Insulin Receptor-Related Receptor Activation by Artificial Double-ER Mutations in the Transmembrane Domain

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• The insulin receptor-related receptor (IRR) is activated by alkaline extracellular pH.
• IRR activation involves synergistic sites in its extracellular domain.
• The role of the transmembrane domain in IRR pH sensing is not well understood.

Purpose of the Study:

• To investigate the role of the IRR transmembrane domain in receptor activation and pH sensing.
• To engineer mutations in the transmembrane domain to modulate IRR activity and pH sensitivity.

Main Methods:

• Molecular modeling based on NMR-derived structure to predict amino acid substitutions.
• Site-directed mutagenesis to introduce specific mutations in the IRR transmembrane domain.
• Phosphorylation assays to assess receptor activation across a pH range.

Main Results:

• Identified mutations that enhance non-covalent interactions within the IRR transmembrane dimer.
• A double mutant (A938E-A939R) showed high phosphorylation at neutral pH and retained alkaline sensitivity.
• Another double mutant (V929E-G930R) exhibited strong basal phosphorylation across the tested pH range.

Conclusions:

• The transmembrane domain is critical for IRR activation and pH sensing.
• Targeted mutations in the transmembrane domain allow for precise control over IRR function and pH sensitivity.
• These findings provide insights into the mechanism of IRR activation and potential therapeutic strategies.