enhancement of immobilized enzyme over free GOx and about a 30% increase in catalytic rates when the enzyme was moved on the same molecular scaffold to a microenvironment with a higher local concentration of polyanions, which suggests a role of local pH on the enzymatic activity.

Silicon Nanowire Field Effect Transistor Sensors with Minimal Sensor-to-Sensor Variations and Enhanced Sensing Characteristics.

Silicon nanowire field effect transistor (FET) sensors have demonstrated their ability for rapid and label-free detection of proteins, nucleotide sequences, and viruses at ultralow concentrations with the potential to be a transformative diagnostic technology. Their nanoscale size gives them their ultralow detection ability but also makes their fabrication challenging with large sensor-to-sensor variations, thus limiting their commercial applications. In this work, a combined approach of nanofabrication, device simulation, materials, and electrical characterization is applied toward identifying and improving fabrication steps that induce sensor-to-sensor variations. An enhanced complementary metal-oxide-semiconductor-compatible process for fabricating silicon nanowire FET sensors on 8 in. silicon-on-insulator wafers is demonstrated. The fabricated nanowire (30 nm width) FETs with solution gates have a Nernst limit subthreshold swing (SS) of 60 ± 1 mV/decade with ∼1.7% variations, whereas literature values for SS are ≥80 mV/decade with larger (>10 times) variations. Also, their threshold voltage variations are significantly (∼3 times) reduced, compared to literature values. Furthermore, these improved FETs have significantly reduced drain current hysteresis (∼0.6 mV) and enhanced on-current to off-current ratios (∼106). These improvements resulted in nanowire FET sensors with the lowest (∼3%) reported sensor-to-sensor variations, compared to literature studies. Also, these improved nanowire sensors have the highest reported sensitivity and enhanced signal-to-noise ratio with the lowest reported defect density of 2.1 × 1018 eV-1 cm-3, in comparison to literature data. In summary, this work brings the nanowire sensor technology a step closer to commercial products for early diagnosis and monitoring of diseases.

Comparison between Field Effect Transistors and Bipolar Junction Transistors as Transducers in Electrochemical Sensors.

Field effect transistors (FET) have been widely used as transducers in electrochemical sensors for over 40 years. In this report, a FET transducer is compared with the recently proposed bipolar junction transistor (BJT) transducer. Measurements are performed on two chloride electrochemical sensors that are identical in all details except for the transducer device type. Comparative measurements show that the transducer choice significantly impacts the electrochemical sensor characteristics. Signal to noise ratio is 20 to 2 times greater for the BJT sensor. Sensitivity is also enhanced: BJT sensing signal changes by 10 times per pCl, whereas the FET signal changes by 8 or less times. Also, sensor calibration curves are impacted by the transducer choice. Unlike a FET sensor, the calibration curve of the BJT sensor is independent of applied voltages. Hence, a BJT sensor can make quantitative sensing measurements with minimal calibration requirements, an important characteristic for mobile sensing applications. As a demonstration for mobile applications, these BJT sensors are further investigated by measuring chloride levels in artificial human sweat for potential cystic fibrosis diagnostic use. In summary, the BJT device is demonstrated to be a superior transducer in comparison to a FET in an electrochemical sensor.

Mechanistic Influence of Nanometer Length-Scale Surface Chemistry on DNA Hybridization.

Hybridization of surface-immobilized oligonucleotides to their complementary counterparts is central to the rational design of novel nanodevices and DNA sensors. In this study, we have adopted a unified approach of combining sensing experiments with molecular dynamics simulations to characterize the hybridization of a 23 nucleotide long single-strand probe DNA tethered to a gold surface. Experiments indicate significant conformational changes of DNA in close vicinity (∼1 nm) of the gold surface upon hybridization and also conformational heterogeneity within hybridized DNA, consistent with simulation results. Simulations show that the conformational heterogeneity on a gold surface arises due to stabilization of surface-adsorbed partial and full duplexes, resulting in impeded hybridization in comparison to what observed on a repulsive surface. Furthermore, these simulations indicate that hybridization could be improved by tuning the nonspecific adsorption on a nanopatterned surface with an optimal patterning length. Simulations were performed on the probe tethered to gold nanodots of varying (2-8 nm) diameter. An improved hybridization of the present probe sequence was only observed for the 6 nm gold dots patterned on a repulsive surface. Results reveal that the 2D nanoconfinement provided by the 6 nm gold dot is optimal for reducing conformational heterogeneity for the specific sequence used in this study. Thus, improved DNA hybridization can be achieved on a gold nanodot patterned repulsive surface, where the optimal dot diameter will depend on the probe length and sequence. In summary, this study provides mechanistic insights onto hybridization on gold and offers a unique method toward improved hybridization on a nanopatterned surface with an optimized patterning length.

Optimization of pH sensing using silicon nanowire field effect transistors with HfO2 as the sensing surface.

Silicon nanowire field effect transistor sensors with SiO(2)/HfO(2) as the gate dielectric sensing surface are fabricated using a top down approach. These sensors are optimized for pH sensing with two key characteristics. First, the pH sensitivity is shown to be independent of buffer concentration. Second, the observed pH sensitivity is enhanced and is equal to the Nernst maximum sensitivity limit of 59 mV/pH with a corresponding subthreshold drain current change of ∼ 650%/pH. These two enhanced pH sensing characteristics are attributed to the use of HfO(2) as the sensing surface and an optimized fabrication process compatible with silicon processing technology.

Specificity of ligand binding to transport sites: Ca2+ binding to the Ca2+ transport ATPase and its dependence on H+ and Mg2+.

Ligand binding to transport sites constitutes the initial step in the catalytic cycle of transport ATPases. Here, we consider the well characterized Ca2+ ATPase of sarcoplasmic reticulum (SERCA) and describe a series of Ca2+ binding isotherms obtained by equilibrium measurements in the presence of various H+ and Mg2+ concentrations. We subject the isotherms to statistical mechanics analysis, using a model based on a minimal number of mechanistic steps. The analysis allows satisfactory fits and yields information on occupancy of the specific Ca2+ sites under various conditions. It also provides a fundamental method for analysis of binding specificity to transport sites under equilibrium conditions that lead to tightly coupled catalytic activation.

Statistical mechanics model for the interaction between the neurotransmitter gamma-aminobutyric acid and GABAA receptors.

Interactions between the neurotransmitter gamma-aminobutyric acid (GABA) and GABAA receptor ion channels play an important role in the central nervous system. A statistical mechanics model is proposed for the interaction between GABA and GABAA receptors. The model provides good fits to the electrophysiology data as well as an estimation of receptor activation energies, and predicts the temperature dependence consistent with measurements. In addition, the model provides insights into single channel conductance measurements. This model is also applicable to other ligand-gated ion channels with similar pentameric structures.