Geranyltranstransferase

 

Geranyltranstransferase is a member of the Isoprenoid Synthase Type I superfamily. It catalyses the metal-dependent formation of (2E,6E)-farnesyl diphosphate from geranyl diphosphate + isopentenyl diphosphate. It is known that the residue at the fifth position before the first DDxxD motif [PMID:12135472], in this case a phenylalanine, is responsible for determining product chain length.

 

Reference Protein and Structure

Sequence
P08836 UniProt (2.5.1.1, 2.5.1.10) IPR000092 (Sequence Homologues) (PDB Homologues)
Biological species
Gallus gallus (Chicken) Uniprot
PDB
1fps - CRYSTAL STRUCTURE OF RECOMBINANT FARNESYL DIPHOSPHATE SYNTHASE AT 2.6 ANGSTROMS RESOLUTION (2.6 Å) PDBe PDBsum 1fps
Catalytic CATH Domains
1.10.600.10 CATHdb (see all for 1fps)
Cofactors
Magnesium(2+) (3) Metal MACiE
Click To Show Structure

Enzyme Reaction (EC:2.5.1.10)

isopentenyl diphosphate(3-)
CHEBI:128769ChEBI
+
geranyl diphosphate(3-)
CHEBI:58057ChEBI
→
diphosphate(3-)
CHEBI:33019ChEBI
+
2-trans,6-trans-farnesyl diphosphate(3-)
CHEBI:175763ChEBI
Alternative enzyme names: Farnesyl pyrophosphate synthetase, Farnesyl-diphosphate synthase, Farnesylpyrophosphate synthetase, Geranyl transferase I, Prenyltransferase, FPP synthetase, Geranyltranstransferase,

Enzyme Mechanism

Introduction

The substrate undergoes heterolysis. The diphosphate product remains associated with the active site, and the cabocation is delocalised over the three terminal carbon atoms of the intermediate. The double bond of isopentenyl diphosphate adds to the terminal carbocation in an electrophilic addition. The diphosphate formed in the initial heterolysis deprotonates the carbon adjacent to the newly formed carbocation, forming a new double bond in the trans,trans-farnesyl diphosphate product.

Catalytic Residues Roles

UniProt PDB* (1fps)
Asp121 Asp121(102)A Forms part of both the Mg1 and Mg2 binding sites. Also helps stabilise the partially closed conformation. metal ligand
Asp257 Asp257(238)A Forms part of the Mg3 binding site. metal ligand
Asp258 Asp258(239)A Forms part of the Mg3 binding site via a water molecule. activator
Phe112 Phe112(93)A The residue at the fifth position before the first DDxxD motif, in this case Phe112 (Phe240 in E. coli) is responsible for determining product chain length [PMID:12135472]. steric role
Phe253 Phe253(234)A Helps hold the reactive intermediates in the correct position to ensure erroneous products aren't formed. steric role
Lys71 Lys71(52)A Stabilizes fully closed conformation via electrostatic interaction with C-terminal carboxylate of IPP. electrostatic stabiliser
Asp117 Asp117(98)A Forms part of both the Mg1 and Mg2 binding sites. metal ligand
Arg126 Arg126(107)A Hydrogen bonds to substrate diphosphate stabilising the negative charge. electrostatic stabiliser
Lys214, Asp188 Lys214(195)A, Asp188(169)A Helps stabilise the partially closed conformation. electrostatic stabiliser
*PDB label guide - RESx(y)B(C) - RES: Residue Name; x: Residue ID in PDB file; y: Residue ID in PDB sequence if different from PDB file; B: PDB Chain; C: Biological Assembly Chain if different from PDB. If label is "Not Found" it means this residue is not found in the reference PDB.

Chemical Components

dephosphorylation, heterolysis, overall reactant used, charge delocalisation, bimolecular electrophilic addition, proton transfer, overall product formed, native state of enzyme regenerated

References

  1. Hosfield DJ et al. (2004), J Biol Chem, 279, 8526-8529. Structural Basis for Bisphosphonate-mediated Inhibition of Isoprenoid Biosynthesis. DOI:10.1074/jbc.c300511200. PMID:14672944.
  2. Aaron JA et al. (2010), Pure Appl Chem, 82, 1585-1597. Trinuclear Metal Clusters in Catalysis by Terpenoid Synthases. DOI:10.1351/PAC-CON-09-09-37. PMID:21562622.
  3. Liang PH (2009), Biochemistry, 48, 6562-6570. Reaction kinetics, catalytic mechanisms, conformational changes, and inhibitor design for prenyltransferases. DOI:10.1021/bi900371p. PMID:19537817.
  4. Rondeau JM et al. (2006), ChemMedChem, 1, 267-273. Structural basis for the exceptional in vivo efficacy of bisphosphonate drugs. DOI:10.1002/cmdc.200500059. PMID:16892359.
  5. Liang PH et al. (2002), Eur J Biochem, 269, 3339-3354. Structure, mechanism and function of prenyltransferases. DOI:10.1046/j.1432-1033.2002.03014.x. PMID:12135472.
  6. Koyama T et al. (2000), Biochemistry, 39, 463-469. Intersubunit Location of the Active Site of Farnesyl Diphosphate Synthase:  Reconstruction of Active Enzymes by Hybrid-Type Heteromeric Dimers of Site-Directed Mutants†. DOI:10.1021/bi991621b. PMID:10631008.
  7. Tarshis LC et al. (1996), Proc Natl Acad Sci U S A, 93, 15018-15023. Regulation of product chain length by isoprenyl diphosphate synthases. DOI:10.1073/pnas.93.26.15018. PMID:8986756.

Step 1. The substrate undergoes heterolysis. The diphosphate product remains associated with the active site, and the cabocation is delocalised over the three terminal carbon atoms of the intermediate.

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Catalytic Residues Roles

Residue Roles
Asp258(239)A activator
Arg126(107)A electrostatic stabiliser
Asp121(102)A metal ligand
Asp117(98)A metal ligand
Asp257(238)A metal ligand
Lys214(195)A electrostatic stabiliser
Lys71(52)A electrostatic stabiliser
Asp188(169)A electrostatic stabiliser
Phe112(93)A steric role
Phe253(234)A steric role

Chemical Components

dephosphorylation, heterolysis, overall reactant used, charge delocalisation

Step 2. The double bond of isopentenyl diphosphate adds to the terminal carbocation in an electrophilic addition. The positive charge in the geranyl carbocation intermediate is delocalised over the three terminal carbons.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Asp188(169)A electrostatic stabiliser
Asp121(102)A metal ligand
Asp117(98)A metal ligand
Asp257(238)A metal ligand
Lys71(52)A electrostatic stabiliser
Arg126(107)A electrostatic stabiliser
Lys214(195)A electrostatic stabiliser
Phe112(93)A steric role
Phe253(234)A steric role

Chemical Components

ingold: bimolecular electrophilic addition

Step 3. The diphosphate formed in the initial heterolysis deprotonates the carbon adjacent to the newly formed carbocation, forming a new double bond in the trans,trans-farnesyl diphosphate product. The product of the reaction moves from the top Mg site to the bottom Mg site upon dissociation of the diphosphate product, ready for another round of elongation. The chain elongation reaction will continue until product chain has reached the residue located at the fifth position before the first DDxxD motif (Phe112 in this case)

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Asp121(102)A metal ligand
Asp117(98)A metal ligand
Asp257(238)A metal ligand
Phe112(93)A steric role
Phe253(234)A steric role
Lys71(52)A electrostatic stabiliser
Arg126(107)A electrostatic stabiliser
Asp188(169)A electrostatic stabiliser
Lys214(195)A electrostatic stabiliser

Chemical Components

proton transfer, overall product formed, native state of enzyme regenerated
Download: Image, Marvin File

Step 1. The substrate undergoes heterolysis. The diphosphate product remains associated with the active site, and the cabocation is delocalised over the three terminal carbon atoms of the intermediate.

Step 2. The double bond of isopentenyl diphosphate adds to the terminal carbocation in an electrophilic addition. The positive charge in the geranyl carbocation intermediate is delocalised over the three terminal carbons.

Step 3. The diphosphate formed in the initial heterolysis deprotonates the carbon adjacent to the newly formed carbocation, forming a new double bond in the trans,trans-farnesyl diphosphate product. The product of the reaction moves from the top Mg site to the bottom Mg site upon dissociation of the diphosphate product, ready for another round of elongation. The chain elongation reaction will continue until product chain has reached the residue located at the fifth position before the first DDxxD motif (Phe112 in this case)

Products.

Contributors

Gemma L. Holliday, Craig Porter, James W. Murray, Charity Hornby