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Catalytic Site Atlas Version 2.2.12
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CSA entry for 1ptd
Original Entry
Title:
Hydrolase (phosphoric diester)
Compound:
Phosphatidylinositol-specific phospholipase c
Mutant:
No
UniProt/Swiss-Prot:
P14262-PLC_BACCE
EC Class:
4.6.1.13
Other CSA Entries:
Overview of all sites for 1ptd
Homologues of 1ptd
Entries for UniProt/Swiss-Prot: P14262
Entries for EC: 4.6.1.13
Other Databases:
PDB entry: 1ptd
PDBsum entry: 1ptd
UniProt/Swiss-Prot: P14262
IntEnz entry: 4.6.1.13
Literature Report:
Introduction:
This pdb contains the structure of Phosphatidylinositol-specific phospholipase C (PI-PLC) from Bacillus cereus. PI-PLC is a ubiquitous enzyme involved in a vast range of cellular signalling cascades. Prokaryotic PLCs act as virulence factors in some bacteria. PI-PLC catalyses the hydrolysis of the sn-3 phosphodiester bond of phosphatidylinositol (PI) producing diacylglycerol (DAG) and inositol 1-phosphate [I(1)P]. In the case of the bacterial enzyme the main product seems to be the cyclic intermediate, myo-inositol 1,2-cyclic phosphate, which is rapidly hydrolysed to I(1)P by the eukaryotic PI-PLCs.

PI-PLC from Bacillus cereus, and the nearly identical enzyme from B. thuringiensis have been used as model systems for the study of IP-PLCs as a whole. Bacterial PI-PLCs are metal-ion-independent. This is in contrast to mammalian PI-PLCs which are Ca2+ dependent. The R69D mutant of B. thuringiensis PI-PLC is calcium dependent and has been used to investigate the role of calcium in catalysis by PI-PLCs (PMID: 16042375).

Mechanism:
The first step of the is an intramolecular nucleophilic attack by the C2 hydroxyl of the substrate on the phosphorus producing IP and a myo-inositol 1,2-cyclic phosphate. This is catalysed by His32 and His82 acting as general base and general acid catalysts respectively.
The second step is reverse of the first step but with water acting as the nucleophile. This results in the hydrolysis of a phosphorus-oxygen bond in myo-inositol 1,2-cyclic phosphate to give D-myo-inositol 1-phosphate. However, for the bacterial enzyme, this step is slow and the main product is myo-inositol 1,2-cyclic phosphate.

Sites:

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Found by:
Literature reference 

ResidueChainNumberUniProt numberFunctional part FunctionTargetDescription
HISA 32 63Sidechain
Acid/baseSubstrate
In the first step His32 is the general base catalyst deprotonating the hydroxyl group on C2 of the substrate so that it may act as an intramolecular nucleophile.
Evidence from paper Evidence concerns Evidence type
PubMed ID 11583175 Current protein
PubMed ID 9335537 Current protein Residue is positioned appropriately (ligand position known)
PubMed ID 9335537 Current protein Mutagenesis of residue
PubMed ID 11583175 Current protein Kinetic studies
PubMed ID 9335537 Current protein Kinetic studies
PubMed ID 9335537 Current protein Conservation of residue

ResidueChainNumberUniProt numberFunctional part FunctionTargetDescription
ARGA 69 100Sidechain
ElectrostaticTransition state
The positive charge on Arg69 stabilises the negatively charged transition state.
Evidence from paper Evidence concerns Evidence type
PubMed ID 9335537 Current protein Residue is positioned appropriately (ligand position known)
PubMed ID 9335537 Current protein Mutagenesis of residue
PubMed ID 11583175 Current protein
PubMed ID 9335537 Current protein Kinetic studies
PubMed ID 11583175 Current protein Kinetic studies
PubMed ID 9335537 Current protein Conservation of residue

ResidueChainNumberUniProt numberFunctional part FunctionTargetDescription
HISA 82 113Sidechain
Acid/baseSubstrate
His82 is the general acid catalyst which protonates the sn-3 oxygen of the DAG leaving group.
Evidence from paper Evidence concerns Evidence type
PubMed ID 11583175 Current protein Kinetic studies
PubMed ID 9335537 Current protein Kinetic studies
PubMed ID 11583175 Current protein
PubMed ID 9335537 Current protein Residue is positioned appropriately (ligand position known)
PubMed ID 9335537 Current protein Conservation of residue
PubMed ID 9335537 Current protein Mutagenesis of residue

ResidueChainNumberUniProt numberFunctional part FunctionTargetDescription
ASPA 274 305Sidechain
ElectrostaticResidue
Asp274 forms a hydrogen bond to with the side chain of His32 and acts as an electrostatic stabiliser.
Evidence from paper Evidence concerns Evidence type
PubMed ID 11583175 Current protein
PubMed ID 9335537 Current protein Kinetic studies
PubMed ID 9335537 Current protein Residue is positioned appropriately (ligand position known)
PubMed ID 9335537 Current protein Mutagenesis of residue
PubMed ID 9335537 Current protein Conservation of residue
PubMed ID 11583175 Current protein Kinetic studies
Notes:


References:
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