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Catalytic Site Atlas

CSA LITERATURE entry for 2sqc

E.C. namesqualene---hopene cyclase
SpeciesAlicyclobacillus acidocaldarius (Bacteria)
E.C. Number (IntEnz) 5.4.99.17
CSA Homologues of 2sqcThere are 15 Homologs
CSA Entries With UniProtID P33247
CSA Entries With EC Number 5.4.99.17
PDBe Entry 2sqc
PDBSum Entry 2sqc
MACiE Entry M0254

Literature Report

IntroductionSqualene-hopene cyclase (SHC) from Alicyclobacillus acidocaldarius catalyses the cyclation of linear triterpene squalene. There are two possible products; hop-22(29)-ene or hopan-22-ol (aka diplopterol) which are formed in a roughly 90:10 ratio. SHC is a membrane protein with characteristics similar to those of prostaglandin-H2 synthase, the only other reported protein of this type. Enzymes like SHC are the targets for the development of anticholesteremic and antifungal drugs.
Mechansim1. The general acid (protonated) Asp 376 (coupled with His 451) protonates the substrate squalene at C3, forming a carbocation, and leaving the Asp:His diad as a stable salt bridge. The acidity of Asp 376 is increased by Tyr 495. 2. The positive charge on the Asp:His couple is stabilised by a negative charge on the Asp 374, Asp 377 couple. 3. There is a cyclization cascade, and the cation makes its way down the molecule. Atoms C4, C10, C8, C13, C17, C18 and C22 are stabilised by the residues Trp 312, Trp 489, Phe 365, Trp 169 and Phe 601 respectively (Phe 601 stabilising 3 positions.) 4. At C29, a water molecule acts as a base by accepting a proton from the substrate to form hopene. 5. This water molecule is polarised by other waters which are in turn polarised by a network of hydrogen bonds between Gln 262, Glu 45, Glu 93 and Arg 127. 6. Diplopterol is formed if the front water adds as hydroxyl to the last carbocation instead of accepting the proton. 7. Asp 376 is re-protonated by Tyr 495 via a water molecule, the proton originating from solvent water.
(the carbon numbering used above is for hopene)
Reaction

Catalytic Sites for 2sqc

Annotated By Reference To The Literature - Site 7 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
PheA605605macie:sideChainStabilises the carbocation in the C17 and C22 positions.
PheA601601macie:sideChainStabilises the carbocation in the C18 position.
GlnA262262macie:sideChainHelps to polarise the water molecule which acts as a general base at the end of the mechanism.
AspA374374macie:sideChainWorks with Asp 377 in stabilising the positive charge of the Asp 376, His 451 couple.
AspA377377macie:sideChainWorks with Asp 374 in stabilising the positively charged Asp 376, His 451 couple.
GluA4545macie:sideChainHelps to polarise the water molecule which acts as a general base at the end of the mechanism.
HisA451451macie:sideChainAssists Asp 376 in protonating the C3 atom of the substrate.
ArgA127127macie:sideChainHelps to polarise the water molecule which acts as a general base at the end of the mechanism.
TrpA312312macie:sideChainStabilises the carbocation in the C4 position.
TyrA495495macie:sideChainTyr 495 increases the acidity of Asp 376, making it a better general acid. It also re-protonates Asp 376 via a water molecule.
TrpA489489macie:sideChainStabilises the carbocation in the C10 position.
TrpA169169macie:sideChainStabilises the carbocation in the C13 position.
CysA376376macie:sideChainActs as a general acid (along with His 451) to protonate the C3 atom of the substrate.
GluA9393macie:sideChainHelps to polarise the water molecule which acts as a general base at the end of the mechanism.
PheA365365macie:sideChainStabilises the carbocation in the C8 position.

Annotated By Reference To The Literature - Site 8 (Perform Site Search)
ResidueChainNumberUniProtKB NumberFunctional PartFunctionTargetDescription
PheB605605macie:sideChainStabilises the carbocation in the C17 and C22 positions.
PheB601601macie:sideChainStabilises the carbocation in the C18 position.
GlnB262262macie:sideChainHelps to polarise the water molecule which acts as a general base at the end of the mechanism.
AspB374374macie:sideChainWorks with Asp 377 in stabilising the positive charge of the Asp 376, His 451 couple.
AspB377377macie:sideChainWorks with Asp 374 in stabilising the positively charged Asp 376, His 451 couple.
GluB4545macie:sideChainHelps to polarise the water molecule which acts as a general base at the end of the mechanism.
HisB451451macie:sideChainAssists Asp 376 in protonating the C3 atom of the substrate.
ArgB127127macie:sideChainHelps to polarise the water molecule which acts as a general base at the end of the mechanism.
TrpB312312macie:sideChainStabilises the carbocation in the C4 position.
TyrB495495macie:sideChainTyr 495 increases the acidity of Asp 376, making it a better general acid. It also re-protonates Asp 376 via a water molecule.
TrpB489489macie:sideChainStabilises the carbocation in the C10 position.
TrpB169169macie:sideChainStabilises the carbocation in the C13 position.
CysB376376macie:sideChainActs as a general acid (along with His 451) to protonate the C3 atom of the substrate.
GluB9393macie:sideChainHelps to polarise the water molecule which acts as a general base at the end of the mechanism.
PheB365365macie:sideChainStabilises the carbocation in the C8 position.

Literature References

Notes:The conserved residues Tyr 609 and Tyr 612 could also act to stabilise the carbocation at the earlier positions.
Wendt KU
Structure and function of a squalene cyclase.
Science 1997 277 1811-1815
PubMed: 9295270
Wendt KU
The structure of the membrane protein squalene-hopene cyclase at 2.0 A resolution.
J Mol Biol 1999 286 175-187
PubMed: 9931258
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