An isoleucine-based allosteric switch controls affinity and shape shifting in integrin CD11b A-domain

Area of Science:

• Biochemistry
• Structural Biology
• Cell Biology

Background:

• Integrins mediate cell adhesion by switching between low and high affinity states upon activation.
• Ligand binding typically occurs via the von Willebrand Factor A-type domain of integrins.
• Previous crystallographic studies suggested two conformations (closed and open) of this domain, but direct structure-function links were missing.

Purpose of the Study:

• To establish structure-function correlates for the integrin CD11b A-domain.
• To define the allosteric switch controlling integrin conformation and ligand-binding affinity.
• To investigate the role of a specific C-terminal isoleucine residue in affinity regulation.

Main Methods:

• Expression of stable low and high affinity forms of the integrin CD11b A-domain.
• Determination of binding isotherms to quantify ligand affinity.
• Crystallographic analysis to resolve the structures of different affinity states.
• Solution-based structural difference analysis.

Main Results:

• A low affinity form, lacking an N-terminal extension, adopted a closed conformation and did not bind ligands.
• High affinity forms, generated by modifying a C-terminal isoleucine (Ile316), adopted an open conformation.
• The Ile316 residue is crucial, wedged in a hydrophobic socket in the closed (low affinity) state and displaced in the open (high affinity) state.
• Structural differences between low and high affinity states were observed in solution.

Conclusions:

• A ligand-independent, isoleucine-based allosteric switch intrinsic to the CD11b A-domain controls its conformation and affinity.
• This switch provides a molecular mechanism for integrin activation.
• The findings establish direct structure-function relationships for integrin affinity modulation.