PACTS-Assisted Thermal Proteome Profiling for Use in Identifying Peptide-Interacting Proteins

Area of Science:

• Biochemistry
• Proteomics
• Molecular Biology

Background:

• Bioactive peptides are crucial in biological processes.
• Traditional methods for identifying peptide-interacting proteins often require peptide modification, potentially causing false positives.
• Changes in protein abundance upon peptide-ligand interaction offer a signature for identifying these interactions.

Purpose of the Study:

• To develop a novel strategy, Peptide-ligand-induced Abundance Change of proTeinS (PACTS)-assisted Thermal Proteome Profiling (TPP), for identifying peptide-interacting proteins.
• To demonstrate the efficacy of PACTS-assisted TPP using known interactions and large-scale profiling.
• To investigate the interactome of amyloid beta (Aβ) 1-42 in THP-1 cells and its implications for Alzheimer's disease (AD).

Main Methods:

• Development of the PACTS-assisted TPP strategy based on peptide-ligand-induced changes in protein abundance.
• Validation of the method using the 3-phosphoinositide-dependent protein kinase 1 (PDPK1) and PDPK1-interacting fragment (PIF) interaction.
• Application of PACTS-assisted TPP to profile Aβ1-42 interactome in THP-1 cells.

Main Results:

• The PACTS-assisted TPP strategy successfully identified peptide-interacting proteins based on abundance changes.
• The interaction of PDPK1 and PIF was confirmed.
• Large-scale profiling of PIF identified interacting proteins.
• The Aβ1-42 interactome in THP-1 cells was described, identifying 103 interacting proteins.
• Aβ1-42 was found to directly interact with and inhibit fatty acid synthase.

Conclusions:

• PACTS-assisted TPP is an efficient method for identifying peptide-interacting proteins without requiring peptide modification.
• The identified Aβ-interacting proteins, including fatty acid synthase, offer new insights into fatty acid metabolic disorders in Alzheimer's disease.
• This approach provides valuable resources for AD research.