Bornyl diphosphate synthase

 

Boronyl diphosphate synthase catalyses the conversion of geranyl diphosphate to boronyl diphosphate. It is a member of the monoterpene synthases, a group of enzymes that catalyse the metal-dependent cyclisation of geranyl diphosphate to a variety of monoterpene products via carbocation intermediates.

 

Reference Protein and Structure

Sequence
O81192 UniProt (4.2.3.116, 4.2.3.121, 5.5.1.8) IPR008949 (Sequence Homologues) (PDB Homologues)
Biological species
Salvia officinalis (garden sage) Uniprot
PDB
1n20 - (+)-Bornyl Diphosphate Synthase: Complex with Mg and 3-aza-2,3-dihydrogeranyl diphosphate (2.3 Å) PDBe PDBsum 1n20
Catalytic CATH Domains
1.10.600.10 CATHdb (see all for 1n20)
Cofactors
Magnesium(2+) (3) Metal MACiE
Click To Show Structure

Enzyme Reaction (EC:5.5.1.8)

geranyl diphosphate(3-)
CHEBI:58057ChEBI
→
(+)-bornyl diphosphate(3-)
CHEBI:57293ChEBI
Alternative enzyme names: (+)-bornylpyrophosphate cyclase, Geranyl-diphosphate cyclase, Bornyl pyrophosphate synthase, Bornyl pyrophosphate synthetase, (+)-bornyl-diphosphate lyase (decyclizing),

Enzyme Mechanism

Introduction

Boronyl diphosphate synthase catalyses its reaction by promoting ionisation of the substrate and subsequently using both the departed pyrophosphate group and cation-pi interactions from aromatic residues to stabilise and orientate carbocation intermediates. Initial substrate ionisation is promoted by three Mg2+ ions that stabilise the departing pyrophosphate leaving group. Recapture of the pyrophosphate at C3 to give (3R)-linalyl diphosphate allows rotation around the C2-C3 bond. Re-ionisation now facilitates SN' cyclisation to give the (4R)-alpha-terpinyl cation. The C8 carbocation is likely to be stabilised by cation-pi interactions involving Phe 578 and Trp 323. Further anti-Markovnikov cyclisation gives the 2-boronyl cation that is quenched by pyrophosphate to yield the final product.

Catalytic Residues Roles

UniProt PDB* (1n20)
Asp351, Asp355, Asp496, Thr500, Glu504 Asp351(302)A, Asp355(306)A, Asp496(447)A, Thr500(451)A, Glu504(455)A Coordinate Mg ions metal ligand
Trp323, Phe578 Trp323(274)A, Phe578(529)A Proposed to stabilise accumulation of positive charge on C8 carbocation using cation-pi interactions. van der waals interaction, polar/non-polar interaction, steric role, electrostatic stabiliser
Ile344, Val452 Ile344(295)A, Val452(403)A Play an important role in ensuring the correct reaction occurs by restricting the conformations that the intermediates can take. van der waals interaction, steric role
*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

heterolysis, overall reactant used, charge delocalisation, dephosphorylation, intermediate formation, bimolecular nucleophilic addition, intramolecular electrophilic addition, cyclisation, overall product formed

References

  1. Whittington DA et al. (2002), Proc Natl Acad Sci U S A, 99, 15375-15380. Nonlinear partial differential equations and applications: Bornyl diphosphate synthase: Structure and strategy for carbocation manipulation by a terpenoid cyclase. DOI:10.1073/pnas.232591099. PMID:12432096.
  2. O’Brien TE et al. (2018), ACS Catal, 8, 3322-3330. Predicting Productive Binding Modes for Substrates and Carbocation Intermediates in Terpene Synthases—Bornyl Diphosphate Synthase As a Representative Case. DOI:10.1021/acscatal.8b00342.
  3. Wise ML et al. (1998), J Biol Chem, 273, 14891-14899. Monoterpene synthases from common sage (Salvia officinalis). cDNA isolation, characterization, and functional expression of (+)-sabinene synthase, 1,8-cineole synthase, and (+)-bornyl diphosphate synthase. PMID:9614092.

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
Phe578(529)A steric role, van der waals interaction
Trp323(274)A steric role, van der waals interaction
Val452(403)A steric role, van der waals interaction
Ile344(295)A steric role, van der waals interaction
Asp351(302)A metal ligand
Asp355(306)A metal ligand
Asp496(447)A metal ligand
Thr500(451)A metal ligand
Glu504(455)A metal ligand

Chemical Components

heterolysis, overall reactant used, charge delocalisation, dephosphorylation, intermediate formation

Step 2. The diphosphate initiates nucleophilic attack at the C4 carbon, which causes double bond rearrangement and the formation of the linalyl diphosphate intermediate.

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Catalytic Residues Roles

Residue Roles
Phe578(529)A steric role, van der waals interaction
Trp323(274)A steric role, van der waals interaction
Val452(403)A steric role, van der waals interaction
Ile344(295)A steric role, van der waals interaction
Asp351(302)A metal ligand
Asp355(306)A metal ligand
Asp496(447)A metal ligand
Thr500(451)A metal ligand
Glu504(455)A metal ligand

Chemical Components

ingold: bimolecular nucleophilic addition, intermediate formation

Step 3. Isomerisation to linalyl diphosphate allows rotation of the newly formed C2-C3 bond to the cisoid conformer. The linalyl diphosphate 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.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Phe578(529)A steric role, van der waals interaction
Trp323(274)A steric role, van der waals interaction
Val452(403)A steric role, van der waals interaction
Ile344(295)A steric role, van der waals interaction
Asp351(302)A metal ligand
Asp355(306)A metal ligand
Asp496(447)A metal ligand
Thr500(451)A metal ligand
Glu504(455)A metal ligand

Chemical Components

heterolysis, overall reactant used, charge delocalisation, dephosphorylation, intermediate formation

Step 4. The terminal double bond adds to the newly formed carbocation in an intramolecular electrophilic addition resulting in a cyclic intermediate.

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Catalytic Residues Roles

Residue Roles
Phe578(529)A steric role, electrostatic stabiliser, polar/non-polar interaction
Trp323(274)A steric role, electrostatic stabiliser, polar/non-polar interaction
Val452(403)A steric role, van der waals interaction
Ile344(295)A steric role, van der waals interaction
Asp351(302)A metal ligand
Asp355(306)A metal ligand
Asp496(447)A metal ligand
Thr500(451)A metal ligand
Glu504(455)A metal ligand

Chemical Components

ingold: intramolecular electrophilic addition, cyclisation, intermediate formation

Step 5. The remaining double bond adds to the newly formed carbocation in an intramolecular electrophilic addition resulting in a bicyclic intermediate.

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Catalytic Residues Roles

Residue Roles
Phe578(529)A steric role, electrostatic stabiliser, polar/non-polar interaction
Trp323(274)A steric role, electrostatic stabiliser, polar/non-polar interaction
Val452(403)A steric role, van der waals interaction
Ile344(295)A steric role, van der waals interaction
Asp351(302)A metal ligand
Asp355(306)A metal ligand
Asp496(447)A metal ligand
Thr500(451)A metal ligand
Glu504(455)A metal ligand

Chemical Components

ingold: intramolecular electrophilic addition, cyclisation, intermediate formation

Step 6. The diphosphate initiates nucleophilic attack at the carbocation, forming the final product.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Phe578(529)A steric role, van der waals interaction
Trp323(274)A steric role, van der waals interaction
Val452(403)A steric role, van der waals interaction
Ile344(295)A steric role, van der waals interaction
Asp351(302)A metal ligand
Asp355(306)A metal ligand
Asp496(447)A metal ligand
Thr500(451)A metal ligand
Glu504(455)A metal ligand

Chemical Components

ingold: bimolecular nucleophilic addition, overall product formed
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 diphosphate initiates nucleophilic attack at the C4 carbon, which causes double bond rearrangement and the formation of the linalyl diphosphate intermediate.

Step 3. Isomerisation to linalyl diphosphate allows rotation of the newly formed C2-C3 bond to the cisoid conformer. The linalyl diphosphate 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 4. The terminal double bond adds to the newly formed carbocation in an intramolecular electrophilic addition resulting in a cyclic intermediate.

Step 5. The remaining double bond adds to the newly formed carbocation in an intramolecular electrophilic addition resulting in a bicyclic intermediate.

Step 6. The diphosphate initiates nucleophilic attack at the carbocation, forming the final product.

Products.

Contributors

Gemma L. Holliday, Steven Smith, James Willey