Ligand binding energy and enzyme efficiency from reductions in protein dynamics

Area of Science:

• Biochemistry
• Enzymology
• Protein Dynamics

Background:

• Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is a tetrameric enzyme crucial for glycolysis.
• Cofactor binding significantly influences enzyme activity and stability.
• Understanding ligand-induced conformational changes is key to enzyme mechanism elucidation.

Purpose of the Study:

• To investigate the relationship between protein dynamics and cofactor binding affinity in rabbit muscle GAPDH.
• To quantify the effect of successive NAD+ binding on GAPDH's internal dynamics.
• To explore the contribution of dynamic changes to binding energy and catalytic efficiency.

Main Methods:

• Characterization of tetrameric rabbit muscle glyceraldehyde 3-phosphate dehydrogenase (GAPDH).
• Analysis of successive nicotinamide adenine dinucleotide (NAD+) binding events and their affinities.
• Assessment of protein dynamics changes upon cofactor binding.

Main Results:

• GAPDH binds four NAD+ molecules with decreasing affinities (10^11 to 10^5 M^-1).
• Protein dynamics are most reduced upon binding the first NAD+, with smaller reductions for subsequent molecules.
• The fourth NAD+ molecule binds without significant dynamic change.

Conclusions:

• Reduced protein dynamics, coupled with improved internal bonding, directly account for the enhanced NAD+ binding affinity of GAPDH.
• This mechanism of dynamic modulation of binding energy may be applicable to other enzyme-ligand interactions.
• Understanding these dynamics offers insights into enzyme catalytic efficiency and ligand binding.