A conserved P-loop anchor limits the structural dynamics that mediate nucleotide dissociation in EF-Tu.

Area of Science:

• Biochemistry
• Structural Biology
• Molecular Biophysics

Background:

• The phosphate-binding loop (P-loop) is a conserved motif in nucleotide-binding proteins, but its structural dynamics and role in binding are not fully understood.
• Understanding these principles is crucial for biomolecular engineering.

Purpose of the Study:

• To investigate the relationship between P-loop structural dynamics and GTP-binding properties in Elongation Factor (EF) Tu.
• To explore the biomolecular engineering applications of these findings.

Main Methods:

• Utilized in vitro rapid-kinetics measurements and in silico molecular dynamics (MD) simulations.
• Analyzed wild-type EF-Tu and P-loop variants using Steered Molecular Dynamics (SMD) simulations.

Main Results:

• A correlation was found between P-loop flexibility and the entropy of activation for GTP dissociation.
• Increased backbone flexibility in N-terminal P-loop amino acids was observed during force-induced GTP dissociation.
• Identified Gly18 and His19 interactions with helix C as a P-loop anchoring module.

Conclusions:

• P-loop flexibility directly influences GTP binding and dissociation kinetics.
• A conserved P-loop anchoring module, involving helix C interactions, is proposed.
• This anchoring module is suggested to be conserved across various GTPases and ATPases.