Single-Molecule Kinetics

Single-molecule kinetics provides an unparalleled perspective on enzymatic activity by examining individual enzyme molecules rather than ensemble averages. This approach enables the observation of transient states, conformational heterogeneity, and stochastic behaviors that are often obscured in bulk measurements. By leveraging advanced single-molecule detection techniques, Creative Enzymes empowers researchers to gain precise, mechanistic insights into enzyme function, regulation, and dynamics.

Understanding Single-Molecule Kinetics

Single-molecule kinetics is a cutting-edge approach that studies enzymatic behavior at the individual molecule level, providing insights hidden in ensemble-averaged measurements. Unlike traditional bulk assays, which observe the average activity of millions of molecules, single-molecule techniques include:

• Single-Molecule Fluorescence (e.g., FRET, TIRF microscopy): Tracks conformational changes, binding events, or catalytic turnover by monitoring fluorescent signals at the level of individual enzyme molecules.
• Optical Tweezers / Magnetic Tweezers: Apply mechanical forces to measure enzyme-driven motions such as DNA/RNA unwinding or protein folding in real time.
• Atomic Force Microscopy (AFM): Provides nanoscale resolution to monitor structural changes or mechanical properties during enzyme catalysis.
• Nanopore Sensors: Detect substrate conversion or product release as changes in ionic current when molecules pass through a nanopore.

Uncovering Insights

• Heterogeneity: Enzymes of the same type can exhibit distinct catalytic rates, conformational dynamics, and transient pauses due to stochastic fluctuations.
• Intermediate States: Direct observation of short-lived intermediates and rare events (e.g., substrate binding, conformational changes).
• Mechanistic Details: Real-time tracking of catalytic cycles, processivity (e.g., in polymerases), and force-dependent kinetics (e.g., motor proteins).

Applications

• Resolving dynamic disorder in enzyme turnover.
• Studying allosteric regulation and cofactor interactions.
• Designing precision biocatalysts with tailored properties.

By bypassing ensemble averaging, single-molecule kinetics unveils the full spectrum of enzymatic behavior, refining models of catalysis and aiding drug discovery.

Our Service Offerings

Service Workflow

Service Description

Creative Enzymes offers comprehensive single-molecule kinetic analysis services, combining high-resolution instrumentation with proprietary assay development. We specialize in:

• Real-time monitoring of individual enzyme turnover events.
• Detection and quantification of intermediate states and transient conformations.
• Analysis of heterogeneity within enzyme populations.
• Investigation of processivity, pausing, and catalytic rate distributions.
• Integration with fluorescent, FRET-based, and other single-molecule labeling strategies.

Our platform accommodates both natural and engineered enzymes across diverse families and substrates, tailored to the specific research needs of each client.

Samples and Deliveries

Contact Our Team

Advantages of Choosing Creative Enzymes

Representative Case Studies

FAQs

• Q: What enzyme types are suitable for single-molecule kinetic studies?

  A: Our platform accommodates a wide range of enzymes including hydrolases, transferases, ligases, polymerases, and motor proteins, provided the enzyme-substrate system can be suitably labeled and monitored.

• Q: How much sample is required?

  A: Single-molecule studies generally require low nanomolar to picomolar concentrations, significantly reducing the amount of enzyme needed compared to bulk assays.

• Q: Can you provide kinetic parameters comparable to classical Michaelis-Menten analysis?

  A: Yes, we derive traditional kinetic metrics, including turnover rates and catalytic efficiencies, while also providing insights into heterogeneity and transient states not accessible in ensemble measurements.

• Q: Are labeling strategies included in the service?

  A: Yes, we offer guidance on fluorescent or other labeling methods, and can integrate labeling as part of the assay development process if required.

• Q: How long does a typical single-molecule kinetics study take?

  A: Timelines vary depending on assay complexity, enzyme type, and labeling strategy, but most studies are completed within 4–8 weeks from sample submission to final report.