Structural studies of geranylgeranylglyceryl phosphate synthase, a prenyltransferase found in thermophilic Euryarchaeota

Abstract

Archaea are uniquely adapted to thrive in harsh environments, and one of these adaptations involves the archaeal membrane lipids, which are characterized by their isoprenoid alkyl chains connected via ether linkages to glycerol 1-phosphate. The membrane lipids of the thermophilic and acidophilic euryarchaeota Thermoplasma volcanium are exclusively glycerol dibiphytanyl glycerol tetraethers. The first committed step in the biosynthetic pathway of these archaeal lipids is the formation of the ether linkage between glycerol 1-phosphate and geranylgeranyl diphosphate, and is catalyzed by the enzyme geranylgeranylglyceryl phosphate synthase (GGGPS). The 1.72 Å resolution crystal structure of GGGPS from T. volcanium (TvGGGPS) in complex with glycerol and sulfate is reported here. The crystal structure reveals TvGGGPS to be a dimer, which is consistent with the absence of the aromatic anchor residue in helix 5a that is required for hexamerization in other GGGPS homologs; the hexameric quaternary structure in GGGPS is thought to provide thermostability. A phylogenetic analysis of the Euryarchaeota and a parallel ancestral state reconstruction investigated the relationship between optimal growth temperature and the ancestral sequences. The presence of an aromatic anchor residue is not explained by temperature as an ecological parameter. An examination of the active site of the TvGGGPS dimer revealed that it may be able to accommodate longer isoprenoid substrates, supporting an alternative pathway of isoprenoid membrane-lipid synthesis.

Hexamerization and thermostability emerged very early during geranylgeranylglyceryl phosphate synthase evolution

“…We conclude that histidine with its heterocyclic aromatic ring also functions as an “aromatic anchor” to stabilize the hexamer, and suspect that all extant sulfolobic GGGPS proteins are hexamers as well. In conclusion, almost all group II GGGPS enzymes must be considered to be hexameric, except the bacterial proteins from Flavobacteriales and a very small number of archaeal proteins, mainly from Thermoplasmatales, 4,16,17 as summarized in Table S1.…”

“…To our knowledge, group II GGGPS enzymes from only two phyla have been reported to be dimeric based on experimental data (Table S1). These are the bacterial Flavobacteriales (fjGGGPS, zpGGGPS 4 ; fsGGGPS, this work) and the archaeal Thermoplasmatales (taGGGPS 4,16,19 ; tvGGGPS 17 ). To study the emergence of these dimeric group II GGGPS, we expressed synthetic genes for predecessors of the Flavobacteriales and analyzed the purified proteins.…”

“…GGGPS sequences from Thermoplasmatales are missing in our ASR, because their somewhat diverse sequences were filtered out within the process of identification of wandering sequences to obtain robust ASR results 12 . An alternative ASR of GGGPS sequences including Thermoplasmatales has been published recently 17 . Based on computational data only, the authors postulate a convergent evolution for the hexamerization of group II GGGPS in the different phyla, which goes along with dimeric predecessors at the deep nodes in the phylogenetic tree.…”

The catalytic and structural basis of archaeal glycerophospholipid biosynthesis

“…This reaction marks the first committing step into the biosynthesis of archaeal phospholipids. Presumably, this biosynthesis step occurs in the cytosol as GGGPS enzymes do not contain transmembrane helices and are purified as soluble proteins from the soluble fraction of cell lysates (Payandeh et al 2006 ; Peterhoff et al 2012 , 2014 ; Linde et al 2018 ; Nemoto et al 2019 ; Blank et al 2020 ; Kropp et al 2021 ).…”

“…To date, the protein data bank (PDB) contains 6 reported GGGPS crystal structures. Notably, most structures are from thermophilic archaea such as: Archaeoglobus fulgidu s (AfGGGPS, group Ia, (Payandeh et al 2006 )), Methanothermobacter thermoautotrophicus (MtGGGPS, group IIa, (Peterhoff et al 2014 )), Thermoplasma acidophilum (TaGGGPS, group IIa, (Nemoto et al 2019 )) and Thermoplasma volcanium (TvGGGPS, group IIa, (Blank et al 2020 )). The structure of a bacterial GGGPS (group IIb, (Peterhoff et al 2014 )) from Flavobacterium johnsoniae was also reported (FjGGGPS).…”