Conformational distributions of protease-serpin complexes: a partially translocated complex

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• Serpins are crucial regulators of serine proteases.
• They form metastable covalent complexes, trapping target proteases.
• This involves reactive center loop cleavage and protease translocation.

Purpose of the Study:

• To investigate the conformational dynamics of protease-serpin covalent complexes.
• To understand the role of protease stability and labeling in complex formation.
• To determine if full protease translocation is essential for metastable complex formation.

Main Methods:

• Single-pair Förster resonance energy transfer (spFRET) was employed.
• Conformational distributions of bovine trypsin (BTryp) and rat trypsin (RTryp) complexes with alpha(1)-proteinase inhibitor (alpha(1)PI) were measured.
• Protease active site stability was modulated via mutagenesis.

Main Results:

• BTryp complexes showed full translocation (E(full)I*), but with mobility.
• Most RTryp complexes also exhibited E(full)I*, with narrower distributions.
• RTryp complexes with cationic dyes showed two conformations: E(full)I* and partial translocation (E(part)I*).
• Active site destabilization increased the E(full)I* population.

Conclusions:

• Protease stability significantly impacts conformational distributions in serpin complexes.
• Interactions between proteases and labels can influence translocation.
• Full translocation of the protease is not a prerequisite for forming metastable serpin-protease complexes.