Purification and Characterization of Geranylgeranyl Diphosphate Synthase from Methanobacterium thermoformicicum SF-4.

Geranylgeranyl diphosphate (GGPP) synthase [EC 2.5.1.29] was purified to homogeneity from Methanobacterium thermoformicicum SF-4. The enzyme was a dimeric protein consisting of two identical subunits (Mr = 39,000) and catalyzed prenyl transfer reactions using isopentenyl diphosphate (Km = 30.8 μM) and either dimethylallyl diphosphate (Km=16.8 μM), geranyl diphosphate (Km=12.6 μM), or farnesyl diphosphate (Km=14.7 μM) as allylic partners. During a sequential elongation, C5→C10→C15→C20' intermediates were accumulated with various ratios to the final product GGPP. In the presence of 0.8 M KCl, GGPP synthease activity was greatly enhanced, stabilized to heat treatment at 65°C for 30 min, and protected from inhibition by p-chloromercuribenzoic acid. No other prenyltransferase synthesizing C20 or shorter prenyl diphosphate was observed in M. thermoformicicum SF-4. These suggest that GGPP synthase alone is important in the biosynthetic pathways to squalene and membrane polar lipids at a chain elongation stage in this strain.