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  5. Enzyme Activity Measurement for Hydrolases Acting on Ester Bonds

Enzyme Activity Measurement for Hydrolases Acting on Ester Bonds

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Creative Enzymes is now ranked among the top leaders of the biochemical service industry. Over the past several years, we have earned the trust of tens of thousands of clients by consistently delivering test results of exceptional quality. Our success is founded on a strong commitment to research and innovation: we maintain a highly skilled team of enzymologists and continuously invest in advanced assay development. Combined with our in-house analytical facilities, we have accumulated extensive expertise in testing hydrolases with unparalleled accuracy and reliability.

Understanding Hydrolases Acting on Ester Bonds

Hydrolases are enzymes that catalyze hydrolysis reactions and can be classified into several subclasses. The subclass EC 3.1 includes enzymes that act on ester bonds—commonly known as esterases. These enzymes hydrolyze esters, thioesters, and phosphates, playing vital roles in genetics, cell signaling, metabolism, and structural biology.

The importance of ester bond hydrolases is reflected in their wide-ranging applications:

  • Industrial Applications : Used in detergents and cleaning agents for efficient stain removal under mild conditions, and in waste management for degrading organic residues.
  • Medical and Pharmaceutical Research : Enzymes such as acetylcholinesterase regulate neurotransmission by breaking down acetylcholine.
  • Biotechnology : Many hydrolases process nucleic acids, catalyzing cleavage of phosphoester and phosphodiester bonds—crucial for DNA/RNA analysis and synthesis.
  • Environmental Protection : Applied in bioremediation and eco-friendly degradation processes.

Despite significant advances, the catalytic mechanisms of many esterases remain incompletely understood. Accurate enzyme activity measurement is therefore essential for research and application development. Creative Enzymes provides comprehensive solutions to meet this need.

Comprehensive Service Offerings

Workflow and Process

Workflow of enzyme activity measurement for hydrolases acting on ester bonds

Our Service Offerings

At Creative Enzymes, we offer a complete suite of enzyme activity measurement services for hydrolases acting on ester bonds, including:

Standardized Enzyme Activity Assays

Using spectrophotometric, chromatographic, and fluorometric techniques to deliver reproducible and precise activity results.

Custom Enzyme Assays

Tailored solutions for unique research needs, ensuring accurate measurement under specific conditions such as pH, temperature, inhibitors, or substrates.

Kinetic Characterization

Determination of kinetic parameters ( K m , V max , k cat ) to reveal catalytic efficiency and mechanism.

Comparative Activity Profiling

Benchmarking enzyme variants or engineered hydrolases to identify the most effective candidates.

Substrate Specificity Testing

Comprehensive testing across a variety of natural and synthetic substrates.

Enzyme Classes Covered

Structure of ubiquitin carboxy-terminal hydrolase L1 (UCHL1, PDB: 2etl) Carboxylic and Thioester Hydrolases
Structure of calcineurin heterodimer composed of the catalytic (PPP3CA) and regulatory (PPP3R1) subunits Phosphoric Ester Hydrolases
Structure of phosphonoacetate hydrolase (PDB:1e16), zinc ions and phosphonoformate are shown in the active site Nucleases

Contact Our Team

Why Choose Creative Enzymes

Expertise

Decades of combined enzymology experience, particularly in esterases and nucleic acid-related hydrolases.

Cutting-edge Technologies

Advanced analytical platforms ensure reliable and highly reproducible results.

Customization

Assays adapted to your specific research requirements.

Efficiency

Timely delivery of results with full technical support.

Global Trust

Recognized and praised by thousands of clients worldwide for accuracy and professionalism.

Efficient Delivery

Streamlined project management for timely results.

Case Studies and Real-World Applications

Case 1: Structural and Functional Insights into an Atypical Carboxylesterase

Researchers characterized a novel carboxylesterase from Bacillus coagulans (BCE), notable for its high enantioselectivity toward solketal esters, enabling efficient production of enantiopure (S)-IPG—an important precursor for β-blockers, glycerophospholipids, and prostaglandins. BCE remains catalytically active up to 65 °C. Crystal structures of BCE in apo- and glycerol-bound forms, combined with docking studies, revealed that enantioselectivity arises from improved stabilization of the S-enantiomer intermediate. Interestingly, BCE adopts a lipase-like fold with a "lid" domain, though it lacks lipase-like interfacial activation. This unique enzyme expands opportunities for cost-effective biocatalysis in pharmaceutical intermediate synthesis.

Biochemical activity of Bacillus coagulans carboxylesterase (BCE) Figure 1. BCE activity toward p -nitrophenyl esters. Relative activity refers to the activity in the presence of p -nitrophenyl caproate (100%). (De Vitis et al ., 2018)

Case 2: Design of Donepezil Analogs as Potent AChE Inhibitors

To explore new treatments for Alzheimer's disease, researchers designed and synthesized eco-friendly donepezil analogs and evaluated their acetylcholinesterase (AChE) inhibitory activity. Substituting the phenyl ring of donepezil with a pyridine moiety produced equally potent inhibitors against electric eel AChE. Kinetic studies revealed that the lead compound exhibited a mixed inhibition mode, paralleling donepezil's dual-binding behavior. Molecular modeling further indicated additional residue interactions in the human AChE active site, suggesting potential improvements in binding efficiency. These findings highlight promising structural modifications for next-generation AChE inhibitors in Alzheimer's therapy.

Inhibitory potency profile of donepezil-based analog 2 against Acetylcholinesterase (AChE) Figure 2. Lineweaver-Burke double reciprocal plot for Analog 2. (Makarian et al ., 2022)

FAQs

  • Q: What information should I provide when requesting enzyme activity measurement?

    A: We recommend including the type of enzyme, source, sample condition, research purpose, and specific challenges you face. This enables us to tailor the assay precisely.

  • Q: How long does a typical assay project take?

    A: Turnaround times depend on project complexity but generally range from two to four weeks. Custom assays may require additional development time.

  • Q: Can you test engineered or mutant hydrolases?

    A: Yes, we routinely test both natural and engineered variants, providing comparative activity profiles.

  • Q: What assay formats do you offer?

    A: We provide spectrophotometric, fluorometric, and chromatographic assays, with the flexibility to integrate other techniques when necessary.

  • Q: Do you assist with data interpretation?

    A: Absolutely. Our team provides not just raw data, but also analytical insights to help guide your next research step.

References:

  1. De Vitis V, Nakhnoukh C, Pinto A, et al . A stereospecific carboxyl esterase from Bacillus coagulans hosting nonlipase activity within a lipase‐like fold. The FEBS Journal . 2018;285(5):903-914. doi:10.1111/febs.14368
  2. Makarian M, Gonzalez M, Salvador SM, Lorzadeh S, Hudson PK, Pecic S. Synthesis, kinetic evaluation and molecular docking studies of donepezil-based acetylcholinesterase inhibitors. Journal of Molecular Structure . 2022;1247:131425. doi:10.1016/j.molstruc.2021.131425
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