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Updated: May 25, 2026
12:07
Sequence-specific Labeling of Nucleic Acids and Proteins with Methyltransferases and Cofactor Analogues
Published on: November 22, 2014
Encoding substrates with mass tags to resolve stereospecific reactions using Nimzyme
Kai Deng1, Kevin W George, Wolfgang Reindl
1Joint BioEnergy Institute, Physical Biosciences Division, Lawrence Berkeley National Laboratory, Emeryville, CA 94608, USA.
Rapid Communications in Mass Spectrometry : RCM
|February 14, 2012
Summary
This study introduces a modified Nimzyme assay using unique mass tags to rapidly screen stereospecific glycan reactions. This method allows simultaneous resolution of different stereoisomers from crude samples, advancing high-throughput enzyme analysis.
Area of Science:
- Glycobiology
- Enzymology
- Analytical Chemistry
Background:
- The nanostructure-initiator mass spectrometry (Nimzyme) assay offers rapid screening of glycan-modifying enzymes.
- Existing Nimzyme methods struggle to resolve stereospecific reactions common in glycobiology.
- Higher-throughput methods for stereospecific reactions are needed, especially from crude samples.
Purpose of the Study:
- To adapt the Nimzyme assay for resolving stereospecific reactions.
- To enable high-throughput screening of glycan-modifying enzymes with stereochemical resolution.
- To utilize unique mass tags for encoding substrate identity within the Nimzyme assay.
Main Methods:
- Employed nanostructure-initiator mass spectrometry (NIMS) with an accurate mass tagging approach.
- Reactants were tagged with unique perfluorinated tails for mass spectrometric analysis.
- Utilized conventional MALDI-TOF instrumentation for mass spectrometry.
Main Results:
- Successfully resolved stereospecific reaction pathways of maltose, lactose, and cellobiose simultaneously.
- Demonstrated the assay's capability with unique fluorous tags on reactants and products.
- Showed that the assay is effective with both purified enzymes and crude cell lysates.
Conclusions:
- The modified Nimzyme assay with accurate mass tagging enables rapid screening of targeted stereospecific reactions.
- This approach is suitable for high-throughput screening and functional annotation of glycan-modifying enzymes.
- The method facilitates the analysis of complex biological samples, including crude cell lysates.
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