The conformational landscape of a serpin N-terminal subdomain facilitates folding and in-cell quality control.
Upneet KaurKyle C KihnHaiping KeWeiwei KuoLila M GieraschDaniel N HebertPatrick L WintrodeDaniel DeredgeAnne GershensonPublished in: bioRxiv : the preprint server for biology (2023)
Many multi-domain proteins including the serpin family of serine protease inhibitors contain non-sequential domains composed of regions that are far apart in sequence. Because proteins are translated vectorially from N-to C-terminus, such domains pose a particular challenge: how to balance the conformational lability necessary to form productive interactions between early and late translated regions while avoiding aggregation. This balance is mediated by the protein sequence properties and the interactions of the folding protein with the cellular quality control machinery. For serpins, particularly α 1 -antitrypsin (AAT), mutations often lead to polymer accumulation in cells and consequent disease suggesting that the lability/aggregation balance is especially precarious. Therefore, we investigated the properties of progressively longer AAT N-terminal fragments in solution and in cells. The N-terminal subdomain, residues 1-190 (AAT190), is monomeric in solution and efficiently degraded in cells. More ý-rich fragments, 1-290 and 1-323, form small oligomers in solution, but are still efficiently degraded, and even the polymerization promoting Siiyama (S53F) mutation did not significantly affect fragment degradation. In vitro, the AAT190 region is among the last regions incorporated into the final structure. Hydrogen-deuterium exchange mass spectrometry and enhanced sampling molecular dynamics simulations show that AAT190 has a broad, dynamic conformational ensemble that helps protect one particularly aggregation prone ý-strand from solvent. These AAT190 dynamics result in transient exposure of sequences that are buried in folded, full-length AAT, which may provide important recognition sites for the cellular quality control machinery and facilitate degradation and, under favorable conditions, reduce the likelihood of polymerization.
Keyphrases
- quality control
- molecular dynamics simulations
- induced apoptosis
- single molecule
- cell cycle arrest
- mass spectrometry
- molecular docking
- molecular dynamics
- cell death
- endoplasmic reticulum stress
- stem cells
- amino acid
- bone marrow
- small molecule
- oxidative stress
- mesenchymal stem cells
- ionic liquid
- cell proliferation
- pi k akt
- blood brain barrier