Multidimensional Engineering of Cyberlindnera jadinii for Efficient Invertase Production

The yeast Cyberlindnera jadinii has long been used in food and feed industries. While it shows strong intracellular protein synthesis, its limited secretion capacity hampers its use as a protein production host. To overcome this, we used the endogenous invertase CjINV1 as a model and applied multidimensional engineering to the tetraploid wild-type strain NBRC0988. This involved multicopy integration of gene expression cassettes into the chromosome via a landing pad strategy, transcriptional enhancement through combinatorial optimization of promoter and terminator elements, and chitin-based cell wall remodeling to facilitate protein release. The engineered strain showed markedly enhanced protein secretion in a 5-L bioreactor, yielding 3456 ± 81 mg/L extracellular protein after 168 h cultivation, with invertase specific activity of 789.3 ± 52.3 U/mg. To our knowledge, this represents the highest reported yield for yeast-based invertase production to date, highlighting C. jadinii as a promising platform for recombinant protein production.