Red blood cells (RBCs) are transcriptionally silent yet dynamically remodel metabolism in response to oxygen tension. Using ultra-pure human RBCs, we generated the deepest contamination-free proteome to date (3,775 proteins) and mapped the oxygen-dependent interactome. These datasets reveal an oxygen-responsive metabolon centered on the Band 3 (SLC4A1) N-terminus. We identify biliverdin reductase B (BLVRB) as a previously unrecognized Band 3 interactor that dissociates under hypoxia, coincident with increased Band 3–deoxyhemoglobin contacts. This reversible assembly functions as an oxygen-sensitive switch coordinating redox and glycolytic remodeling. Humanized mice lacking Band 3 N-terminal segments exhibit impaired glycolytic activation, reduced 2,3-bisphosphoglycerate synthesis, and diminished exercise tolerance, demonstrating physiological relevance. Population-scale cis-pQTLs for SLC4A1 and BLVRB suggest functions beyond canonical heme catabolism. Mechanistically, BLVRB serves as a nitric-oxide relay that trans-nitrosates the glycolytic enzyme GAPDH at C152, transiently inhibiting glycolysis. Together, these findings define a Band 3–BLVRB–hemoglobin axis integrating oxygen sensing and metabolic control.