Aspartate aminotransferase is potently inhibited by copper complexes: Exploring copper complex-binding proteome

Area of Science:

• Biochemistry
• Proteomics
• Cellular Metabolism

Background:

• Copper complexes can interact with cellular components, potentially influencing metabolism.
• The existence of a specific copper complex-binding proteome is hypothesized.
• Understanding these interactions is crucial for cellular health and disease research.

Purpose of the Study:

• To explore the copper complex-binding proteome in primary rat hepatocytes.
• To identify proteins that bind to copper complexes.
• To investigate the inhibitory effects of copper complexes on specific enzymes.

Main Methods:

• Copper chelating ion-immobilized affinity chromatography (Cu-IMAC) and mass spectrometry were employed.
• Purification of high-abundance proteins using SP- and Q-Sepharose Fast Flow columns.
• Enzyme inhibition assays and fluorescence titration were used to study protein-copper complex interactions.

Main Results:

• 97 putative copper-binding proteins were identified.
• Aspartate aminotransferase (AST), malate dehydrogenase (MDH), and catalase (CAT) were purified and characterized.
• Schiff-based copper complexes 6 and 7 potently inhibited AST (IC50 values of 3.6 and 7.2 μM) with high selectivity.
• Copper complex 6 entered HepG2 cells and inhibited intracellular AST activity.

Conclusions:

• A set of 97 copper-binding proteins was identified, providing insights into cellular copper interactions.
• Specific copper complexes demonstrate potent and selective inhibition of AST.
• Copper complex 6 shows potential for targeting intracellular AST activity, suggesting therapeutic possibilities.