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Catalytic Site Atlas

CSA LITERATURE entry for 1j00

E.C. namelysophospholipase
SpeciesEscherichia coli (Bacteria)
E.C. Number (IntEnz) 3.1.1.5
CSA Homologues of 1j00
CSA Entries With UniProtID P0ADA1
CSA Entries With EC Number 3.1.1.5
PDBe Entry 1j00
PDBSum Entry 1j00
MACiE Entry 1j00

Literature Report

IntroductionEscherichia coli thioesterase I (TAP) is a multifunctional enzyme possessing the activities of thioesterase, esterase, arylesterase, protease and lysophospholipase. This enzyme is also known as protease I and lysophospholipase L1, and is responsible for the degradation of thioester, amide and ester bonds. In particular, TAP catalyses the hydrolytic cleavage/deacylation of fatty acyl-CoA thioesters from fatty acyl-acyl carrier proteins, especially those with long acyl groups.
TAP has stereoselectivity for amino acid derivative substrates, and hence is useful for the kinetic resolution of racemic mixtures of industrial chemicals.
MechansimThe catalytic triad (Ser 10 - Asp 154 - His 157) and the oxyanion hole (Ser 10 - Gly 44 - Asn 73) are involved in the deacylation catalytic mechanism.
1. The His 157 imidazole ring becomes deprotonated, and its Ne2 lone pair becomes available for hydrogen bonding to the Ser 10 hydroxyl. 2. Asp 154 is positioned close to the entrance of the substrate tunnel and the O-delta-2 atom attracts the proton of N-delta-1-His 157. This draws the His backwards to provide a larger space during substrate binding. 3. Ser 10 OH acts as a nucleophile towards the carbonyl C of the ester substrate, forming an oxyanion intermediate. 4. The oxyanion is stabilised by the oxyanion hole, formed from Ser 10, Gly 44 and Asn 73. The amide protons of ser 10 and Gly 44 acts as proton donors, stabilising the tetrahedral adduct. 5. The thioester bond is cleaved, resulting in product formation and release.
Reaction

Catalytic Sites for 1j00

Annotated By Reference To The Literature - Site 1 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
GlyA4470macie:mainChainAmideGly 44 forms part of the oxyanion hole, to stabilise the oxyanion, and additionally plays a role in acyl-enzyme intermediate transformation. The Gly 44 amide proton is devoted to hydrogen bonding to the oxyanion, thereby acting as a proton donor. The minimally restrained movement of Gly 44, due to its small H side chain, is important in conformational fine-tuning during oxyanion formation.
SdpA1036macie:ptmSer 10 is the catalytic nucleophile. Hydrogen bonding to His 157 activates the serine towards nucleophilic attack of the carbonyl C in the substrate ester. Ser 10 is also involved in formation of the oxyanion hole, which stabilises the oxyanion intermediate. The Ser 10 amide proton is devoted to hydrogen bonding with the oxyanion.
AsnA7399macie:sideChainAsn 73 stabilises the oxyanion by contributing to the formation of the oxyanion hole. The asparagine residue offers its gamma-H for hydrogen bonding to the oxyanion. Asn 73 is also involved in the loop 75-80 switch-move motion, essential for accommodating longer acyl-chain-length substrates.
AspA154180macie:sideChainAsp 154 forms part of the catalytic triad. Asp 154 is positioned close to the entrance of the substrate tunnel and the O-delta-2 atom attracts the proton of N-delta-1-His 157. This draws the His backwards to provide a larger space during substrate binding.
HisA157183macie:sideChainThe His 157 imidazole ring becomes deprotonated, and its Ne2 lone pair becomes available for hydrogen bonding to the Ser 10 hydroxyl (side chain H-bonding). Thus, His 157 is responsible for the activation of the catalytic serine. Hydrogen bonding between the amide-N of His 157 and the O-gamma-1 of Asp 154 strengthens co-ordination of the catalytic triad and acts as a connector between Ser 10 and Asp 154. His 157 also participates in hydrogen bonding of the acyl-enzyme intermediate.

Annotated By Reference To The Literature - Site 2 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
GlyA4470macie:mainChainAmideGly 44 forms part of the oxyanion hole, to stabilise the oxyanion, and additionally plays a role in acyl-enzyme intermediate transformation. The Gly 44 amide proton is devoted to hydrogen bonding to the oxyanion, thereby acting as a proton donor. The minimally restrained movement of Gly 44, due to its small H side chain, is important in conformational fine-tuning during oxyanion formation.
SdpA1036macie:ptmSer 10 is the catalytic nucleophile. Hydrogen bonding to His 157 activates the serine towards nucleophilic attack of the carbonyl C in the substrate ester. Ser 10 is also involved in formation of the oxyanion hole, which stabilises the oxyanion intermediate. The Ser 10 amide proton is devoted to hydrogen bonding with the oxyanion.
AsnA7399macie:sideChainAsn 73 stabilises the oxyanion by contributing to the formation of the oxyanion hole. The asparagine residue offers its gamma-H for hydrogen bonding to the oxyanion. Asn 73 is also involved in the loop 75-80 switch-move motion, essential for accommodating longer acyl-chain-length substrates.
AspA154180macie:sideChainAsp 154 forms part of the catalytic triad. Asp 154 is positioned close to the entrance of the substrate tunnel and the O-delta-2 atom attracts the proton of N-delta-1-His 157. This draws the His backwards to provide a larger space during substrate binding.
HisA157183macie:sideChainThe His 157 imidazole ring becomes deprotonated, and its Ne2 lone pair becomes available for hydrogen bonding to the Ser 10 hydroxyl (side chain H-bonding). Thus, His 157 is responsible for the activation of the catalytic serine. Hydrogen bonding between the amide-N of His 157 and the O-gamma-1 of Asp 154 strengthens co-ordination of the catalytic triad and acts as a connector between Ser 10 and Asp 154. His 157 also participates in hydrogen bonding of the acyl-enzyme intermediate.

Literature References

Notes:
Lee LC
Functional role of catalytic triad and oxyanion hole-forming residues on enzyme activity of Escherichia coli thioesterase I/protease I/phospholipase L1.
Biochem J 2006 397 69-76
PubMed: 16515533
Lo YC
Crystal structure of Escherichia coli thioesterase I/protease I/lysophospholipase L1: consensus sequence blocks constitute the catalytic center of SGNH-hydrolases through a conserved hydrogen bond network.
J Mol Biol 2003 330 539-551
PubMed: 12842470
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