Structural basis of protein substrate processing by human mitochondrial high-temperature requirement A2 protease

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• Human high-temperature requirement A2 (HtrA2) is a trimeric protease vital for cellular stress response by degrading misfolded proteins.
• Dysfunctional HtrA2 is implicated in neurodegenerative disorders.
• Previous studies revealed HtrA2 activation via peptide ligands but lacked insight into protein substrate interactions.

Purpose of the Study:

• To investigate the molecular mechanisms of HtrA2 binding to protein substrates.
• To elucidate the role of distinct conformational states in protein substrate processing.
• To understand how HtrA2's trimeric structure facilitates its protease function.

Main Methods:

• Solution-based Nuclear Magnetic Resonance (NMR) spectroscopy.
• Utilized a model protein substrate mimic (drkN SH3-PDZbm).
• Investigated thermodynamic, kinetic, and structural aspects of the interaction.

Main Results:

• HtrA2 binds protein substrates via a two-pronged interaction: a C-terminal PDZ-binding motif and a central hydrophobic region.
• Binding occurs preferentially with an unfolded ensemble of substrate molecules.
• Multivalent substrate binding to the HtrA2 trimer significantly enhances its catalytic activity.

Conclusions:

• Binding avidity is a key regulator of HtrA2 substrate processing.
• The trimeric architecture of HtrA2 is essential for its function as a stress-protective protease.
• This study provides a comprehensive understanding of HtrA2-protein substrate interactions, relevant to neurodegenerative disease mechanisms.