Crystal Structure of Human Squalene Synthase

Abstract

Squalene synthase catalyzes the biosynthesis of squalene, a key cholesterol precursor, through a reductive dimerization of two farnesyl diphosphate (FPP) molecules. The reaction is unique when compared with those of other FPP-utilizing enzymes and proceeds in two distinct steps, both of which involve the formation of carbocationic reaction intermediates. Because FPP is located at the final branch point in the isoprenoid biosynthesis pathway, its conversion to squalene through the action of squalene synthase represents the first committed step in the formation of cholesterol, making it an attractive target for therapeutic intervention. We have determined, for the first time, the crystal structures of recombinant human squalene synthase complexed with several different inhibitors. The structure shows that SQS is folded as a single domain, with a large channel in the middle of one face. The active sites of the two half-reactions catalyzed by the enzyme are located in the central channel, which is lined on both sides by conserved aspartate and arginine residues, which are known from mutagenesis experiments to be involved in FPP binding. One end of this channel is exposed to solvent, whereas the other end leads to a completely enclosed pocket surrounded by conserved hydrophobic residues. These observations, along with mutagenesis data identifying residues that affect substrate binding and activity, suggest that two molecules of FPP bind at one end of the channel, where the active center of the first half-reaction is located, and then the stable reaction intermediate moves into the deep pocket, where it is sequestered from solvent and the second half-reaction occurs. Five ␣ helices surrounding the active center are structurally homologous to the active core in the three other isoprenoid biosynthetic enzymes whose crystal structures are known, even though there is no detectable sequence homology.The isoprenoid biosynthetic pathway yields a structurally diverse family of low molecular weight molecules with a variety of physiological functions (1-3). In humans, the pathway produces such critical end-products as cholesterol, bile acids, dolichol, ubiquinone, steroid hormones, and prenylated proteins. Chain length-selective prenyltransferases catalyze the successive head-to-tail addition of the 5-carbon (C5-) isopentenyl diphosphate to the growing isoprene chain to form a series of linear C10-, C15-, C20-and C25-isoprenoid diphosphates. Cyclic terpenes are formed from these linear isoprenoid diphosphates through the actions of numerous class I terpene cyclases (3). The longer 30-carbon linear terpene, squalene, of cholesterol biosynthesis and the 40-carbon linear terpene, phytoene, of carotenoid biosynthesis are formed by head-to-head condensations, respectively, of two 15-carbon and 20-carbon isoprenoid diphosphates, through the actions of the prenyltransferases squalene synthase and phytoene synthase (4) and are subsequently converted into their respective cyclic terpenes through the action of specific class II terpene c...

Structure of trichodiene synthase from Fusarium sporotrichioides provides mechanistic inferences on the terpene cyclization cascade

“…The x-ray crystal structures of three cyclases have been solved to date: pentalenene synthase from Streptomyces UC5319 (12), 5-epi-aristolochene synthase from Nicotiana tabacum (13), and aristolochene synthase from Penicillium roqueforti (14). Despite a lack of significant overall sequence identity, these enzymes all share the ''terpenoid synthase fold'' (12), which is also shared by avian farnesyl diphosphate synthase (15) and human squalene synthase (16). Structural similarity maintained in the face of broad synthetic diversity indicates divergence from a common ancestor early in the evolution of terpene biosynthesis.…”

Unearthing the roots of the terpenome

“…Accordingly, naturally occurring enzymes catalyzing these four isoprenoid coupling reactions likely evolved from an FPPase-like ancestor [20]. For example, squalene synthase catalyzes a cyclopropanation reaction ( Figure 1) and exhibits the FPPase fold despite insignificant amino acid sequence identity [21] ( Figure 3A). Clearly, the permissiveness of the biosynthetic chaperone is easily cajoled by mutagenesis in nature or in the laboratory to allow for the generation of new terpenoid products.…”

SQ109, a New Drug Lead for Chagas Disease

“…In addition, it acts as an uncoupler, collapsing pH gradients (⌬pH) and membrane potentials (⌬) in bacterial systems (24), thereby reducing ATP synthesis. In unrelated work, we also reported (25) that SQ109 was an inhibitor of dehydrosqualene synthase (from Staphylococcus aureus), a protein whose three-dimensional structure (26) and mechanism of action (27) are very similar to those of squalene synthase (SQS) (28), which catalyzes the first step of sterol biosynthesis in eukaryotes, such as T. cruzi, suggesting that SQ109 might also inhibit SQS.…”