This study reports engineered protease-stable, gramicidin-inspired, peptide-based proton transporter LD8Δ, with alternating L- and D-amino acid residues, to treat the highly chemoresistant, radioresistant, immunosuppressive metastatic renal cell carcinoma (RCC) cell line SK-RC-45. Exploiting the potential of the LD8Δ proton transporter, this study proposes mechanistically rational combination therapy with high translational potential. Current treatment methods involve multi-targeted tyrosine kinase inhibitors, which are accompanied by serious side-effects and exhibit an alarmingly low median overall survival. Computational and experimental data suggested that LD8Δ induced considerable membrane deformation, which supported the destabilization of the intracellular pH regulating mechanism observed in LD8Δ-treated RCC. This mechanistically rational combination therapy using LD8Δ and HIF-2α silencing siRNA induced mitochondrial depolarization, cell cycle arrest, apoptosis, reduction in angiogenesis and disruption of the major oncogenic signalling pathways in SK-RC-45 cells. The designed therapy upregulated the VHL tumour suppressor protein and downregulated HIF-2α protein expression, thus decreasing the activity of the HIF transcriptional factor, which is the master regulator causing clear cell RCC. Additionally, it upregulated DAB2IP and facilitated radiosensitization in the radioresistant SK-RC-45 cells. To the best of our knowledge, this is the first study demonstrating that a designed biocompatible proton transporter can potentiate immunotherapy against RCC by inducing the downregulation of the dual checkpoint proteins CD47, PD-L1 and ganglioside GM2, resulting in enhanced phagocytosis and preventing T cell inactivation and T cell apoptosis.