Squalene to Lanosterol

Cyclization of squalene to lanosterol. Supernatant protein factor (SPF), a cytosolic protein, promotes both stages of the cyclization. 

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Steroids are heavily modified triterpenoids that are biosynthesized in living organisms from farnesyl diphosphate (Cl5) by a reductive dimerization to the acyclic hydrocarbon squalene (C30), which is converted into lanosterol (Figure 27.12). Further rearrangements and degradations then take place to yield various steroids. The conversion of squalene to lanosterol is among the most... 

Synthesis of endogenic cholesterol is also controlled by exogenous cholesterol supplied in food the more dietary cholesterol is digested, the less endogenic cho-lesterol is produced in the liveV. Exogenous cholesterol inhibits the activity of hydroxymethylglutaryl-CoA reductase and the cyclization of squalene to lanosterol. 

The sequence of transformations from squalene to lanosterol begins by the enzymatic oxidation of the 2,3-double bond of squalene to form (3S)-2,3-oxidosqualene [also called squalene 2,3-epoxide]. 

The application of this procedure to the fused polycyclic compound E, which already has a linear dual and only the last two steps (iii-iv) apply to it, leads to a linear acyclic structure F which may be traced back to the biogenetic cyclisation of squalene to lanosterol via cationic intermediates, as well as to the stereospecific cationic cyclisation of polyolefins studied by Johnson [18]. 

A general type of chemical reaction between two compounds, A and B, such that there is a net reduction in bond multiplicity (e.g., addition of a compound across a carbon-carbon double bond such that the product has lost this 77-bond). An example is the hydration of a double bond, such as that observed in the conversion of fumarate to malate by fumarase. Addition reactions can also occur with strained ring structures that, in some respects, resemble double bonds (e.g., cyclopropyl derivatives or certain epoxides). A special case of a hydro-alkenyl addition is the conversion of 2,3-oxidosqualene to dammara-dienol or in the conversion of squalene to lanosterol. Reactions in which new moieties are linked to adjacent atoms (as is the case in the hydration of fumarate) are often referred to as 1,2-addition reactions. If the atoms that contain newly linked moieties are not adjacent (as is often the case with conjugated reactants), then the reaction is often referred to as a l,n-addition reaction in which n is the numbered atom distant from 1 (e.g., 1,4-addition reaction). In general, addition reactions can take place via electrophilic addition, nucleophilic addition, free-radical addition, or via simultaneous or pericycUc addition. 

Squalene is an important biological precursor of many triterpenoids, one of which is cholesterol. The first step in the conversion of squalene to lanosterol is epoxidation of the 2,3-douhle bond of squalene. Acid-catalysed ring opening of the epoxide initiates a series of cyclizations, resulting in the formation of protesterol cation. Elimination of a C-9 proton leads to the 1,2-hydride and 1,2-methyl shifts, resulting in the formation of lanosterol, which in turn converted to cholesterol by enzymes in a series of 19 steps. 

Figure 22-6 The cyclization of all-frans squalene to lanosterol and cycloartenol.

Isotopic labeling experiments show that cholesterol is derived from ethanoate by way of squalene and lanosterol. The evidence for this is that homogenized liver tissue is able to convert labeled squalene to labeled lanosterol and thence to labeled cholesterol. The conversion of squalene to lanosterol is particularly interesting because, although squalene is divisible into isoprene units, lanosterol is not—a methyl being required at C8 and not C13 ... 

As a result, some kind of rearrangement must be required to get from squalene to lanosterol. The nature of this rearrangement becomes clearer if we write the squalene formula so as to take the shape of lanosterol ... 

The series of processes involved in the biosynthetic conversion of squalene to lanosterol in Figure 27.6 is written in a stepwise format for convenience, but the cyclization sequence in steps 2-4 appears to take plaa at one time without intermediates. Similarly, the carbocation rearrangements (Section 6.12) in steps 7-10 take place at essentially the same time without intermediates. 

This step comprises cyclization of squalene to lanosterol (the first sterol to be formed) and conversion of lanosterol to cholesterol. The cyclization begins with conversion of squalene to squalene-2,3-epoxide by a microsomal mixed-function oxidase that requires O2, NADPH, and FAD (Figure 19-15). Cyclization of squalene-2,3-epoxide to lanosterol occurs by a series of concerted 1,2-methyl group... 

Investigations conducted in our laboratory [3] and by Ritter and Dempsey [11] resulted in the proposal that a sterol carrier protein, present in rat liver cytosol, was required for the conversion of squalene to cholesterol by liver microsomal enzymes. Later, it was shown that rat liver cytosol contains two proteins which are required for the microsomal conversion of squalene to cholesterol [5,12]. Sterol carrier proteinj (SCPi) participates in the microsomal conversion of squalene to lanosterol, while sterol carrier protein 2 (SCP2) participates in the microsomal conversion of lanosterol to cholesterol. In addition, SCP2 also participates in key steps in the utilization of cholesterol as well as in the intracellular transfer of cholesterol between cellular organelles. 

SCPi participates in the enzymatic conversion of squalene to lanosterol by rat liver microsomes (cf. Chapter 1). It has been purified both in our laboratory [13] and by Ferguson and Bloch [14]. In Dr. Bloch s laboratory, the protein has been called supernatant protein factor or SPF [14-20]. It is a single polypeptide chain with a molecular weight of approximately 47000 dalton [14] (Table 1). The protein has an acidic p7. Recent evidence suggests that SCP] and SPF are one and the same protein [21]. 

By means of high resolution chromatographic techniques and mass spectrometry, it was shown that the major product (90%) formed when squalene was incubated with purified SCP, microsomes and cofactors (NADPH and FAD) was lanosterol [13]. While SCP] specifically activated the enzymatic conversion of squalene to lanosterol by liver microsomal membranes, it was not capable of activating reactions subsequent to the formation of lanosterol. Thus, the microsomal conversion of... 

These results demonstrate that SCPi is required for the conversion of squalene to lanosterol, even when squalene is enzymatically generated in situ bound to the microsomal membranes [24]. Furthermore, the results show excellent correspondence between experiments in which small quantities (less than 2 vol%) of organic solvent are used to add squalene to enzyme incubations [2,3,12,13] and experiments in which no organic solvent is employed. 

Fig. 8 is a schematic diagram of a cell which shows the known sites in which sterol carrier proteins are involved in cholesterol biosynthesis, utilization and intracellular transfer. SCP, participates in the conversion of squalene to lanosterol and SCP2 participates in the conversion of lanosterol to cholesterol, the conversion of cholesterol to cholesterol ester by ACAT, and probably also in the conversion of cholesterol to 7a-hydroxycholesterol. SCPj transfers cholesterol from cytoplasmic lipid inclusion droplets to mitochondria in the adrenal and SCPj also translocates cholesterol from the outer to the inner mitochondrial membrane. 

Structure Voser et at., ibid. 35 2414(1952) Barnes et af.t J. Chem. Soc. 1953 571, Stereochemistry eidem, ibid. 1953, 576, Prepn from isocholesterol Bloch, Urcch, Bio-Chem. Prepn. 6, 32 (t958). Prepn by cyclization of squalener Cornforth et at, Ciba Foundation Symposium of Terpenes and Sterols 1958 119 van Tamelen et a 1. J. Am. Chem. Soc. 88 4752 (1966). Mechanism of the squalene to lanosterol conversion eidem, ibid, l04 6479, 64 0 (1982). 

Stage 3 Cyclization of Squalene to Lanosterol and Its Conversion to Cholesterol (Figure 19.22)... 

The first step in the conversion of squalene to lanosterol is epoxidation of the... 

So far we have dwelt on the over-all reaction, based on the concept of titanium (II), and the chemistry at this point appears very complicated. One would like to see some clarification of the over-all mechanism. We were pleased to find that one can, in this titanium series, eflFectively separate the complexation and the reduction steps (li). This is as important as the separation of the oxidation and the cyclization steps in the over-all conversion of squalene to lanosterol, a very knotty and complicated problem in the biological province. 

Shibata, N., Arita, M., Misaki, Y., Dohmae, N., Takio, K., Ono, T., Inoue, K., and Arai, H., Supernatant protein factor, which stimulates the conversion of squalene to lanosterol, is a cytosolic squalene transfer protein and enhances cholesterol biosynthesis, Proc. Natl. Acad. Sci. U.SA. 98 (5), 2244-2249., 2001. 

The road from squalene to lanosterol, the fully cyclized precursor to cholesterol, and other steroids passes through (35)-2,3-oxidosqualene (squalene oxide). The... 

The electrophilic additions of water and hydrogen halides all involve nucleophilic attack on a carbenium ion intermediate. Many nucleophiles can add in a similar manner, and in fact, other alkenes can act as nucleophiles and add to carbenium ions, creating another carbenium ion. This is the basis of cationic polymerization (see Chapter 13). Nature exploits the addition of one alkene to another to create complex polycyclic natural products. As shown below, the conversion of squalene to lanosterol involves a cascade of alkene additions. The enzyme squalene oxi-... 

Cyclic Forms of Saccharides and Concerted Proton Transfers 545 Squalene to Lanosterol 550... 

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