Non-adherence is a major hurdle for the effective treatment of HIV, a global epidemic even after 30 years of medical advancements. Long-acting therapeutics have the potential to improve adherence through simplified regimens. One such investigational medication is MK-8591, a nucleoside reverse transcriptase translocation inhibitor with sub-nanomolar antiviral activity and a long intracellular half-life. To support this program, a world-class manufacturing synthesis which is robust, green, and sustainable was targeted. In this presentation, I will describe how our team has addressed the long standing challenge of stereoselective glycosylation with a novel four-enzyme biocatalytic cascade. I will also discuss various routes developed to efficiently complete the synthesis, and the catalytic methods developed to enabled each approach. Highlights include two new methods for enantioselective ketone alkynylation, and a route consisting of nine enzymes carried through one pot.