Mapping ERK2-MKP3 binding interfaces by hydrogen/deuterium exchange mass spectrometry

Area of Science:

• Cellular signaling
• Protein-protein interactions
• Enzymology

Background:

• ERK2 (extracellular signal-regulated kinase 2) is a key regulator of cell growth and differentiation within the MAPK pathway.
• MKP3 (mitogen-activated protein kinase phosphatase 3) is an ERK2-specific phosphatase that terminates ERK2 signaling.

Purpose of the Study:

• To elucidate the molecular mechanisms underlying the specific recognition and dephosphorylation of ERK2 by MKP3.
• To map the interaction surfaces involved in the ERK2-MKP3 complex formation.

Main Methods:

• Hydrogen/deuterium exchange mass spectrometry (HDX-MS) was employed to map protein-protein interaction surfaces.
• Analysis of specific peptide sequences and their binding sites on ERK2.

Main Results:

• MKP3 recognizes ERK2 via two distinct protein-protein interactions, ensuring high specificity.
• A common docking site on ERK2, opposite the active site, binds MKP3 motifs for initial tethering.
• A second interaction at the ERK2 substrate-binding site allosterically activates MKP3 for efficient dephosphorylation.

Conclusions:

• A bipartite interaction model explains the specific recognition and regulation of ERK2 by MKP3.
• This model, involving both docking and substrate-site interactions, is crucial for productive dephosphorylation.
• The identified interaction principles may extend to other MAPK-regulator and MAPK-substrate interactions.