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Catalytic Site Atlas

CSA LITERATURE entry for 1n29

E.C. namephospholipase A2
SpeciesHomo sapiens (Human)
E.C. Number (IntEnz) 3.1.1.4
CSA Homologues of 1n29There are 229 Homologs
CSA Entries With UniProtID P14555
CSA Entries With EC Number 3.1.1.4
PDBe Entry 1n29
PDBSum Entry 1n29
MACiE Entry 1n29

Literature Report

IntroductionPLA2 catalyses the hydroylsis of the 2-acyl ester of 1,2-diacylphosphatides. Two forms of PLA2 are known: the small secretory PLA2s which are amphipathic molecules usually found associated with lipid membranes, and the larger cytosolic forms. This annotation is for the small secretory form. Secretory PLA2s form part of the neurotoxic component of many snake and bee venoms due to its ability to block acetylcholine release.
Secretory PLA2s have been divided into 3 main groups: I, II, and III. Class I (present in some snake venoms and in mammalian exocrine pancrease) and II (present in some snake venoms and also broadly distributed among a variety of mammalian cell types) are closely related to each other. The class III enzymes (including venom enzymes from the honeybee and the Gila monster) appear to form a separate divergent group though their active site and mechanism is similar. The key catalytic histidine and aspartate are in very similar orientations however the residues making up the rest of the hydrogen bond network are not conserved spatially.
MechansimA standard numbering system exists for class I and class II enzymes; in this system the catalytic residues are Gly 30, His 48, and Asp 99. In this entry however 1n29 numbering is used, in which the catalytic residues are Gly 29, His 47, and Asp 91.
The reaction involves attack by a water molecule on the ester carbonyl to give a tetrahedral intermediate which then collapses with loss of the alcoholate leaving group. A Ca2+ ion and the backbone NH of Gly 29 form an oxyanion hole to stabilise negative charge on the tetrahedral intermediate. There has been some debate however on exactly how the nucleophilic water molecule is activated, see references pubmed ID 11749391 or pubmed ID 12501175 for a discussion. In one proposed mechanism, His 47 deprotonates the attacking water molecule and later protonates the departing alcoholate leaving group. Asp 91 functions to modify the pKa of His 47 as in the serine proteases. A second proposed mechanism involves two water molecules at the active site. One of these (w6) deprotonates the other (w5) and is itself deprotonated by His 47, while w5 attacks the ester carbonyl. The departing alcoholate leaving group is protonated by w6 which is itself reprotonated by His 47. In this second proposed mechanism, which has been used for this annotation, the Ca2+ ion coordinates w5 and lowers its pKa as well as stabilising the tetrahedral intermediate.
The existence of a second Ca2+ ion in stabilising the tetrahedral intermediate has also been proposed, see references pubmed ID 8203286 and pubmed ID 1201175. This is suggested to interact with the amide oxygen of the peptide bond between residue 29 and Gly 30, hyperpolarising this peptide bond and so increasing oxyanion stabilisation by the NH group of Gly 30. However some sPLA2 enzymes do not seem to contain this second calcium ion or even a potential coordination site for it.
Reaction

Catalytic Sites for 1n29

Annotated By Reference To The Literature - Site 1 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
HisA4767macie:sideChainDeprotonates a water molecule (w6) that deprotonates the nucleophilic water w5. Reprotonates w6 as the alcoholate leaving group takes a proton from w6.
AspA91111macie:sideChainModifies the pKa of His 47.
GlyA2949macie:mainChainAmideBackbone NH forms part of the oxyanion hole that stabilises the tetrahedral intermediate resulting from nucleophilic attack by water on the substrate carbonyl.

Literature References

Notes:Clarification is needed on the role of catalytic water molecules in the active site and on the possible significance of the second calcium ion.
Berg OG
Interfacial enzymology: the secreted phospholipase A(2)-paradigm.
Chem Rev 2001 101 2613-2654
PubMed: 11749391
Edwards SH
The crystal structure of the H48Q active site mutant of human group IIA secreted phospholipase A2 at 1.5 A resolution provides an insight into the catalytic mechanism.
Biochemistry 2002 41 15468-15476
PubMed: 12501175
Scott DL
Structure and catalytic mechanism of secretory phospholipases A2.
Adv Protein Chem 1994 45 53-88
PubMed: 8154374
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