Enzyme Conjugation with Nucleic Acids

inquiry

Combining enzymes with nucleic acids creates versatile molecular tools that transform biological research, diagnostics, and biosensing applications. Enzyme-nucleic acid conjugates facilitate sensitive detection of DNA or RNA sequences, enabling breakthroughs in medical diagnostics, gene expression analysis, infectious disease detection, and even biodefense. At Creative Enzymes, we specialize in the development of enzyme-linked oligonucleotides, HRP- or AP-labeled probes, and conjugates tailored for both direct and indirect detection systems. By leveraging state-of-the-art protein expression, purification, and conjugation technologies, we deliver high-quality, well-characterized conjugates that empower precise, reproducible, and high-sensitivity assays for research and commercial applications.

Background: Understanding the Significance of Enzyme-Nucleic Acid Conjugation

Enzymes and nucleic acids are two fundamental biomolecules, each with unique biological functions. Enzymes catalyze biochemical reactions with high specificity and efficiency, while nucleic acids store, transmit, and regulate genetic information. When combined, their properties create multifunctional conjugates capable of signal amplification, target recognition, and molecular labeling.

Applications of Enzyme-Nucleic Acid Conjugates

Medical Diagnostics: Detection of specific DNA or RNA sequences for disease diagnosis, mutation analysis, or pathogen identification.
Biosensing and Bioelectronics: Probe-enzyme conjugates are central to genosensor platforms, enabling rapid, sensitive, and selective detection.
Gene Expression Analysis: Quantitative measurement of mRNA or DNA copy numbers using conjugated probes in PCR-based or hybridization-based assays.
Biological Defense & Environmental Monitoring: Detection of infectious agents or genetic material from environmental samples with high specificity.

Enzyme-nucleic acid conjugates as genosensors Figure 1. A. Genosensor signal amplification approaches using probe-HRP conjugates; B. Hybrid –dsDNA detection probe containing three scCro DNA binding sites (in dark green) with HRP–scCro conjugate bound. (Aktas et al., 2015)

Mechanisms of Conjugation

Direct Conjugation: The enzyme is chemically linked to a nucleic acid probe via covalent bonds. For example, ssDNA-HRP conjugates are widely used in both colorimetric and electrochemical assays.
Indirect Conjugation: Biotinylated nucleic acid probes are recognized by enzyme-labeled proteins such as streptavidin-HRP or alkaline phosphatase (AP), facilitating signal generation through enzymatic reactions.

Recent innovations include hybrid detection systems, where a DNA-binding protein tagged with HRP interacts with a hybrid ssDNA-dsDNA probe, significantly amplifying detection signals. These approaches are becoming central to next-generation biosensors and highly sensitive molecular diagnostics.

What We Offer: Comprehensive Enzyme-Nucleic Acid Conjugation Services

Creative Enzymes provides a full spectrum of enzyme-nucleic acid conjugation solutions for both research and commercial applications. Our offerings include:

Development of Enzyme-Linked Oligonucleotides

Custom synthesis of ssDNA, dsDNA, or RNA probes conjugated to signal enzymes.
Tailored conjugates for direct detection, sandwich assays, or hybridization-based platforms.

Antibody-Enzyme Conjugate (AEC) Preparation

Complementary service for nucleic acid detection systems that require antibody recognition.
Supports multiplexed detection platforms by combining nucleic acid probes with antibody-enzyme complexes.

Protein Expression, Purification, and Conjugation

Production of high-purity enzymes (HRP, AP, glucose oxidase, etc.) suitable for conjugation.
Controlled chemical or enzymatic conjugation with nucleic acid probes to ensure functional integrity.

Characterization of Target Conjugates

Analytical validation including spectrophotometry, SDS-PAGE, electrophoretic mobility shift assays, and activity assays.
Ensures consistent quality, activity, and reproducibility for downstream applications.

Custom Signal Amplification Solutions

Design of advanced probe-enzyme systems for amplified detection in biosensors or diagnostic assays.
Optimization of conjugate ratios, linker chemistry, and detection modalities.

Get a quote

Service Workflow

Workflow of enzyme conjugation with nucleic acids service

Technical Capabilities and Conjugation Strategies

Creative Enzymes combines advanced biochemistry techniques with practical experience to deliver reliable enzyme-nucleic acid conjugates. Key technical details include:

Direct Conjugation Methods	Chemical Crosslinking: Using bifunctional linkers that covalently bond enzyme amino groups to nucleic acid functional groups.
	Polymer-Mediated Conjugation: Enzymes are modified with polymers carrying multiple primary amino groups, allowing multivalent DNA binding.
	Click Chemistry: Bioorthogonal reactions providing highly specific, mild, and efficient conjugation.
Indirect Conjugation Methods	Biotin-Streptavidin Systems: Biotin-labeled nucleic acids detected via streptavidin-HRP or streptavidin-AP complexes.
Indirect Conjugation Methods	Hapten-Antibody Systems: Modified nucleic acids with haptens recognized by enzyme-labeled antibodies for signal generation.

Why Choose Us: Advantages of Creative Enzymes' Conjugation Services

Multidisciplinary Conjugation Expertise

Our team integrates protein chemistry, nucleic acid chemistry, and assay development across direct and indirect conjugation platforms.

High-Quality, Application-Specific Customization

We design tailored conjugates with flexible enzyme and linker choices to match specific assays and research objectives.

Advanced Instrumentation and Technical Capabilities

State-of-the-art chromatography and analytical tools ensure precise conjugation, validation, and performance consistency.

Rigorous Quality Control and Characterization

Enzyme activity, nucleic acid integrity, and conjugate stability are strictly monitored with comprehensive analytical reporting.

Proven Signal Amplification Strategies

We optimize conjugates for enhanced sensitivity in genosensors, electrochemical assays, and low-abundance target detection.

Comprehensive Technical and Application Support

Our experts provide end-to-end support, from assay design and optimization to troubleshooting and scale-up.

Case Studies: Real-World Applications of Enzyme-Nucleic Acid Conjugates

Case 1: Horseradish Peroxidase-Encapsulated DNA Nanoflowers

A versatile colorimetric biosensing platform was developed for rapid and sensitive detection of cancer-derived exosomes using horseradish peroxidase–encapsulated DNA nanoflowers (HRP-DFs). HRP was directly immobilized within DNA nanoflowers during rolling circle amplification, providing both biorecognition and signal generation. HRP-DFs encoded with CD63 aptamer sequences specifically captured exosomes and formed sandwich complexes with magnetic beads for easy separation. The bound HRP catalyzed a visible color change of ABTS in the presence of hydrogen peroxide, enabling naked-eye and spectrometric detection. The platform achieved high sensitivity, low detection limits, strong anti-interference capability, and good reproducibility for exosome analysis.

Horseradish peroxidase-encapsulated DNA nanoflowers are an innovative signal generation tag for a colorimetric biosensor Figure 2. Schematic illustration of colorimetric biosensor for the detection of target exosomes on cholesterol -modified DNA probes-conjugated magnetic bead (MB) and CD63 aptamer-encoded HRP-DF, based on catalyzing ABTS-H₂O₂ color-changing system. (Zheng et al., 2022)

Case 2: Novel Signal Amplification Approach for HRP-Based Colorimetric Genosensors Using DNA Binding Protein Tags

Sensitive DNA detection is increasingly important for applications in disease diagnosis, food safety, and biosecurity. Traditional hybridization assays rely on ssDNA probes directly conjugated to enzymes such as HRP or on nanoparticle-based amplification strategies, which can present limitations in sensitivity and flexibility. This study introduces a novel DNA sensing platform based on an HRP–DNA-binding protein tag (scCro) conjugate combined with a hybrid ssDNA–dsDNA detection probe. The probe contains multiple scCro binding sites, enabling efficient signal amplification. Using this approach, detection sensitivity and limit of detection for high-risk HPV16 were improved more than threefold compared with conventional ssDNA–HRP conjugates, demonstrating its potential for point-of-care diagnostics.

Design and characterization of DNA detection probes with scCro DNA binding sites Figure 3. EMSA assay for the binding of the HRP–scCro conjugate to the HPV16 detection probes with one and three DNA binding sites. Bands with asterisks correspond to protein–DNA complexes. (Aktas et al., 2015)

FAQs: Comprehensive Guidance on Enzyme-Nucleic Acid Conjugation

Q: What types of enzymes can be conjugated to nucleic acids?

A: Common enzymes include horseradish peroxidase (HRP), alkaline phosphatase (AP), glucose oxidase, and luciferase. We also accommodate custom enzymes depending on assay requirements.
Q: Can both DNA and RNA be conjugated to enzymes?

A: Yes. Both single-stranded and double-stranded DNA, as well as RNA molecules, can be conjugated using direct or indirect methods. The choice depends on stability and assay design considerations.
Q: What is the difference between direct and indirect conjugation?

A: Direct conjugation involves covalent attachment of the enzyme to the nucleic acid, providing a single-molecule detection tool. Indirect conjugation uses intermediate recognition systems (biotin-streptavidin or hapten-antibody) to amplify the signal.
Q: How is the quality of the conjugates verified?

A: Conjugates are validated through spectrophotometric analysis, SDS-PAGE, electrophoretic mobility shift assays, and enzyme activity assays to ensure functionality, purity, and stability.
Q: Are your conjugates suitable for commercial diagnostic assays?

A: Yes. We adhere to stringent quality standards and provide detailed characterization data suitable for research, preclinical, and commercial assay development.
Q: Can you assist in assay optimization using these conjugates?

A: Absolutely. Our team provides technical support for optimizing assay conditions, conjugate ratios, and detection methods to maximize sensitivity and reproducibility.

References:

Aktas GB, Skouridou V, Masip L. Novel signal amplification approach for HRP-based colorimetric genosensors using DNA binding protein tags. Biosensors and Bioelectronics. 2015;74:1005-1010. doi:10.1016/j.bios.2015.07.077
Zeng R, Wang J, Wang Q, Tang D, Lin Y. Horseradish peroxidase-encapsulated DNA nanoflowers: An innovative signal-generation tag for colorimetric biosensor. Talanta. 2021;221:121600. doi:10.1016/j.talanta.2020.121600

First Name:

Last Name:

Email *

Phone Number:

Company/Institution:

Country or Region:

Quantity:

Services & Products of Interested

Project Description:

For research and industrial use only. Not intended for personal medicinal use. Certain food-grade products are suitable for formulation development in food and related applications.

Services

Online Inquiry