Dolichylphosphate mannose synthase (DPMS) performs an essential function by synthesizing the activated lipid-linked mannose intermediate used in protein glycosylation pathways. In eukaryotes and archaea, DPMS catalyzes the transfer of mannose from GDP-mannose to dolichylphosphate to generate dolichylphosphate mannose (Dol-P-Man). Type-III DPMS from Pyrococcus furiosus (PfDPMS) has a catalytic domain attached to a GtrA-like transmembrane (TM) domain with an unusual topology. Here, we present crystallographic data from a crystal complex determined from an enzymatic reaction mixture that provides detailed information about donor- and acceptor binding in the active site prior to mannosyl transfer. We also present a new, unexpected structural state for the TM domain in which a Dol-P-Man molecule is bound “upside-down” with its mannosylphosphate headgroup positioned in a polar pocket between the TM helices. By generating a panel of TM-domain mutants, we confirm that the TM domain does not participate directly in the catalysis of mannosyl transfer and discuss the possibility of this domain providing moonlighting function to PfDPMS by translocating the Dol-P-Man product to the cell exterior.