The biosynthesis of mannosyl-1-phosphoryl-polyisoprenol in Micrococcus lysodeikticus and its role in mannan synthesis.

Previous studies from this laboratory have shown that crude, cell-free extracts of Micrococcus lysodeikticus catalyze the incorporation of mannose from GDP- mannose into three lipids1 and a mannan.2Two of the lipids have been identified as a-D-mannosyl-(l -# 3')-diglyceride and a-D-mannosyl-(l 3)-a-D-mannosyl-(1 3')-diglyceride.1In this communication, the results of studies on the structure of the third mannose-containing lipid are presented.All the available evidence indicates that the structure of this compound is mannosyl-1-phosphorylpolyisoprenol (MPP).Preliminary experiments suggest that IMPP serves as an intermediate in the enzymatic transfer of mannosyl groups from GDP-mannose to mannan.Methods.-Enzyme preparation: A particulate preparation that catalyzes the synthesis of MPP, mannan, and mannosyldiglyceride was obtained by centrifugation of a crude, cell-free extract of M. lysodeikticus' at 100,000 X g for 1 hr.The MPP synthetase was prepared from the crude extract as follows: Crude extract (60 ml) was heated at 40°f or 20 min.The pH of the solution (at 00) was adjusted to 5.5 with 1 N acetic acid, and 15.7 gm of (NH4)2SO4 was added with stirring.After 30 min, the precipitate was collected by centrifugation, dissolved in 0.02 M Tris-HCI, pH 7.6 (buffer A), and dialyzed overnight against 2 liters of buffer A.