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AMP2/GMP2 Assay for Screening Ligases, Synthetases and Phosphodiesterases

Ligase & Synthetase Assay - A Transcreener AMP2/GMP2 Assay Application

Transcreener AMP2/GMP2 Assay directly measures AMP/GMP produced by ligases & synthetases. These AMP/GMP measurements effectively determine the activity of the ligase & synthetase enzyme. The assay provides a powerful tool to screen compound libraries for ligase/synthetase modulators to help find new therapies for disease.

The kit comes complete with the detection reagents required to measure activity. Ligase/Synthetase enzyme is not included in the inhibitor screening asay kit. Please contact us for questions related to acquiring the enzyme.

How Does This Ligase & Synthetase Assay Work?

The Transcreener Ligase & Synthetase Assay determines ligase or synthetase enzyme activity by directly measuring the AMP/GMP formed by the enzyme using the Transcreener AMP2/GMP2 assay. By detecting AMP/GMP output, the assay provides a universal method to assess the activity of any AMP/GMP-producing enzyme in real-time. 

Transcreener Ligase & Synthetase Assay technology uses a simple but highly effective method that consists of an antibody selective to AMP/GMP and a far-red fluorescent tracer. AMP/GMP produced in the reaction competes with the tracer changing the fluorescent properties and providing fluorescent readout.

The ligase & synthetase assay is available with FP and TR-FRET. It is a simple mix-and-read format. Perform your enzyme reaction, add the detection reagent, and measure. The simplicity of the system yields robust results that also makes it extremely amiable to HTS.

Example Assay Target Applications

 Enzyme Target
LigasesUb Ligases
SUMO Ligases
DNA ligases
RNA Ligases
SynthetasesAcetyl CoA Synthetase
Amino-Acyl tRNA Synthetases
NAD Synthetase

Direct Detection of AMP/GMP to Measure Ligase & Synthetase Enzymatic Activity

Fluorescent Polarization (FP)

Transcreener Ligase & Synthetase Assay Schematic

Time-Resolved Förster Resonance Energy Transfer (TR-FRET)

Ligase & Synthetase TR-FRET Readout

Applications

  • Measure Enzymatic Activity of Ligases & Synthetases 
  • Screen Compound Libraries for Ligase/Synthetase Modulators
  • Quantify Inhibitor Potency
  • Inhibitor Selectivity Profiling
  • Measure Drug-Target Residence Time

Features

  • Direct detection of unlabeled AMP or GMP
  • Easy to use, homogenous, one-step format
  • Robust Assay Z'>0.7 under initial velocity reactions
  • Far-red fluorescent readouts minimize compound interference
  • A safe, non-radioactive assay technique
  • Wide substrate concentration range: 1 μM to 1000 μM

Easy-to-Use, Mix-and-Read, HTS-Ready Assay

Run your enzyme reaction, add Transcreener reagents, and read your plates. The ligase & synthetase assay is compatible with 96, 384, and 1536-well formats. 

Transcreener Mix and Read Assay

Continuous Kinetic Mode Compatible

Transcreener Assays can be used in kinetic mode, allowing the assay to be used in applications not possible with endpoint only assay methods.

Measure Residence Times

Measure the residence times for inhibitors with a simple biochemical assay to characterize leads with the greatest potential to become drugs.

Streamlined Assay Development

Using the assay in kinetic mode reduces the number of experiments required to optimize conditions for a new enzyme.

Ligase & Synthetase Assay Services

Interested in moving your program forward, but don't want to bring an assay in-house? Our scientists can help! BellBrook scientists will use their extensive biochemistry and enzymology expertise to work with you to accelerate your ligase/synthetase program. 

Assay Development Services

Lead Discovery Services Include:

  • Inhibitor Screening - To identify or confirm activity with the target.
  • Inhibitor Potency Profiling - Dose-response with target and/or related proteins. Fast IC50 results.
  • Residence Time Measurements - Determination of koff using 'jump dilution' enzymatic assay method.
  • Mechanism of Action Studies - Kinetic analysis to define the mode of inhibition.

Contact Us to Learn More

Please fill out the form below. We will respond quickly to get the conversation moving and learn how we can help. We keep things discrete, confidential, and professional. 

Far Red FP & TR-FRET Readouts Validated on Major Multimode Readers

Supplier Instrument FP Assays
TR-FRET Assays
bioteklogo TriStar²S LB 942 in review validated
Mithras² LB 943 in review validated
bioteklogo Cytation™ 5 validated validated
Cytation™ 3 validated validated
Cytation™ 1 validated validated
Synergy™ H1 validated validated
Synergy™ 2/H4/4 validated validated
Synergy™ HTX not capable not capable
Synergy™ Neo 2 validated validated
BMGLABTECH Logo POLARstar® Omega validated validated
FLUOstar® Omega not capable validated
PHERAstar® FSX validated validated
PHERAstar® Plus/FS validated validated
CLARIOstar® /Plus validated validated
VANTAstar validated validated
MDS AT logo Analyst® GT/HT validated validated
Gemini® XPS/EM not capable not capable
SpectraMax® M2/M2e not capable not capable
SpectraMax® M5/M5e/FlexStation® 3 not capable validated
SpectraMax® Paradigm validated contact us
SpectraMax® iD3/iD5 in review validated
perkinElmerLogo EnVision®/EnVision® Xcite validated validated
tecanLogo Infinite® M1000/M1000Pro/Safire2™ validated validated
Infinite® M200 not capable not capable
Infinite® F500 validated validated
Infinite® F200/Ultraevolution contact us contact us
Spark™ 10M validated validated

What's Included

What You Will Need

Transcreener AMP²/GMP² Assay Kits

* For custom or bulk orders (over 100,000 wells) please contact us (info@bellbrooklabs.com) for a quote.

The Role of Ligases & Synthetases As Therapeutic Targets

The physiological role of ligases and synthetases is dependent on the respective family. Here, we explore roles of aminoacyl-tRNA synthetases as therapeutic targets.

Aminoacyl-tRNA synthetases ligate amino acids to tRNAs and translate the genetic code for protein synthesis. Because of their role in protein synthesis, these synthetases are needed for cell survival, and therefore are popular drug targets. Diseases they are targeted for include infectious diseases, cancers, and inflammatory diseases (Kim et al, 2020).  

The Transcreener AMP2/GMP2 Ligase & Synthetase Assay is an excellent tool for researchers examining the therapeutic effects of ligases and synthetases.

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