The modular organization of the type I polyketide synthases (PKSs) is propitious for rational engineering of desirable analogous. By combining multiple, state-of-the-art approaches including modification of docking domains, use of modules of varying domain composition, alternative interdomain fusion sites, and targeted adaptation of key domain-domain interfaces, we were able to reprogram the stambomycin PKS from Streptomyces ambofaciens ATCC23877 and to produce the target 37-membered mini-stambomycin metabolites, a reduction in chain length of 14 carbons relative to the 51-membered parental compounds. Shunt metabolites released from the multienzyme subunit upstream of the newly-installed junction were also produced.