Computer Simulations of NAD Channeling between GAPDH and LDH

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Biophysical Journal




Functional protein-protein interactions are essential for many physiological processes and may play important roles in substrate channeling, coenzyme transfer, and compartmentation in glycolysis. Herein, Brownian dynamics (BD) elucidates the interactions between the glycolytic enzymes glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and lactate dehydrogenase (LDH) and the transfer of the cofactor nicotinamide adenine dinucleotide (NAD) between LDH and GAPDH. BD channeling simulation results strongly depend on choice of reactive atom set. When the reactive atoms set comprise of atoms in the vicinity of the NAD binding site of either enzyme, short NAD trajectories between enzyme subunits occur. If the reactive atoms set were chosen from atoms belonging to NAD binding sites, the efficiency of reaching LDH decreased significantly, and even the shortest trajectories spent time equilibrating with solvent before binding the next active site. Transfer of NAD from GAPDH to LDH is sensitive to overall structure of enzyme-enzyme complex. Small variations in orientation of one enzyme relative to the other cause changes in channeling efficiency. The process of NAD release from GAPDH and LDH binding sites was studied with steered molecular dynamics. The GAPDH/LDH complex with two LDH subunits facing two GAPDH subunits was chosen as initial structure. Six NAD molecules were included - two molecules in GAPDH active subunits and 4 NAD molecules bound to each LDH subunit. External forces were applied to O3 NAD atom of each NAD molecule. MD trajectories (2 ns) with external force (≤ 1000 pN) were able to pull the NAD out of GAPDH, but not out of LDH. Such strong binding of NAD by LDH is because the nicotinic moiety is buried deep inside LDH subunit globule. Additional studies are needed to confirm the hypothesis that intermolecular contacts soften GAPDH subunit and its affinity to NAD reduces.


2, Supplement 1

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