In June 2023, the “Computational Protein Engineering” (CPE) group was established at DTU Biosustain with the aim of supporting the development of novel bioprocesses for the sustainable manufacturing of natural products, biochemicals, and microbial foods. Current efforts in the CPE group include the application of machine learning methods based on generative AI and active learning as well as the development of experimental (e.g., biosensors) and computational (e.g., software) tools for improving enzymes as single biocatalysts, or as parts of metabolic pathways for the development of cell factories. In this talk, I will present how methodology development was (and continues to be) crucial for protein and metabolic engineering and directed evolution. As example, I will focus on the multiparametric optimisation of cytochrome P450 mono-oxygenase and formate dehydrogenase, enzymes relevant for the biosynthesis of natural products, biochemicals and C1 metabolism. Finally, I will also summarize efforts towards the development of microbial cell factories to produce B vitamins, earlier work carried out before switching from industry to academia.