Are acidic and basic groups in buried proteins predicted to be ionized?

Area of Science:

• Protein biochemistry
• Computational biophysics
• Structural biology

Background:

• Ionizable residues (Asp, Glu, Arg, Lys) are vital for protein structure, stability, and function, comprising ~25% of protein composition.
• Understanding residue ionization states is key to predicting protein behavior and interactions.

Purpose of the Study:

• To predict the ionization states of acidic and basic residues in a large protein dataset.
• To quantify the impact of solvation energy and protein microenvironment on residue ionization.
• To investigate the conservation patterns of ionized residues.

Main Methods:

• Utilized Multi-Conformation Continuum Electrostatics (MCCE) calculations.
• Analyzed ionization states of 36,192 ionizable residues across 490 proteins.
• Quantified solvation energy changes (DeltaDeltaG(rxn)) and their effect on pKa shifts.

Main Results:

• Predicted 93.5% of ionizable residues to be ionized.
• Identified significant solvation energy losses for buried ionized residues, shifting pKa values.
• Observed that 85% of buried residues with substantial pKa shifts were ionized, with specific propensities for Arg, Asp, Glu, and Lys.
• Demonstrated differential stabilization effects of backbone dipoles and polar side-chains (Asn, Gln, Ser, Thr, Tyr) on charged residues.
• Found buried ionized residues to be more conserved than surface-exposed ones.

Conclusions:

• Protein microenvironments significantly impact residue ionization, often favoring ionization even for buried residues.
• Ion-pair interactions and specific side-chain/backbone interactions play critical roles in stabilizing ionization states.
• The ionization state and burial status of residues correlate with their evolutionary conservation, highlighting their functional importance.