Farnesylation and Geranylation

The isoprenylation occurs at the Cys-residue of the consensus sequence Cys-A-A-X-COOH, whereby the nature of the C-terminal X-residue determines whether farnesylation or geranylation occurs (Fig.3.14). After the isoprenoid residue is appended, the three C-terminal residues are removed by a prenylation-dependent endopeptidase and the new COOH-group of the Cys-residue is methylated to increase the hydrophobicity of the C-terminus. 

The isoprenoid modification can be found, among others, on the Ras protein and other members of the Ras superfamily (see Chapter 9), as well as with the a-subunit of 

G-protein (see Chapter 5). The / -complex of G-proteins is also associated with the membrane via geranylation. A twofold geranylation is found on two Cys residues of the Rab protein (see Chapter 9). 

In addition to promoting membrane association, other functional aspects of protein prenylation have been appreciated (review Sinensky, 2000). Prenylation can also serve to mediate protein-protein interactions and has a role in protein trafficking. 

A lipid anchor can also be used to attach proteins to the extracellular side of cells. Cell surface proteins that mediate cell-cell interactions can be specifically anchored to the outward side of cells by means of a glycosl-phosphatidyl-inositol moiety. 

---

Certain proteins such as Ras and transducin undergo the addition of iso-prenoid groups attached to cysteine residue of the protein. Isoprenoids such as farnesyl and geranyl are 15- and 20-carbon substances, respectively. 

Farnesyl and geranyl groups can form covalent bonds with proteins, particularly the G proteins and certain protooncogene products involved in signal transduction. These hydrophobic groups anchor the proteins in the ceU membrane. 

Cholesterol, which is present in brain and in almost all tissues, is synthesised from isopentenyl pyrophosphate via squalene, farnesyl and geranyl pyrophosphates. The synthesis of squalene commences with the isomerisation of isopentenyl pyrophosphate to dimethylallyl pyrophosphate, after which successive condensations take place according to Equation 11.118. The hydrocarbon squalene is then transformed into the tetracyclic steroidal configuration of lanosterol by appropriate enzymes, and this is followed by conversion into cholesterol. Cholesterol is the precursor to most other steroids in the body. 

Whiie the esters (and lactones) mentioned above may (or may not) be mixtures, the farnesyl and geranyl esters found in Andrena are not. They appear to all be of all-tra/is-farnesol and devoid of esters of the other farnesol isomers. Simiiarly the geranyl esters are free of their neryl counterparts. The specific isomer of dihydrofarnesyl acetate from Melissodes is unknown although 2,3-dihydro-6-tra 5-farnesol has been found in a Bombus jonellus cephalic gland (Bergstrom and Svensson, 1973a). 

Bergstrdm, G. and Tengd, J. (1974) Studies on natural odoriferous compounds IX. Farnesyl- and geranyl esters as main volatile constituents of the secretion from Dufour s gland in 6 species of Andrena (Hymenoptera, Apidae). Chem, Scr.y 5, 28-38. 

C-racemization of phosphorothioylated/phosphorylated amino acids (Scheme 23). An oxathiaphospholane approach to one-pot phosphorothioylation of isoprenoid alcohols such as allyl, geranyl, isopentenyl, citronellyl, farnesyl, and phytyl alcohols has also been reported (Scheme 24). ... 

In two important papers Poulter et al. have published their results on the compelling evidence for a stepwise mechanism for the 1 -4 condensation reaction between isopentenyl pyrophosphate and geranyl pyrophosphate to form farnesyl... 

TengO, J. and BergstrOm, G. (1975) PAX-trans farnesyl hexanoate and geranyl octanoate in the Dufour gland secretion of Andrena (Hymenoptera Apidae). J. Chem. EcoL, 1, 253-68. 

Mevalonate kinase [E.C.2.7.1.36 (5) -> (6)] preparations from several plants are inhibited by geranyl, geranylgeranyl, farnesyl, and phytyl pyrophosphates, the most potent inhibitors being the latter two compounds.This enzyme may, therefore, be a control point of isoprenoid biosynthesis in plants. A cell-free system for conversion of mevalonic acid (5) into mevalonic acid S-pyro-phosphate (7) has been prepared from the sapogenin-producing plant Agave americana. ... 

C2]-Squalene, 80, has been produced71 in the reaction sequence shown in equation 31 which involves alkylation of 3-13C-ethyl acetoacetate with geranyl bromide, followed by hydrolysis, decarboxylation and treatment with triethyl phosphonoacetate and then reduction of the ester 82 with LiAlHr, bromination with CBr4/PPh3 and coupling the farnesyl bromide with Cul/Li-pyrrolidine. Epoxidation of 80 has been effected by... 

FIGURE 1.4 Proposed biosynthetic route for the biosynthesis of (A) squalene oxide (squalene-2,3-oxide) via the isoprenoid pathway and (B) triterpene saponins of the dammarane-type and oleanane-type from squalene oxide. PP, diphosphate group GPS, geranyl phosphate synthase FPS, farnesyl phosphate synthase NADPH, nicotinamide adenine dinucleotide phosphate. 

In Scheme 26 a selection of lipidated peptide sequences synthesized via this approach is shown. A variety of N- and H-Ras-derived peptide sequences was obtained bearing different types of lipids like farnesyl, palmitoyl, and geranylgeranyl and a fluorescent-labeled geranyl group. Furthermore, fluorescent markers like nitroben-zoxadiazole and photoactivatable groups were introduced. 

Geranyl diphosphate and farnesyl diphosphate are analogues of dimethylallyl diphosphate that contain two and three C5 subunits respectively they can undergo exactly the same SnI reactions as does dimethylallyl diphosphate. In all cases, a carbocation mechanism is favoured by the resonance stabilization of the allylic carbocation. Dimethylallyl diphosphate, geranyl diphosphate, and farnesyl diphosphate are precursors for natural terpenoids and steroids. 

An exactly analogous process can then occur, in which geranyl diphosphate provides the allylic cation, and a further molecule of isopentenyl diphosphate adds on, giving farnesyl diphosphate this can subsequently yield geranylgeranyl diphosphate. 

The compounds geranyl diphosphate, farnesyl diphosphate, and geranylgeranyl diphosphate are biochemical precursors of monoterpenes, sesquiterpenes, and diterpenes respectively, and virtually all subsequent modifications of these precursors involve initial formation of an allylic cation through loss of diphosphate as the leaving group. 

Formation of squalene. Isopentenyl diphosphate undergoes isomerization to form dimethylallyl diphosphate. The two C5 molecules condense to yield geranyl diphosphate, and the addition of another isopentenyl diphosphate produces farnesyl diphosphate. This can then undergo dimerization, in a head-to-head reaction, to yield squalene. Farnesyl diphosphate is also the starting-point for other polyisoprenoids, such as doli-chol (see p. 230) and ubiquinone (see p. 52). 

---