Unveiling the Intramolecular Thermodynamics of Multivalent Proteins: Exploratory Study on Engineered Protein Model.

Area of Science:

• Biochemistry
• Structural Biology
• Protein Engineering

Background:

• Multidomain proteins mediate crucial biological processes through multivalent interactions.
• Understanding the thermodynamics of these domain interactions is essential but challenging.
• Current knowledge lacks detailed thermodynamic insights into intramolecular interactions within protein complexes.

Purpose of the Study:

• To investigate the temperature-dependent kinetics and thermodynamics of multidomain protein interactions.
• To develop a theoretical model for extracting thermodynamic quantities of inter- and intramolecular interactions.
• To provide insights into the thermodynamics of intramolecular interactions for protein design and engineering.

Main Methods:

• Utilized surface plasmon resonance (SPR) to measure temperature-dependent binding kinetics.
• Screened protein-peptide binding pairs with multivalent scaffolds to minimize nonspecific binding.
• Employed initial rate analysis within a theoretical framework to quantify thermodynamic parameters.

Main Results:

• Successfully selected protein-peptide pairs with suitable monomer kinetics for higher valence studies.
• Developed and validated a theoretical model to extract thermodynamic quantities for both inter- and intramolecular interactions.
• Quantified thermodynamic parameters governing complex multivalent protein interactions.

Conclusions:

• Novel insights into the thermodynamics of intramolecular interactions in multivalent protein complexes were obtained.
• The developed methodology enables the study of complex protein interactions across varying valences.
• Findings have significant implications for the rational design and engineering of proteins.