Conformational equilibria and intrinsic affinities define integrin activation.

Area of Science:

• Biochemistry
• Cell Biology
• Structural Biology

Background:

• Integrins are crucial cell surface receptors mediating cell adhesion and signaling.
• Integrin α5β1 plays a key role in various biological processes, including cell migration and wound healing.
• Understanding integrin conformational states is vital for deciphering their activation mechanisms.

Purpose of the Study:

• To define the discrete free energies of integrin α5β1 conformational states.
• To quantify the affinities of each state for its ligand.
• To elucidate the energetic requirements and regulation of integrin activation.

Main Methods:

• Measurement of intrinsic affinities for ligand across different conformational states.
• Determination of equilibrium constants linking these states.
• Analysis of free energy requirements using protein fragments and intact integrin α5β1.
• Characterization of integrin α5β1 conformation on K562 cells.

Main Results:

• Three distinct conformational states of integrin α5β1 possess unique free energies.
• The extended-open state exhibits a 5,000-fold higher ligand affinity compared to bent-closed and extended-closed states.
• Integrin α5β1 exists predominantly in the bent-closed conformation (99.8%) on K562 cells.
• Cellular energy input is required to overcome stabilization of the bent conformation by transmembrane and cytoplasmic domains.
• N-glycans and leg domains contribute to headpiece opening by regulating conformational equilibria.

Conclusions:

• Integrin activation is a tightly regulated process involving discrete energy states and profound affinity modulation.
• Cellular energy and intrinsic structural features govern the conformational transitions of integrins.
• These findings offer new principles for understanding signaling in receptors with extracellular domains linked to single-span transmembrane domains.