Dynamic Mechanisms of Neutral Amino Acid Exchanger ASCT2 Transporting Substrate Glutamine

Tumor cells rely on the high expression of transporter proteins to meet their nutrient demands, with alanine-serine-cysteine transporter 2 (ASCT2) being a key player in glutamine (Gln) uptake. Glutamine, a conditionally essential amino acid abundant in protein-rich foods, such as meat, dairy, and legumes, serves as a critical nitrogen and carbon source for cellular biosynthesis. ASCT2-mediated Gln transport not only fuels cancer progression but also plays a role in nutrient absorption in healthy tissues, particularly the gut, where dietary amino acids are assimilated. Despite its dual significance in physiology and pathology, the molecular mechanisms of Gln transport by ASCT2 remain poorly understood, hindering the development of targeted therapies and dietary interventions. In this study, microsecond classical and Gaussian accelerated molecular dynamics (CMD/GaMD) were conducted to investigate the dynamic mechanism of glutamine transportation by ASCT2. A Markov State Model (MSM) was built based on the enhanced sampling trajectories to search for the communication pathways and critical transition states during the allosteric movement of helical hairpin 2 (HP2), a structural gatekeeper of transport. Additionally, the C467R mutation was found to disrupt HP2 dynamics, impair Gln binding, and hinder Gln transport. Intriguingly, substrate-bound ASCT2 exhibited prolonged HP2 opening compared to its unloaded state, suggesting that food-derived Gln may stabilize transporter conformations. The important transition states of the transporter opening process were also identified from the MSM. The key pathway from the “close” to “open” state is S2 → S15 → S10 → S6 → S5 → S7 → S19, with a maximum probability of 16.76%. These findings not only advance ASCT2-targeted drug discovery but also offer a framework for designing functional foods or nutraceuticals that modulate amino acid bioavailability, potentially leveraging natural compounds to fine-tune ASCT2 activity for cancer prevention or metabolic health optimization.