Protein prenylation: enzymes, therapeutics, and biotechnology applications

Abstract

Protein prenylation is a ubiquitous covalent post-translational modification found in all eukaryotic cells, comprising attachment of either a farnesyl or a geranylgeranyl isoprenoid. It is essential for the proper cellular activity of numerous proteins, including Ras family GTPases and heterotrimeric G-proteins. Inhibition of prenylation has been extensively investigated to suppress the activity of oncogenic Ras proteins to achieve antitumor activity. Here, we review the biochemistry of the prenyltransferase enzymes and numerous isoprenoid analogs synthesized to investigate various aspects of prenylation and prenyltransferases. We also give an account of the current status of prenyltransferase inhibitors as potential therapeutics against several diseases including cancers, progeria, aging, parasitic diseases, and bacterial and viral infections. Finally, we discuss recent progress in utilizing protein prenylation for site-specific protein labeling for various biotechnology applications.

Figure 1

(A) Structures of 1 (farnesyl diphosphate, FPP) and 2 (geranylgeranyl diphosphate, GGPP). (B) Reactions catalyzed by prenyltransferase enzymes.

Figure 2

Three-step prenylation processing of proteins. Proteins undergo farnesylation and proteolytic cleavage of aaX residues, followed by carboxymethylation, and then get localized at the plasma membrane. Some proteins, shown here N-Ras, undergo palmitoylation and then localize to plasma membrane, while other proteins, shown here K-Ras, have a polybasic sequence upstream of the “CaaX box” facilitating membrane localization.

Figure 3

Crystal structures of prenyltransferase enzymes. (A) Crystal structure of FTase in complex with a nonhydrolyzable FPP analog and a peptide substrate based on KRas-4B (PDB 1D8D): magenta, α-subunit; blue, β-subunit; cyan, isoprenoid analog; green, CaaX peptide. (B) Binding pocket of FPP showing interaction of protein and isoprenoid substrates over a large surface area: gray, space-fill structure of β-subunit of FTase; cyan, isoprenoid analog; green, CaaX peptide. (C) Crystal structure of GGTase-II in complex with Rab escort protein and FPP (PDB 1LTX): magenta, GGTase-II α-subunit; blue, GGTase-II β-subunit; green, Rab escort protein; cyan, FPP.

Figure 4

Structures of isoprenoid analogs used to probe mechanism of prenyltransferase enzymes.

Figure 5

Key features of catalysis by protein farnesyltransferase. (A) Schematic representation of transition state showing thiol activation by Zn²⁺, diphosphate stabilization by Mg²⁺, and partial bonding to leaving group and incoming nucleophile (adapted from ref (23)). (B) Structural model for transition state based on kinetic isotope effect measurements and DFT calculations. The model reaction used for computation (shown in these images) employed ethanethiol and dimethylallyl diphosphate. (C) Electrostatic potential map of transition state based on the same model shown in panel B (images B and C images adapted from ref (24)). Color scheme for B: carbon (green), hydrogen (white), oxygen (red), phosphorus (magenta), and sulfur (yellow). Color scheme for C: red represents more negative potential, blue represents less negative potential, and green is intermediate.

Figure 6

Structures of isoprenoid analogs used to study structure, mechanism, and isoprenoid substrate specificity of FTase and GGTase-I.

Figure 7

Chemical proteomic strategy for analysis of the prenylated proteome.

Figure 8

Structures of isoprenoid analog used to analyze prenylated proteome.

Figure 9

Structures of FTIs and GGTI investigated in clinical trials against cancer or HGPS.