Label-Free Deacetylase Assays

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Epigenetic modifications of histones such as acetylation cause conformational changes in the chromatin structure, thereby regulating transcriptional activity. Histone acetyltransferases (HATs) and histone deacetylases (HDACs) are enzymes responsible for such post-translational modification of core histones. HDACs remove acetyl groups from lysine residues and are separated into four categories: Class I (HDAC1, 2, 3 & 8), Class II (HDAC4-9), Class III (SIRT1-7) and Class IV (HDAC11). Class I, II, and IV are zinc-dependent deacetylases while class III is NAD-dependent. HDACs have been shown to play a significant role in the development of neurodegenerative and metabolic disorders. Thus, HDAC modulators could provide viable treatment options and are currently targeted for drug discovery.

What We Offer

SAMDI Tech offers a plug-and-play format for quantitative analysis of deacetylase activity using native substrates. Native peptide and protein substrates can be captured and purified onto SAMDI 384 or 1536 biochip arrays for analysis by mass spectrometry. No need for cumbersome fluorescent-labeled substrates that can interfere with activity and lead to increase false positive rates. SAMDI has the capability to monitor all acetylation states simultaneously in a single assay.

SAMDI Advantages

  • Fast - Eliminates labels and reduces assay development time
  • Efficient - Multi-analyte detection and 10,000s of samples analyzed per day
  • Quantitative - Measures enzyme kinetics, mechanism of action, and IC50s

Types of Deacetylase Assays

  • HDACs 1-11/li>
  • Sirtuins 1, 2, 3, 5, 6, 7

Assay Format
SAMDI surface chemistry for purification of peptide analytes prior to mass spectrometry. Deacetylase is reacted with peptide substrate and both substrate and product are captured to SAMDI arrays.

SAMDI Mass Spectrometry
SAMDI mass spectrum of Deacetylation event where 30% of substrate is converted to product.

Assay Development
SAMDI is quantitative and can measure enzyme kinetics including: Vo vs [Enzyme], enzyme linearity, Km determination, plate uniformity, and IC50 analysis.