Squalene cyclases

Head-to-head condensation of two farnesylpyrophosphate (C]3—PP) molecules yields a G13-cyclopropane (C3)-CH intermediate which is then reduced to yield squalene H(CH2-C(CH3)=CH-CH2)3-(CH2-CH=C(CH3)CH2)3 (C30), that is, if one represents the isoprene polarities as IP and PI, one could represent squalene as (IP)3—(PI)3. Squalene is subsequently oxidized [via a squalene monooxygenase] to yield squalene 2,3-epoxide which is cyclized to the tetracyclic sterol terpene lanosterol (C30) [via squalene cyclase]. 

Cationic cyclizations of polyolefins to oligocyclic terpenes such as that catalyzed by squalene-cyclases in the biosynthesis of steroids have long fascinated chem-... 

Table I. Properties of Purified (Oxido)Squalene Cyclase Enzymes...

Squalene monooxygenase, an enzyme bound to the endoplasmic reticulum, converts squalene to squalene-2,3-epoxide (Figure 25.35). This reaction employs FAD and NADPH as coenzymes and requires Og as well as a cytosolic protein called soluble protein activator. A second ER membrane enzyme, 2,3-oxidosqualene lanosterol cyclase, catalyzes the second reaction, which involves a succession of 1,2 shifts of hydride ions and methyl groups. 

The second part of lanosterol biosynthesis is catalyzed by oxidosqualene lanosterol cyclase and occurs as shown in Figure 27.14. Squalene is folded by the enzyme into a conformation that aligns the various double bonds for undergoing a cascade of successive intramolecular electrophilic additions, followed by a series of hydride and methyl migrations. Except for the initial epoxide protonation/cyclization, the process is probably stepwise and appears to involve discrete carbocation intermediates that are stabilized by electrostatic interactions with electron-rich aromatic amino acids in the enzyme. 

CATTEL, L., CERUTI, M., 2,3-Oxidosqualene cyclase and squalene epoxidase Enzymology, mechanism and inhibitors. In Physiology and Biochemistry of Sterols (G.W. Patterson and W.D. Nes, eds,), American Oil Chemists Society, Champaign. 1992, pp. 50-82. 

PORALLA, K HEWELT, A., PRESTWICH, G.D., ABE, I., REIPEN, I., SPRENGER, G., A specific amino acid repeat in squalene and oxidosqualene cyclases, TIBS, 1994, 19,157-158. 

Currently there is no experimentally determined three-dimensional structural information available for OSCs, although studies with a related enzyme, squa-lene-hopene cyclase (SC EC 5.4.99.7) have proved informative. SCs are involved in the direct cyclisation of squalene to pentacyclic triterpenoids known as hopanoids, which play an integral role in membrane structure in prokaryotes [ 51 ]. A number of SC genes have been cloned from bacteria [52 - 54]. The SC and OSC enzymes have related predicted amino acid sequences, and so should have similar spatial structures [55]. The crystal structure of recombinant SC from the Gram-positive bacterium Alicyclobacillus acidocaldarius has established that the enzyme is dimeric [55]. Each subunit consists of two a-a barrel domains that assemble to form a central hydrophobic cavity [55,56]. 

The activity of 2,3-oxidosqualene cyclases is associated with microsomes, indicating their membrane-bound nature. However, the predicted amino acid sequences of these enzymes generally lack signal sequences and obvious transmembrane domains. Addition of hydrophobic membrane-localising regions to OSCs during evolution may have removed selection pressures that maintained alternate mechanisms for membrane localisation [33]. Consistent with this, there is a non-polar plateau on the surface of the A. acidocaldarius SC enzyme which is believed to be immersed in the centre of the membrane. The squalene substrate for SC is likely to diffuse from the membrane interior into the central cavity of the enzyme via this contact region [55,56]. 

Alicyclobacillus Squalene-hopene cyclase (SC). . raveyllscEkdeByI.. mekirryllheBredBt. . ypdvBdBavvvwa.. rraveylkreBkpdBsB.. kaldwveqhBnpdBgB.. acidocaldarius... 

Cattel L, Ceruti M (1992) 2,3-Oxidosqualene cyclase and squalene epoxidase enzymology,... 

Fig. 5.1.2 Cholesterol biosynthesis branch of the isoprenoid biosynthetic pathway. Enzymes are numbered as follows 1 squalene synthase 2 squalene epoxidase 3 2,3-oxidosqua-lene sterol cyclase 4 sterol A24-reductase (desmosterolosis) 5 sterol C-14 demethylase 6 sterol A14-reductase (hydrops-ectopic calcification-moth-eaten, HEM, dysplasia) 7 sterol C-4 demethylase complex (including a 3/ -hydroxysteroid dehydrogenase defective in congenital hemidyspla-sia with ichthyosiform nevus and limb defects, CHILD, syndrome) 8 sterol A8-A7 isomerase (Conradi-Hunermann syndrome CDPX2) 9 sterol A5-desaturase (lathosterolosis) 10 sterol A7-reductase (Smith-Lemli-Opitz syndrome). Enzyme deficiencies are indicated by solid bars across the arrows...

The transformation of squalene into lanosterol. The squalene monooxygenase reaction requires 02, NADPH, FAD, phospholipid, and a cytosolic protein. The cyclase reaction has no known cofactor requirements. The reaction proceeds by means of a protonated intermediate that undergoes a concerted series of trans-1,2 shifts of methyl groups and hydride ions to produce lanosterol. 

Abe I and Prestwich GD (1999) Squalene epoxidase and oxidosqualenedanosterol cyclase-key enzymes in cholesterol biosynthesis. Comprehensive Natural Products Chemistry, Vol 2. Elsevier, Amsterdam, pp 267-298. 

Fig. 14.12. Enzymatic transformation of acyclic squalene oxide (A) into tetracyclic lano-sterol (G). The oxidosqualene-lanosterol cyclase controls the conformation of the substrate so effectively that only one out of 64 possible diastereomers is formed.

VOGELI, U., CHAPPELL, J., Induction of sesquiterpene cyclase and suppression of squalene synthetase activities in plant cell cultures treated with fungal elicitor, Plant Physiol., 1988, 88, 1291-1296. 

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