A Novel Lithium Ion-Assisted TPP Approach for Enhanced Target Discovery

Traditional thermal proteome profiling (TPP) suffers from limitations in detecting membrane and weakly bound proteins, potentially overlooking up to 20% of drug targets. To overcome this, we developed a lithium ion-assisted TPP (Li-TPP) method that significantly enhances protein thermal stability detection. Li-TPP successfully captured thermal shifts in weakly bound proteins like lactate dehydrogenase A (LDHA) and resolved changes within NMDA receptor complexes. Compared to traditional TPP, Li-TPP identified 718 additional proteins (R 2 > 0.8, min_slope ≤ −0.06) and significantly increased thermal displacement values (ΔTm ≥ 1.5 °C) in 368 coidentified proteins. Importantly, Li-TPP identified known mania-associated targets like GSK3β as well as novel regulatory proteins such as CRMP2. Functional enrichment analysis revealed that Li-TPP-identified proteins were significantly enriched in pathways critical to bipolar disorder, including microtubule dynamics (MT3 complex), synaptic plasticity (NMDA receptor assemblies), and energy metabolism (ATP-binding proteins). This provides mechanistic insights into lithium’s therapeutic action, highlighting its role in stabilizing neuronal energy homeostasis and microtubule architecture. This study demonstrates Li-TPP’s superior performance in detecting challenging protein targets and provides a powerful new platform for elucidating drug mechanisms and accelerating the development of multitarget therapeutics for bipolar disorder.