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Improved drug target deconvolution with PISA-DIA using an extended, overlapping temperature gradient.

Samantha J Emery-Corbin1,2, Jumana M Yousef1,2, Subash Adhikari1,2

  • 1Advanced Technology and Biology Division, the Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria, Australia.

Proteomics
|May 20, 2024
PubMed
Summary
This summary is machine-generated.

We developed a novel Proteome Integral Solubility Alteration via label-free DIA (PISA-DIA) method for drug target deconvolution. This approach enhances proteome coverage and sensitivity, improving target identification for novel compounds.

Keywords:
Proteome Integral Solubility Alterationdata independent acquisition (DIA)mass spectrometryprotein stabilityproteomicstarget engagementthermal proteome profiling

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Area of Science:

  • Biochemistry
  • Proteomics
  • Pharmacology

Background:

  • Thermal proteome profiling (TPP) is crucial for drug target deconvolution.
  • Traditional TPP methods often rely on data-dependent acquisition (DDA) and can have limitations in proteome depth and missing data.
  • Data-independent acquisition mass spectrometry (DIA-MS) offers improved proteome coverage and reduced missingness.

Purpose of the Study:

  • To introduce and validate a novel experimental design for TPP using DIA-MS, termed PISA-DIA.
  • To demonstrate the enhanced proteome coverage and sensitivity of PISA-DIA compared to traditional TPP methods.
  • To establish the quantitative and statistical rigor of PISA-DIA for drug target deconvolution.

Main Methods:

  • Development of an extended PISA-DIA workflow utilizing multiple overlapping thermal gradients.
  • Application of label-free DIA-MS for proteome analysis.
  • Validation using A-1331852, a specific inhibitor of BCL-xL, a protein with a high melting temperature.

Main Results:

  • The PISA-DIA approach achieved superior proteome coverage compared to TPP methods requiring DDA-MS and tandem-mass tags (TMT).
  • The extended multiple gradient PISA-DIA workflow successfully identified BCL-xL, a high-melting-temperature target.
  • The method demonstrated quantitative and statistical rigor suitable for drug target deconvolution.

Conclusions:

  • The novel overlapping gradient PISA-DIA-MS approach is ideal for unbiased drug target deconvolution.
  • This method spans a large temperature range, minimizing target dropout and increasing the likelihood of resolving protein targets of novel compounds.
  • PISA-DIA offers enhanced proteome coverage and sensitivity for TPP applications.