Phosphoinositide phospholipase C

 

Mammalian phospholipase C catalyses the hydrolysis of inositol lipid to inositol 1,4,5 trisphosphate and diacyl glycerol, both of which are important second messengers in Ca(II) signalling pathways. It possesses a Triose phosphate isomerase-like catalytic domain, indicating some homology with triosephosphate isomerase, but catalyses a different reaction by a mechanism more similar to the T1 RNAases.

 

Reference Protein and Structure

Sequence
P10688 UniProt (3.1.4.11) IPR028391 (Sequence Homologues) (PDB Homologues)
Biological species
Rattus norvegicus (Norway rat) Uniprot
PDB
1djx - PHOSPHOINOSITIDE-SPECIFIC PHOSPHOLIPASE C-DELTA1 FROM RAT COMPLEXED WITH INOSITOL-1,4,5-TRISPHOSPHATE (2.3 Å) PDBe PDBsum 1djx
Catalytic CATH Domains
3.20.20.190 CATHdb (see all for 1djx)
Cofactors
Calcium(2+) (1) Metal MACiE
Click To Show Structure

Enzyme Reaction (EC:3.1.4.11)

1-phosphatidyl-1D-myo-inositol 4,5-bisphosphate(5-)
CHEBI:58456ChEBI
+
water
CHEBI:15377ChEBI
→
1,2-diacyl-sn-glycerol
CHEBI:17815ChEBI
+
1D-myo-inositol 1,4,5-trisphosphate(6-)
CHEBI:203600ChEBI
+
hydron
CHEBI:15378ChEBI
Alternative enzyme names: 1-phosphatidyl-D-myo-inositol-4,5-bisphosphate inositoltrisphosphohydrolase, 1-phosphatidylinositol-4,5-bisphosphate phosphodiesterase, Monophosphatidylinositol phosphodiesterase, Phosphatidylinositol phospholipase C, Phosphoinositidase C, Triphosphoinositide phosphodiesterase, PI-PLC,

Enzyme Mechanism

Introduction

The OH group on C2 acts as a nucleophile, activated by the general base Glu 341, and forms a cyclic phosphate intermediate, with the release of DAG facilitated by the general acid His 356. The intermediate, stabilised by Ca2+ and His 311, is then hydrolysed by a water molecule activated by His 356 to give 1,4,5,inositol trisphosphate and complete the reaction cycle.

Catalytic Residues Roles

UniProt PDB* (1djx)
Glu390 Glu390(258)A Acts as general base to deprotonate the C2 OH of the inositol so that it can act as a nucleophile and attack the phosphate to form the cyclic intermediate, releasing DAG. hydrogen bond acceptor, hydrogen bond donor, metal ligand, proton acceptor, proton donor, increase acidity
His356 His356(224)A Activates water by deprotonation to allow it to act as a nucleophile and hydrolyse the cyclic phosphate intermediate. hydrogen bond acceptor, hydrogen bond donor, proton acceptor, proton donor
Glu341 Glu341(209)A Could also act as the general acid/base in this reaction. hydrogen bond acceptor, metal ligand, electrostatic stabiliser, increase acidity
His311 His311(179)A Forms hydrogen bonds to stabilise the cyclic intermediate structure which then collapses to release the products. hydrogen bond donor, electrostatic stabiliser
Asn312, Asp343 Asn312(180)A, Asp343(211)A Binds Ca(II) ion. metal ligand
*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

intramolecular nucleophilic substitution, overall reactant used, overall product formed, cyclisation, intermediate formation, proton transfer, bimolecular nucleophilic substitution, decyclisation, intermediate terminated, native state of enzyme regenerated

References

  1. Essen LO et al. (1997), Biochemistry, 36, 1704-1718. Structural Mapping of the Catalytic Mechanism for a Mammalian Phosphoinositide-Specific Phospholipase C†,‡. DOI:10.1021/bi962512p. PMID:9048554.
  2. Apiyo D et al. (2005), Biochemistry, 44, 9980-9989. X-ray Structure of the R69D Phosphatidylinositol-Specific Phospholipase C Enzyme:  Insight into the Role of Calcium and Surrounding Amino Acids in Active Site Geometry and Catalysis. DOI:10.1021/bi047896v. PMID:16042375.
  3. Heinz DW et al. (1998), J Mol Biol, 275, 635-650. Structural and mechanistic comparison of prokaryotic and eukaryotic phosphoinositide-specific phospholipases C. DOI:10.1006/jmbi.1997.1490. PMID:9466937.
  4. Ellis MV et al. (1998), J Biol Chem, 273, 11650-11659. Catalytic Domain of Phosphoinositide-specific Phospholipase C (PLC). MUTATIONAL ANALYSIS OF RESIDUES WITHIN THE ACTIVE SITE AND HYDROPHOBIC RIDGE OF PLCdelta 1. DOI:10.1074/jbc.273.19.11650. PMID:9565585.
  5. Essen LO et al. (1996), Nature, 380, 595-602. Crystal structure of a mammalian phosphoinositide-specific phospholipase Cδ. DOI:10.1038/380595a0. PMID:8602259.

Step 1. Glu390 deprotonates the hydroxyl group adjacent to the phosphate, which initiates a nucleophilic attack on the phosphorus in a substitution reaction that eliminates diacylglycerol which deprotonates His356.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Glu341(209)A hydrogen bond acceptor, increase acidity, electrostatic stabiliser
His356(224)A hydrogen bond donor
Glu390(258)A hydrogen bond acceptor, increase acidity
His311(179)A hydrogen bond donor, electrostatic stabiliser
Glu390(258)A metal ligand
Asp343(211)A metal ligand
Glu341(209)A metal ligand
Asn312(180)A metal ligand
Glu390(258)A proton acceptor
His356(224)A proton donor

Chemical Components

ingold: intramolecular nucleophilic substitution, overall reactant used, overall product formed, cyclisation, intermediate formation, proton transfer

Step 2. His356 deprotonates water, which initiates a nucleophilic attack upon the phosphorus in a substitution reaction. The initial attacking hydroxyl is regenerated and deprotonates Glu390.

Download: Image, Marvin File

Catalytic Residues Roles

Residue Roles
Glu341(209)A hydrogen bond acceptor, electrostatic stabiliser
His356(224)A hydrogen bond acceptor
Glu390(258)A hydrogen bond donor
His311(179)A hydrogen bond donor, electrostatic stabiliser
Glu390(258)A metal ligand
Asp343(211)A metal ligand
Glu341(209)A metal ligand
Asn312(180)A metal ligand
His356(224)A proton acceptor
Glu390(258)A proton donor

Chemical Components

ingold: bimolecular nucleophilic substitution, overall product formed, decyclisation, intermediate terminated, native state of enzyme regenerated, proton transfer
Download: Image, Marvin File

Step 1. Glu390 deprotonates the hydroxyl group adjacent to the phosphate, which initiates a nucleophilic attack on the phosphorus in a substitution reaction that eliminates diacylglycerol which deprotonates His356.

Step 2. His356 deprotonates water, which initiates a nucleophilic attack upon the phosphorus in a substitution reaction. The initial attacking hydroxyl is regenerated and deprotonates Glu390.

Products.

Contributors

Gemma L. Holliday, Gail J. Bartlett, Daniel E. Almonacid, Sophie T. Williams, Peter Sarkies, Katherine Ferris