Isotope Labeling Studies

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Isotope labeling is a powerful and precise tool for dissecting enzymatic reaction pathways and identifying transient intermediates. At Creative Enzymes, our Isotope Labeling Studies enable the detailed investigation of substrate transformation, catalytic residues, and reaction mechanisms. By incorporating stable or radioactive isotopes into substrates or cofactors, combined with advanced analytical techniques, we provide high-resolution mechanistic insights that are essential for enzyme engineering, drug development, and fundamental biochemical research.

Understanding Isotope Labeling

Enzyme-catalyzed reactions often involve complex pathways, with short-lived intermediates that are challenging to detect using conventional methods. Isotope labeling allows researchers to trace the fate of atoms within substrates, revealing bond cleavage, rearrangement events, and active site interactions. Stable isotopes such as ¹³C, ¹⁵N, and ¹⁸O or radioactive isotopes such as ³H and ¹⁴C can be employed depending on experimental requirements. When integrated with spectroscopic, chromatographic, or mass spectrometric analysis, isotope labeling provides unparalleled mechanistic resolution, facilitating rational enzyme modification and substrate design.

Our Service Offerings

Service Workflow

Workflow of isotope labeling study service

Service Details

Creative Enzymes delivers comprehensive Isotope Labeling Studies, combining experimental precision with analytical sophistication:

Service	Details
Design and Synthesis	Custom design of isotopically labeled substrates or cofactors tailored to specific enzymatic reactions. Synthesis of site-specific labels to track individual atoms or functional groups during catalysis.
Experimental Implementation	Incorporation of isotopes into enzyme-catalyzed reactions under controlled conditions. Detection of isotope incorporation or displacement using advanced analytical techniques such as NMR spectroscopy, mass spectrometry, or liquid chromatography. Monitoring transient intermediates and reaction kinetics to identify rate-limiting steps.
Mechanistic Insights	Mapping bond cleavage, formation, and rearrangement within substrates. Identification of active site residues interacting with labeled atoms. Quantitative assessment of isotope effects to elucidate transition states and catalytic mechanisms.
Data Integration and Interpretation	Correlation of experimental findings with computational models for comprehensive mechanistic understanding. Construction of detailed reaction schemes highlighting substrate transformation and intermediate states. Visualization of isotope incorporation patterns and mechanistic pathways for actionable insights.
Reporting and Recommendations	Detailed documentation of experimental procedures, results, and interpretations. Graphical representation of labeled reaction pathways, intermediate states, and isotope effects. Strategic recommendations for enzyme optimization, inhibitor design, or process engineering.

Contact Our Team

Why Choose Creative Enzymes

Expertise

Skilled team with extensive experience in isotope chemistry, enzymology, and mechanistic analysis.

Comprehensive Methodology

Integration of isotope labeling, analytical detection, and computational modeling ensures reliable results.

Customization

Tailored solutions for a wide range of enzyme classes and substrate types.

Advanced Instrumentation

Access to high-resolution NMR, mass spectrometry, and chromatography platforms.

Actionable Insights

Mechanistic data directly support enzyme engineering, inhibitor development, and process optimization.

Regulatory Compliance

Adherence to safety, regulatory, and quality standards for handling isotopes.

Case Studies and Success Stories

Case 1: Tracing Amino Acid Metabolism in Penicillin Production

Client Challenge:

A pharmaceutical company producing penicillin V wanted to understand the contribution of amino acid biosynthesis versus medium uptake in Penicillium chrysogenum. Traditional methods provided only indirect evidence, limiting their ability to optimize yields.

Our Approach:

Applied reciprocal ¹³C labeling, using uniformly ¹³C -labeled glucose as the primary carbon source and naturally labeled amino acids as cosubstrates.
Measured incorporation into biomass using GC–MS analysis to detect differential ¹³C content in key metabolites.
Quantified fluxes through amino acid biosynthetic pathways versus uptake from the medium.

Outcome:

Revealed that phenylalanine and valine were primarily sourced from medium uptake, while other amino acids were largely synthesized de novo.
Provided actionable insights that allowed the client to adjust medium composition, increasing penicillin V production efficiency by 15%.

Case Study 2: Investigating Cofactor Utilization in a Novel Oxidoreductase

Client Challenge:

A biotech startup developing a novel oxidoreductase for industrial biocatalysis needed to understand cofactor turnover and substrate channeling, critical for scaling the reaction efficiently.

Our Approach:

Designed ¹³C- and ¹⁵N-labeled substrate feeding experiments to trace the flow of atoms through enzyme-catalyzed reactions.
Combined mass spectrometry-based isotopologue analysis with kinetic modeling to deduce enzyme mechanism and cofactor regeneration efficiency.
Identified intermediate accumulation and potential rate-limiting steps in the pathway.

Outcome:

Determined that NADPH recycling was limiting turnover and identified a minor side reaction reducing efficiency.
Enabled rational engineering of the enzyme and adjustment of reaction conditions, boosting catalytic efficiency by 3-fold in pilot-scale reactions.

FAQs About Enzyme Activity Measurement Services

Q: Which isotopes can be used in your studies?

A: We utilize both stable isotopes (¹³C, ¹⁵N, ¹⁸O) and radioactive isotopes (³H, ¹⁴C) depending on experimental requirements and safety considerations.
Q: Are special safety precautions required?

A: Yes, radioactive isotopes require strict adherence to safety protocols and regulatory compliance. Our facility is fully equipped to handle such materials safely.
Q: What types of enzymes can be studied?

A: Our service accommodates a wide range of enzymes, including hydrolases, oxidoreductases, transferases, lyases, and ligases.
Q: How long does an isotope labeling study typically take?

A: Depending on enzyme complexity and labeling strategy, studies generally require 6–12 weeks, with expedited options available.
Q: What deliverables are included?

A: Clients receive a detailed report including experimental design, data analysis, isotope incorporation maps, mechanistic interpretation, and strategic recommendations.
Q: Can results guide enzyme engineering or drug development?

A: Yes, isotope labeling studies provide high-resolution mechanistic insights that directly inform rational enzyme design, substrate modification, and inhibitor development.

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