Microorganisms such as Komagataella phaffii (Pichia pastoris) are commonly used in the large-scale manufacturing of proteins and enzymes for biotherapeutic purposes. Researchers from the Love group at the Koch Institute at MIT report a scalable technique which uses an engineered Komagataella phaffii platform to produce receptor binding domain (RBD) proteins, which are expressed on SARS-CoV-2 S proteins and are used as antigens in multiple SARS-CoV-2 vaccine candidates. The manufacturing method developed here does not require the use of methanol to induce bacterial metabolic processes, which simplifies manufacturing and reduces flammability risk. Instead, they integrated additional copies of endogenous transcription factors mit1 and mxr1 to allow constitutive overexpression of the recombinant gene in the presence of sorbitol. In general, this methanol-free manufacturing scheme resulted in a 3- to 5-fold increase in productivity of these engineered strains. The group hypothesizes that this improved productivity is possibly because sorbitol-fed mit1+ strains exhibit less protein folding stresses during RBD transcription. The group compared manufacturing of both the methanol- and sorbitol-induced yeast using their InSCyT biomanufacturing platform—finding that the sorbitol-induced yeast exhibited sustained RBD production with less cellular stress and fewer impurities than the base strain. They then demonstrated that they could scale up to liter scale cultures in a bioreactor, and that the mit1+ cells were capable of producing RBD variants.*