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Functional cooperation of the mitochondrial processing peptidase subunits
P Luciano1, S Geoffroy, A Brandt
1Laboratoire d'Ingéniérie des Systèmes Macromoléculaires, Institut de Biologie Structurale et Microbiologie, CNRS, 31 chemin Joseph Aiguier, Marseille Cedex 20, 13402, France.
Journal of Molecular Biology
|September 23, 1997
Summary
Investigating the mitochondrial processing peptidase (MPP), alanine insertions revealed key residues for beta-MPP activity and subunit interaction. The alpha-subunit binds substrates, suggesting it presents them to the catalytic site.
Area of Science:
- Biochemistry
- Molecular Biology
- Mitochondrial Biology
Background:
- Mitochondrial processing peptidase (MPP) is crucial for protein maturation.
- MPP is a heterodimer composed of alpha-MPP and beta-MPP subunits.
- Understanding subunit interactions and active site domains is vital for MPP function.
Purpose of the Study:
- To investigate domains critical for the activity of the heterodimeric mitochondrial processing peptidase (MPP).
- To elucidate the roles of specific residues in the beta-subunit (beta-MPP) in MPP activity and alpha-subunit (alpha-MPP) interaction.
- To determine the substrate-binding role of the alpha-subunit.
Main Methods:
- Site-directed mutagenesis by alanine insertion in beta-MPP.
- Co-expression and affinity purification of MPP subunits.
- Cross-linking studies.
- Surface plasmon resonance analysis.
Main Results:
- Alanine insertion at Glu70, Ser114, Lys215, and Ser314 in beta-MPP abolished or impaired MPP activity.
- Mutations at Glu70 and Lys215 disrupted alpha-MPP and beta-MPP association.
- Alpha-MPP alone bound precursor proteins and peptide substrates efficiently.
- The HXXEHX76H region, central region (Lys215), and carboxy-terminal region (Ser314) of beta-MPP are critical for activity and/or subunit interaction.
Conclusions:
- The HXXEHX76H region of beta-MPP is essential for active site conformation and potentially alpha-MPP interaction.
- The central region of beta-MPP (around Lys215) mediates alpha-MPP interaction.
- The carboxy-terminal region of beta-MPP (around Ser314) is important for catalysis.
- Alpha-MPP is primarily responsible for binding mitochondrial presequences before their processing by MPP.