GPCR targeting of E3 ubiquitin ligase MDM2 by inactive β-arrestin

Area of Science:

• Molecular and Cellular Biology
• Biochemistry
• Structural Biology

Background:

• E3 ubiquitin ligase Mdm2 targets β-arrestins for ubiquitination, a key step in G protein-coupled receptor (GPCR) internalization.
• The molecular mechanism and structural basis of the β-arrestin-Mdm2 interaction remain largely unknown.

Purpose of the Study:

• To elucidate the molecular architecture of the β-arrestin-Mdm2 complex.
• To identify the β-arrestin-binding region (ABR) on Mdm2 and determine its structural role in GPCR internalization.

Main Methods:

• Crystal structure determination of β-arrestin1 in complex with the Mdm2_ABR peptide.
• Hydrogen/deuterium exchange mass spectrometry (HDX-MS) to analyze conformational changes upon Mdm2 binding.

Main Results:

• The crystal structure reveals that Mdm2_ABR acidic residues bind to the concave N-domain of β-arrestin1, while the β-arrestin1 C-tail remains associated with the N-domain.
• Mdm2 binding occurs to an inactive β-arrestin1 conformation, distinct from the active conformation induced by phosphorylated GPCR C-tails.
• HDX-MS data show Mdm2 binding increases interdomain dynamics and disrupts β-arrestin1 oligomerization.

Conclusions:

• Mdm2 binds to the inactive state of β-arrestin1, suggesting a mechanism where GPCR C-tail binding may displace Mdm2.
• The structural and dynamic insights into the β-arrestin-Mdm2 interaction provide a mechanistic understanding of how Mdm2 facilitates GPCR internalization.