Tiny Tech With Big Impact

Courtesy Photo | Increased local substrate concentrations in enzyme-DNA nanostructures as a result of engineered substrate-DNA interactions. Left: The DNA structures used here to investigate the effects of substrate-DNA binding interactions on enzyme catalysis. Colored strands are indicative of sequence. Right: Representation of enzyme-DNA nanostructures with a double-crossover DNA tile chemically conjugated to an HRP. The mechanism of increased local substrate concentration and enhanced catalysis is shown schematically: binding interactions between enzyme substrates and the DNA scaffold result in increased local concentrations of substrate, giving rise to a decrease in apparent Michaelis-Menten constant (KM) and a corresponding increase in enzyme efficiency (kcat/KM).  

FORT BELVOIR, VA, UNITED STATES

09.12.2017

Courtesy Story

Defense Threat Reduction Agency's Chemical and Biological Technologies Department

Sometimes the smallest technologies provide the biggest impact for protecting warfighters from chemical and biological attacks. Researchers, working for the Defense Threat Reduction Agency’s Toxicant Penetration and Scavenging (TPS) portfolio, are developing enzyme-DNA nanostructures to provide a foundation for understanding catalytic enhancement in nano structured materials. Managed by Brian Pate, Ph.D., the TPS program explores chemical and biological agent countermeasure development as well as the structural relationships. For this project, DTRA’s Chemical and Biological Technologies Department is exploring DNA nanomaterials and their ability to control molecular features of enzyme systems. This has proven effective in enhancing catalysis, creating efficient multistep cascades and improving enzyme stability.

A study published in ChemBioChem, “Mechanisms of Enhanced Catalysis in Enzyme–DNA Nanostructures Revealed through Molecular Simulations and Experimental Analysis,” highlights progress in defining mechanisms of enhanced catalysis in enzyme-DNA nanostructures. Researchers used simulations of enzyme-DNA nanostructures to identify and understand a mechanism of enhanced catalysis through engineered substrate-DNA interactions.

Focusing on the horseradish peroxidase (HPR) enzyme, researchers reengineered HPR to DNA tiles, a basic building block of higher-order DNA structures, and double-stranded (ds) DNA fragments. They then used kinetic analysis, modeling and molecular -level simulations to show that increased substrate concentrations in close proximity to the enzyme and active site resulted in catalytic enhancements.

This approach is applicable to a wide range of enzymes and substrates, and may have future applications for medical countermeasure development and degrading chemical warfare agents, as well as any toxic byproducts for improved warfighter safety. It can also be used to develop environmental decontaminants for detoxifying military equipment.

The molecular simulations and kinetic experiments provide evidence that supports a mechanism of enhanced catalysis in enzyme to DNA nanostructures through increased local concentrations of substrates in close proximity to the enzyme and active site.
Binding of small-molecule compounds to dsDNA is well known, with the major and minor grooves of the double helix structure available as repeat binding sites. Binding interactions can also occur through intercalation between base pairs and through electrostatic interactions with the negatively charged phosphate backbone.

In addition, many other small-molecule compounds are also known to bind to dsDNA such as anticancer drugs, molecular imaging probes and commercially available DNA binding fluorophores and polymer precursors. These findings will help enable the production of more robust products to protect warfighters.

In light of the experimental and simulation results, researchers can propose a design rule for enzyme to DNA nanostructures. In its simplistic form, the design rule states that binding interactions between substrates and enzyme to DNA nanostructures can alter enzyme catalysis and should be considered in the design of hybrid structures. More specifically, this data demonstrates that researchers can use binding interactions in the micromolar range between a substrate and an enzyme- DNA nanostructure to increase the local concentration of substrates. This increase enhances catalysis by promoting substrate association to the active site.

DTRA’s latest nanostructure research continues to advance the TPS portfolio, while exploring preliminary efforts for new medical countermeasures and decontamination methods for the warfighter.

POC: Brian Pate, Ph.D.; [e-mail] 5

• Date Taken: 09.12.2017
• Date Posted: 09.12.2017 14:28
• Story ID: 247926
• Location: FORT BELVOIR, VA, US