Sugar-protein interactions control protein-complex stability in crowded Ficoll and dextran solutions.

Area of Science:

• Biochemistry
• Biophysics
• Physical Chemistry

Background:

• Traditional models of molecular crowding focus on excluded volume effects stabilizing compact protein states.
• High cosolute concentrations involve complex interactions beyond simple volume exclusion.
• Understanding these effects is crucial for protein stability and cellular processes.

Purpose of the Study:

• To systematically investigate the effects of various cosolutes (dextrans, Ficolls, glucose, sucrose) on protein complex thermodynamics.
• To differentiate the crowding mechanisms of polymers from their monomeric constituents.
• To analyze the contributions of excluded volume, chemical interactions, and non-ideal mixing.

Main Methods:

• Studied two variants of streptococcal protein G B1 domain (side-by-side dimer and domain swap dimer).
• Monitored protein complex dissociation using 19F NMR as a function of cosolute properties (molecular weight, concentration) and temperature.
• Applied model fitting to dissect thermodynamic contributions.

Main Results:

• Cosolute interaction distances depend on polymer structure and concentration, unlike sugar monomers.
• At high concentrations, polymer mesh size becomes the effective length scale, reducing the impact of large polymers.
• Excluded volume stabilization is offset by destabilizing chemical interactions (monomers) or non-ideal mixing (polymers).

Conclusions:

• The mechanism of molecular crowding by polymers is distinct from that of their monomers.
• Protein crowding effects are significantly influenced by the structural properties and concentration of the cosolutes.
• These findings refine our understanding of macromolecular crowding in biological systems.