Biocatalytic conversions can involve one enzyme that carries out one specific reaction at a time, or multiple enzymes that carry out a series of conversions to yield a desired product. The use of several enzymes allows the realization of much more complex synthetic schemes. Multi-step synthesis can be carried out in biological systems by utilizing or engineering their metabolic networks for catalysis. Alternatively, multi-enzymatic catalysis can be carried out in vitro using isolated biocatalysts. Both approaches, in vivo or in vitro, have their specific advantages, problems, and challenges that will be illustrated using recent examples.