Biosyntheis of FK506 and its engineering for the generation of novel analogues

FK506 (known as tacrolimus) is a macrocyclic polyketide of microbial origin exhibiting various biological activities, including immunosuppressive, antifungal, antiinflammatory, neuroprotective, and neuroregenerative. It is a clinically important drug used to prevent the rejection of organ transplants and to treat autoimmune diseases such as atopic dermatitis. We have elucidated the detailed biosynthetic pathway of allylmalonyl-coenzyme A from which the FK506 allyl group is derived. Characterization of this discrete pathway facilitated the engineered biosynthesis of novel allyl group-modified FK506 analogues, 36-fluoro-FK520 and 36-methyl-FK506, the latter of which exhibits improved neurite outgrowth activity. We have also detailed the parallel pathways responsible for the post-modification step in the biosynthesis of FK506, demonstrating that substrate-flexible post-PKS modification enzymes, FkbD and FkbM, can provide a potential tool for the combinatorial biosynthesis of novel macrolide derivatives. In addition,  FK506 analogues containing a non-natural starter unit could be obtained through mutasynthesis. Taken together, these results illuminates an efficient strategy for the engineered biosynthesis of FK506 analogues with improved therapeutic properties.