Controlled Protein Activities with Viral Proteases, Antiviral Peptides, and Antiviral Drugs.

Chemical control of protein activity is a powerful tool for scientific study, synthetic biology, and cell therapy; however, for broad use, effective chemical inducer systems must minimally crosstalk with endogenous processes and exhibit desirable drug delivery properties. Accordingly, the drug-controllable proteolytic activity of hepatitis C cis-protease NS3 and its associated antiviral drugs have been used to regulate protein activity and gene modulation. These tools advantageously exploit non-eukaryotic and non-prokaryotic proteins and clinically approved inhibitors. Here, we expand the toolkit by utilizing catalytically inactive NS3 protease as a high affinity binder to genetically encoded, antiviral peptides. Through our approach, we create NS3-peptide complexes that can be displaced by FDA-approved drugs to modulate transcription, cell signaling, and split-protein complementation. With our developed system, we invented a new mechanism to allosterically regulate Cre recombinase. Allosteric Cre regulation with NS3 ligands enables orthogonal recombination tools in eukaryotic cells and functions in divergent organisms to control prokaryotic recombinase activity.

Controlled protein activities with viral proteases, antiviral peptides, and antiviral drugs.

Chemical control of protein activity is a powerful tool for scientific study, synthetic biology, and cell therapy; however, for broad use, effective chemical inducer systems must minimally crosstalk with endogenous processes and exhibit desirable drug delivery properties. Accordingly, the drug-controllable proteolytic activity of hepatitis C cis- protease NS3 and its associated antiviral drugs have been used to regulate protein activity and gene modulation. These tools advantageously exploit non-eukaryotic/prokaryotic proteins and clinically approved inhibitors. Here we expand the toolkit by utilizing catalytically inactive NS3 protease as a high affinity binder to genetically encoded, antiviral peptides. Through our approach, we create NS3-peptide complexes that can be displaced by FDA-approved drugs to modulate transcription, cell signaling, split-protein complementation. With our developed system, we discover a new mechanism to allosterically regulate Cre recombinase. Allosteric Cre regulation with NS3 ligands enables orthogonal recombination tools in eukaryotic cells and functions in divergent organisms to control prokaryotic recombinase activity.

Electrical conductivity and ion diffusion in porcine meniscus: effects of strain, anisotropy, and tissue region.

The purpose of the present study was to investigate the effects of mechanical strain, anisotropy, and tissue region on electrical conductivity and ion diffusivity in meniscus fibrocartilage. A one-dimensional, 4-wire conductivity experiment was employed to measure the electrical conductivity in porcine meniscus tissues from two tissue regions (horn and central), for two tissue orientations (axial and circumferential), and for three levels of compressive strain (0%, 10%, and 20%). Conductivity values were then used to estimate the relative ion diffusivity in meniscus. The water volume fraction of tissue specimens was determined using a buoyancy method. A total of 135 meniscus samples were measured; electrical conductivity values ranged from 2.47mS/cm to 4.84mS/cm, while relative ion diffusivity was in the range of 0.235 to 0.409. Results show that electrical conductivity and ion diffusion are significantly anisotropic (p<0.001), both being higher in the circumferential direction than in the axial direction. Additionally, the findings show that compression significantly affects the electrical conductivity with decreasing conductivity levels corresponding to increased compressive strain (p<0.001). Furthermore, there was no statistically significant effect of tissue region when comparing axial conductivity in the central and horn regions of the tissue (p=0.999). There was a positive correlation between tissue water volume fraction and both electrical conductivity and relative ion diffusivity for all groups investigated. This study provides important insight into the electromechanical and transport properties in meniscus fibrocartilage, which is essential in developing new strategies to treat and/or prevent tissue degeneration.