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Phosphotriesterases (PTEs) represent a class of enzymes capable of efficient neutralization of organophosphates (OPs), a dangerous class of neurotoxic chemicals. PTEs suffer from low catalytic activity, particularly at higher temperatures, due to low thermostability and low solubility. Supercharging, a protein engineering approach via selective mutation of surface residues to charged residues, has been successfully employed to generate proteins with increased solubility and thermostability by promoting charge–charge repulsion between proteins. We set out to overcome the challenges in improving PTE activity against OPs by employing a computational protein supercharging algorithm in Rosetta. Here, we discover two supercharged PTE variants, one negatively supercharged (with −14 net charge) and one positively supercharged (with +12 net charge) and characterize them for their thermodynamic stability and catalytic activity. We find that positively supercharged PTE possesses slight but significant losses in thermostability, which correlates to losses in catalytic efficiency at all temperatures, whereas negatively supercharged PTE possesses increased catalytic activity across 25°C–55°C while offering similar thermostability characteristic to the parent PTE. The impact of supercharging on catalytic efficiency will inform the design of shelf-stable PTE and criteria for enzyme engineering.

During the abrupt and unplanned transition to remote online learning formats due to the COVID-19 outbreak, educators have had to adopt new teaching methods. For instance, online simulations tailored to specific curriculum topics emerged, allowing students to apply their knowledge creatively, with potentially positive effects on engagement and learning efficacy. Here, we examine the implementation of the “Save the World” simulation, created by Wonderville.org, in a high school Advanced Placement Environmental Science classroom in a remote online learning setting. In this module, students determine the most viable renewable energy generation option for given environments. Based on student and teacher feedback, the simulation effectively delivers educational material and promotes student engagement, demonstrating that online simulations can serve as a viable tool to enhance environmental science education and remote learning.