Lanosterol to Cholesterol

3y6-Hydroxysteroid-A -reductase (also known as sterol- i -reductase)  

3j6-Hydroxysteroid-A, A -isomerase (commonly known as sterol-A -isomerase)  

Sterol-A -isomerase deficiency, known as Conradi-HUnermann syndrome (CDPX2), is an X-linked dominant disorder. Clinical manifestations of this disorder include skeletal abnormalities, chondrodysplasia punctata, craniofacial anomalies, cataracts, and skin abnormalities. The 7-dehydrocholesterol reductase deficiency, known as Smith-Lemli-Opitz syndrome (SLO) is an autosomal recessive disorder occurring in about 1 in 20,000 births. Clinical manifestations of affected individuals include craniofacial abnormalities, microcephaly, congenital heart disease, malformation of the limbs, psychomotor retardation, cerebral maldevelopment, and urogenital anomalies. Measurement of 7-dehydrocholesterol in amniotic fluid during second trimester or in neonatal blood specimen has been useful in the identification of the disorder. The sterol-A -reductase deficiency causes a developmental phenotype similar to SLO syndrome and is associated with accumulation of desmosterol. The inability of de novo fetal synthesis of cholesterol combined with its inadequate transport from the mother to the fetus appears 

---

The conversion of lanosterol to cholesterol involves 19 steps and is described in the article Cholesterol Biosynthesis Lanosterol to Cholesterol on pp 377-384 in the March 2002 issue of the Journal of Chemical Education... 

Conversion of Lanosterol to Cholesterol Requires 20 Additional Steps... 

Although lanosterol may appear similar to cholesterol in structure, another 20 steps are required to convert lanosterol to cholesterol (Figure 25.35). The enzymes responsible for this are all associated with the endoplasmic reticulum. The primary pathway involves 7-dehydroeholesterol as the penultimate intermediate. An alternative pathway, also composed of many steps, produces the intermediate desmosterol. Reduction of the double bond at C-24 yields cholesterol. Cholesterol esters—a principal form of circulating cholesterol—are synthesized by acyl-CoA cholesterol acyltransferases (ACAT) on the cytoplasmic face of the endoplasmic reticulum. 

C]-(35)-2,3-epoxysqualene and its racemate have been prepared by two routes in a metabolically non-labile position relative to the demethylation of lanosterol to cholesterol (equation 70 and 71). The racemic [24,30-14C]-2,3-epoxysqualene, 192, has been obtained163 by condensation of (35, 3/ )-2,3-epoxytrisnorsqualene aldehyde 193 with freshly prepared 14C-labelled isopropylidenephosphorane, 194 (equation 70). 

The compensatory effect of cholesterol observed in D407 cells have also been demonstrated in other cell lines (Cho et al. 1998 Holleran et al. 1998) and may well be a consequence of tamoxifen-induced severe inhibition of lanosterol (to cholesterol)-converting enzymes. In rat liver preparations and CHO cells, sterol A8-isomerase (IC50 0.21-0.15. iM) was the most sensitive... 

In addition to the enzymes that are embedded in the membranes of the ER, conversion of lanosterol to cholesterol depends upon soluble cytoplasmic carrier proteins.174 See also Box 21-A. Other sterols formed in... 

Figure 22-7 Conversion of lanosterol to cholesterol. Two of many possible sequences are shown.

The conversion of lanosterol to cholesterol involves removal of the three methyl groups at the 4,4- and 14-positions, shift of the double bond at the B/C junction to between C5 and C6, and reduction of the C24-C25 double bond. The methyl groups are indicated by tracer experiments to be eliminated by oxidation to carbon dioxide. 

From Lanosterol to Cholesterol Takes Approximately 20 Steps Summary of Cholesterol Biosynthesis Lipoprotein Metabolism... 

The last sequence of reactions in the biosynthesis of choles-terol involves approximately 20 enzymatic steps, starting with lanosterol. In mammals the major route involves a series of double-bond reductions and demethylations (fig. 20.10). The sequence of reactions involves reduction of the A24 double bond, the oxidation and removal of the 14a methyl group followed by the oxidation and removal of the two methyl groups at position 4 in the sterol. The final reaction is a reduction of the A7 double bond in 7-dehydro-cholesterol. An alternative pathway from lanosterol to cholesterol also exists. The enzymes involved in the transformation of lanosterol to cholesterol are all located on the endoplasmic reticulum. 

Two pathways for the conversion of lanosterol to cholesterol. The major route in mammals proceeds through 7-dehy drocho l es terol. 

All 27 carbon atoms of cholesterol are derived from acetyl Co A. First acetyl CoA and acetoacetyl CoA combine to form HMG CoA which, in turn, is reduced to mevalonate by HMG CoA reductase. Mevalonate is converted into the five-carbon isoprene compounds 3-isopentenyl pyrophosphate and its isomer dimethylallyl pyrophosphate. These two compounds condense to form the CIO geranyl pyrophosphate, which is elongated to the C15 farnesyl pyrophosphate by the addition of another molecule of isopentenyl pyrophosphate. Two molecules of farnesyl pyrophosphate condense to form the C30 squalene, which is then converted via squalene epoxide and lanosterol to cholesterol. 

Glucose is oxidized in the pentose phosphate pathway (Chap. 11), to produce NADPH that can enter cholesterol synthesis. One molecule of glucose is required per molecule of lanosterol synthesized. (The reactions that convert lanosterol to cholesterol are outside the scope of Chap. 13.)... 

Figure 2 In the first steps of the conversion of lanosterol to cholesterol, an enzyme, cytochrome P-4S0, oxidizes the three methyl groups that will be removed while leaving the rest of the molecule untouched, because of geometric control that is typical in enzymes.

Fig. 20 Steps in the pathway from lanosterol to cholesterol in mammals.

As shown in Figure 6.13, squalene is converted in two steps to lanosterol. Conversion of lanosterol to cholesterol involves 19 steps that are not shown. (Although cholesterol contains several rings, it is not an aromatic compoimd because its rings do not resonate.)... 

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... 

Transformation of lanosterol to cholesterol (Figure 19-16) is a complex, multistep process catalyzed by enzymes of the endoplasmic reticulum (microsomes). A cytosolic sterol carrier protein is also required and presumably functions as a carrier of steroid intermediates from one catalytic site to the next but may also affect activity of the enzymes. The reactions consist of removal of the three methyl groups attached to C4 and C14, migration of the double bond from the 8,9- to the 5,6-position, and saturation of the double bond in the side chain. Conversion of lanosterol to cholesterol occurs principally via 7-dehydrocholesterol and to a minor extent via desmosterol. 

The importance of cholesterol biosynthesis in embryonic development and formation of the central nervous system is reflected in patients with disorders in the pathway for the conversion of lanosterol to cholesterol. Three enzyme deficiencies have been identified (Figure 19-16) ... 

A partial pathway for the conversion of lanosterol to cholesterol. The complete process consists of 19 steps. The C24 = C25 double bond can be reduced by 3 j8-hydroxylsteroid-A -reductase at several steps along the pathway (indicated by I), and deficiency of this enzyme leads to accumulation of desmosterol. Deficiency of enzyme 2 and enzyme 3 results in CDPX2 and SLO syndromes, respectively (see text). 

Cytochrome P450 enzyme system The cytochromes P450 are mixed-function oxidases that require both NADPH and O. They are involved in a number of reactions in the conversion of lanosterol to cholesterol, as well as important steps in the synthesis of steroid hormones. Cytochromes P450 are very important in the detoxification of xenobiotics and in the metabolism of drags. 

Step 4 A series of enzyme-catalyzed reactions converts lanosterol to cholesterol. The three highlighted methyl gronps in the stmctnral formnla of lanosterol are lost via separate multistep operations, the C-8 and C-24 double bonds are reduced, and a new double bond is introduced at C-5. 

There are approximately 20 enzymatic steps from lanosterol to cholesterol or ergosterol, and probably as many from 24-methylenedihydrolanosterol to ergosterol. 

Nineteen discrete reactions are used to convert lanosterol to cholesterol. All of the enzymes necessary for these reactions are embedded in the microsomal membrane. Some of these proteins have been solubilized and purified. However, many questions of the mechanism and enzymology of these reactions remain to be answered. 

For the conversion of lanosterol to cholesterol, there are a number of excellent reviews that consider the sequence of events and the participants therein [1,2,5,105]. 

The enzymes responsible for the conversion of lanosterol to cholesterol, as were those for the conversion of farnesyl pyrophosphate to squalene and lanosterol, are all integral membrane-bound proteins of the endoplasmic reticulum. Many have resisted solubiUzation, some have been partially purified, and several have been obtained as pure proteins. As a consequence, much of the enzymological and mechanistic studies have been done on impure systems and one would anticipate a more detailed and improved understanding of these events as more highly purified enzymes become available. Many approaches have been taken to establish the biosynthetic route that sterols follow to cholesterol. Some examples are synthesis of potential intermediates, the use of inhibitors, both of sterol transformations and of the electron transfer systems, and by isotope dilution experiments. There is good evidence that the enzymes involved in these transformations do not have strict substrate specificity. As a result, many compounds that have been found to be converted to intermediates or to cholesterol may not be true intermediates. In addition, there is structural similarity between many of the intermediates so that alternate pathways and metabolites are possible. For example, it has been shown that side-chain saturation can be either the first or the last reaction in the sequence. Fig. 21 shows a most probable series of intermediates for this biosynthetic pathway. 

Previous investigations from several laboratories have demonstrated that both microsomal membranes and the cytosolic fraction from rat hver are required for the biological synthesis of cholesterol [1-4]. Specifically, the following conversions have been reported to require both microsomes and cytosol acetate to cholesterol [4] squalene to cholesterol [1] squalene-2,3-oxide to lanosterol [3] lanosterol to cholesterol [1,5] A -cholestenol to cholesterol [6] lanosterol to dihydrolanosterol [7] various 4,4-dimethyl sterols to cholesterol [8] and 7-dehydrocholesterol to cholesterol [9,10]. 

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. 

Sterol carrier protein 2 (SCP2) participates in the enzymatic conversion of lanosterol to cholesterol by rat liver microsomal enzymes. 

---