Due to the enzyme's role in
cholesterol biosynthesis, there is interest in lanosterol synthase
inhibitors as potential cholesterol-reducing drugs, to complement existing
statins.[12]
Before the acquisition of the protein's
X-ray crystal structure,
site-directed mutagenesis was used to determine residues key to the enzyme's catalytic activity. It was determined that an
aspartic acid residue (D455) and two
histidine residues (H146 and H234) were essential to enzyme function. Corey et al. hypothesized that the aspartic acid acts by protonating the substrate's
epoxide ring, thus increasing its susceptibility to
intramolecular attack by the nearest
double bond, with H146 possibly intensifying the proton donor ability of the aspartic acid through
hydrogen bonding.[13] After acquisition of the
X-ray crystal structure of the enzyme, the role of D455 as a proton donor to the substrate's epoxide was confirmed, though it was found that D455 is more likely stabilized by hydrogen bonding from two
cysteine residues (C456 and C533) than from the earlier suggested histidine.[12]
Ring formation cascade
Epoxide protonation activates the substrate, setting off a cascade of ring forming reactions. Four rings in total (A through D) are formed, producing the
cholesterol backbone.[12] Though the idea of a concerted formation of all four rings had been entertained in the past, kinetic studies with
(S)-2,3-oxidosqualene analogs showed that product formation is achieved through discrete
carbocation intermediates (see Figure 1). Isolation of monocyclic and bicyclic products from lanosterol synthase mutants has further weakened the hypothesis of a concerted mechanism.[14][15] Evidence suggests that epoxide ring opening and A ring formation is concerted, though.[16]
Structure
Lanosterol synthase is a two-domain monomeric protein[10] composed of two connected (α/α) barrel domains and three smaller
β-structures. The enzyme
active site is in the center of the protein, closed off by a constricted channel. Passage of the (S)-2,3-epoxysqualene substrate through the channel requires a change in
protein conformation. In
eukaryotes, a
hydrophobic surface (6% of the total enzyme surface area) is the
ER membrane-binding region (see Figure 2).[12]
The enzyme contains five fingerprint regions containing
Gln-
Trp motifs, which are also present in the highly analogous bacterial enzyme
squalene-hopene cyclase.[12] Residues of these fingerprint regions contain stacked sidechains which are thought to contribute to enzyme stability during the highly
exergonic cyclization reactions catalyzed by the enzyme.[17]
Lanosterol synthase also catalyzes the cyclization of 2,3;22,23-diepoxysqualene to 24(S),25-epoxylanosterol,[18] which is later converted to 24(S),25-epoxycholesterol.[19] Since the enzyme affinity for this second
substrate is greater than for the monoepoxy (S)-2,3-epoxysqualene, under partial inhibition conversion of 2,3;22,23-diepoxysqualene to 24(S),25-epoxylanosterol is favored over
lanosterol synthesis.[20] This has relevance for disease prevention and treatment.
Clinical significance
Enzyme inhibitors as cholesterol-lowering drugs
Interest has grown in lanosterol synthase inhibitors as drugs to lower blood cholesterol and treat
atherosclerosis. The widely popular
statin drugs currently used to lower
LDL (low-density lipoprotein) cholesterol function by inhibiting
HMG-CoA reductase activity.[6] Because this enzyme catalyzes the formation of precursors far upstream of
(S)-2,3-epoxysqualene and cholesterol,
statins may negatively influence amounts of intermediates required for other biosynthetic pathways (e.g. synthesis of
isoprenoids,
coenzyme Q). Thus, lanosterol synthase, which is more closely tied to cholesterol biosynthesis than
HMG-CoA reductase, is an attractive drug target.[21]
Lanosterol synthase inhibitors are thought to lower
LDL and
VLDL cholesterol by a dual control mechanism. Studies in which lanosterol synthase is partially inhibited have shown both a direct decrease in
lanosterol formation and a decrease in
HMG-CoA reductase activity. The
oxysterol 24(S),25-epoxylanosterol, which is preferentially formed over
lanosterol during partial lanosterol synthase inhibition, is believed to be responsible for this inhibition of
HMG-CoA reductase activity.[22]
Evolution
It is believed that
oxidosqualene cyclases (OSCs, the class to which lanosterol cyclase belongs) evolved from bacterial squalene-hopene cyclase (SHC), which is involved with the formation of
hopanoids.
Phylogenetic trees constructed from the amino acid sequences of OSCs in diverse organisms suggest a single common ancestor, and that the synthesis pathway evolved only once.[23] The discovery of
steranes including
cholestane in 2.7-billion year-old shales from
Pilbara Craton,
Australia, suggests that
eukaryotes with OSCs and complex steroid machinery were present early in earth's history.[24]
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