Linker Dependence of Avidity in Multivalent Interactions Between Disordered Proteins

Area of Science:

• Biochemistry
• Structural Biology
• Protein Dynamics

Background:

• Multidomain proteins utilize multiple binding sites connected by disordered linkers.
• Linker architecture influences protein avidity by altering intramolecular binding effective concentration.
• Existing theoretical models for linker-mediated avidity lack experimental validation due to difficulties in measuring effective concentrations.

Purpose of the Study:

• To develop an experimental system for measuring effective concentrations in bivalent protein interactions.
• To experimentally assess theoretical models of avidity in disordered, multivalent protein systems.
• To investigate the relationship between linker length, interaction strength, and avidity.

Main Methods:

• Developed a model system for bivalent protein interactions with variable disordered linkers.
• Employed competition experiments to directly measure effective concentrations.
• Performed kinetic and thermodynamic characterization of protein interactions across different linker lengths and strengths.

Main Results:

• Established a direct experimental method for quantifying effective concentration in linker-connected protein systems.
• Confirmed that avidity emerges when effective concentration matches the weakest binding affinity.
• Observed that avidity decreases with increasing linker length, but to a lesser extent (one-third) than predicted by current models.

Conclusions:

• Experimental data partially validates theoretical models of avidity in disordered, multivalent interactions.
• The attenuated length dependence of avidity suggests compensatory mechanisms like linker interactions or entanglement.
• Disordered linkers play a direct role in avidity, offering a potential allosteric regulation mechanism for multivalent proteins.