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Literature summary for 5.4.99.17 extracted from

  • Siedenburg, G.; Jendrossek, D.
    Squalene-hopene cyclases (2011), Appl. Environ. Microbiol., 77, 3905-3915.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
DNA and amino acid sequence determination and analysis, sequence comparison, expression in Escherichia coli strain DH5alpha Bradyrhizobium japonicum
DNA and amino acid sequence determination and analysis, sequence comparison, expression in Escherichia coli strain DH5alpha Rhodopseudomonas palustris
DNA and amino acid sequence determination and analysis, sequence comparison, expression in Escherichia coli strain DH5alpha Streptomyces peucetius
DNA and amino acid sequence determination and analysis, sequence comparison, expression in Escherichia coli strain DH5alpha Zymomonas mobilis
DNA and amino acid sequence determination and analysis, sequence comparison, expression in Escherichia coli strain DH5alpha Methylococcus capsulatus
DNA and amino acid sequence determination and analysis, sequence comparison, expression in Escherichia coli strain DH5alpha Tetrahymena thermophila
DNA and amino acid sequence determination and analysis, sequence comparison, expression in Escherichia coli strain DH5alpha Alicyclobacillus acidocaldarius
gene shc, DNA and amino acid sequence determination, expression in Escherichia coli Alicyclobacillus acidocaldarius

Protein Variants

Protein Variants Comment Organism
C435S/D374I/D374V/H451F inactive mutant Alicyclobacillus acidocaldarius
C435S/D374I/D374V/H451F site-directed mutagenesis, inactive mutant Alicyclobacillus acidocaldarius
D376E inactive mutant Alicyclobacillus acidocaldarius
D376E site-directed mutagenesis, inactive mutant Alicyclobacillus acidocaldarius
D377C/D377N/Y612A the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
D377C/D377N/Y612A site-directed mutagenesis, the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
D377E/D376Q/D376R/D377R/E45K/W406V/W417A/D377C inactive mutant Alicyclobacillus acidocaldarius
D377E/D376Q/D376R/D377R/E45K/W406V/W417A/D377C site-directed mutagenesis, inactive mutant Alicyclobacillus acidocaldarius
F365A the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
F365A site-directed mutagenesis, the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
F601A the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
F601A site-directed mutagenesis, the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
F605A the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
F605A site-directed mutagenesis, the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
I261A the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
I261A site-directed mutagenesis, the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
additional information product patterns of mutant enzymes, detailed overview Alicyclobacillus acidocaldarius
Q262G/Q262A/P263G/P263A the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
Q262G/Q262A/P263G/P263A site-directed mutagenesis, the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
V380E inactive mutant Alicyclobacillus acidocaldarius
V380E site-directed mutagenesis, inactive mutant Alicyclobacillus acidocaldarius
V381A/D376C inactive mutant Alicyclobacillus acidocaldarius
V381A/D376C site-directed mutagenesis, inactive mutant Alicyclobacillus acidocaldarius
W169F/W169H/W489A/F605K the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
W169F/W169H/W489A/F605K site-directed mutagenesis, the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
Y420A the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
Y420A site-directed mutagenesis, the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
Y606A/W23V/W495V/W522V/W533A/W591L/W78S/E35Q/E197Q/D530N/T378A the mutant shows the same product pattern and activity as the wild-type Alicyclobacillus acidocaldarius
Y606A/W23V/W495V/W522V/W533A/W591L/W78S/E35Q/E197Q/D530N/T378A site-directed mutagenesis, the mutant shows the same product pattern and activity as the wild-type Alicyclobacillus acidocaldarius
Y609A/Y612A/L607K the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
Y609A/Y612A/L607K site-directed mutagenesis, the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
Y609F the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
Y609F site-directed mutagenesis, the mutant shows an altered product pattern compared to the wild-type enzyme, overview Alicyclobacillus acidocaldarius
Y609F site-directed mutagenesis, the mutant shows an altered product pattern compared to the wild-type enzyme, overview. The phenotype of Y609F mutein is contrarily described in two publications Alicyclobacillus acidocaldarius
Y612F/D376E/D376G/D377E/D377G/D377Q/E45A/E45D/F365W/T41A/E93A/R127Q/W133A/Y267A/F434A/F437A/W258L/D350N/D421N/D442N/H451R/D447N/D377N/D313N/E535Q/D374E the mutant shows the same product pattern as the wild-type with less enzyme activity Alicyclobacillus acidocaldarius
Y612F/D376E/D376G/D377E/D377G/D377Q/E45A/E45D/F365W/T41A/E93A/R127Q/W133A/Y267A/F434A/F437A/W258L/D350N/D421N/D442N/H451R/D447N/D377N/D313N/E535Q/D374E site-directed mutagenesis, the mutant shows the same product pattern as the wild-type with less enzyme activity Alicyclobacillus acidocaldarius

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.003 0.016 squalene pH 6.0, 60°C Alicyclobacillus acidocaldarius

Localization

Localization Comment Organism GeneOntology No. Textmining
plasma membrane enzyme SHC in vivo is a membrane-associated protein and can be solubilized from cell extracts by nonionic detergents, such as Triton X-100 or octylthioglucopyranoside. The enzyme is attached to the inner side of the cytoplasmic membrane by interactions of hydrophobic residues with the phospholipids. The membrane-binding part of the enzyme is a nonpolar region that is encircled by positive-charged amino acids enforcing the anchoring of the enzyme to the negatively charged surface of the phospholipid membrane Alicyclobacillus acidocaldarius 5886
-

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
71600
-
x * 71600, SDS-PAGE Alicyclobacillus acidocaldarius
71600
-
2 * 71600, about, sequence calculation Alicyclobacillus acidocaldarius
72000
-
x * 72000, SDS-PAGE Tetrahymena thermophila
72300
-
x * 72300, SDS-PAGE Rhodopseudomonas palustris
74100
-
x * 74100, SDS-PAGE Streptomyces peucetius
74100
-
x * 74100, SDS-PAGE Zymomonas mobilis
74100
-
x * 74100, SDS-PAGE Methylococcus capsulatus
76300
-
x * 76300, SDS-PAGE Bradyrhizobium japonicum
76300
-
x * 76300, about, sequence calculation Bradyrhizobium japonicum

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
additional information Alicyclobacillus acidocaldarius product pattern of alternative substrates, overview ?
-
?
squalene Bradyrhizobium japonicum
-
hop-22(29)-ene
-
?
squalene Rhodopseudomonas palustris
-
hop-22(29)-ene
-
?
squalene Streptomyces peucetius
-
hop-22(29)-ene
-
?
squalene Zymomonas mobilis
-
hop-22(29)-ene
-
?
squalene Methylococcus capsulatus
-
hop-22(29)-ene
-
?
squalene Tetrahymena thermophila
-
hop-22(29)-ene
-
?
squalene Alicyclobacillus acidocaldarius
-
hop-22(29)-ene
-
?

Organism

Organism UniProt Comment Textmining
Alicyclobacillus acidocaldarius
-
formerly Bacillus acidocaldarius
-
Alicyclobacillus acidocaldarius P33247 formerly Bacillus acidocaldarius, gene shc
-
Bradyrhizobium japonicum
-
-
-
Methylococcus capsulatus
-
-
-
no activity in Methylococcus capsulatus
-
-
-
Rhodopseudomonas palustris
-
-
-
Streptomyces peucetius
-
-
-
Tetrahymena thermophila
-
-
-
Zymomonas mobilis
-
-
-

Purification (Commentary)

Purification (Comment) Organism
native and/or recombinant enzyme, the enzyme in vivo is a membrane-associated protein and can be solubilized from cell extracts by nonionic detergents, such as Triton X-100 or octylthioglucopyranoside Alicyclobacillus acidocaldarius

Reaction

Reaction Comment Organism Reaction ID
squalene = hop-22(29)-ene catalytic mechanism, the initial reaction catalyzed is the protonation of the terminal double bond of squalene. The conserved DXDD motif of SHCs is essential for this protonation reaction, overview Bradyrhizobium japonicum
squalene = hop-22(29)-ene catalytic mechanism, the initial reaction catalyzed is the protonation of the terminal double bond of squalene. The conserved DXDD motif of SHCs is essential for this protonation reaction, overview Rhodopseudomonas palustris
squalene = hop-22(29)-ene catalytic mechanism, the initial reaction catalyzed is the protonation of the terminal double bond of squalene. The conserved DXDD motif of SHCs is essential for this protonation reaction, overview Streptomyces peucetius
squalene = hop-22(29)-ene catalytic mechanism, the initial reaction catalyzed is the protonation of the terminal double bond of squalene. The conserved DXDD motif of SHCs is essential for this protonation reaction, overview Zymomonas mobilis
squalene = hop-22(29)-ene catalytic mechanism, the initial reaction catalyzed is the protonation of the terminal double bond of squalene. The conserved DXDD motif of SHCs is essential for this protonation reaction, overview Methylococcus capsulatus
squalene = hop-22(29)-ene catalytic mechanism, the initial reaction catalyzed is the protonation of the terminal double bond of squalene. The conserved DXDD motif of SHCs is essential for this protonation reaction, overview Tetrahymena thermophila
squalene = hop-22(29)-ene catalytic mechanism, the initial reaction catalyzed is the protonation of the terminal double bond of squalene. The conserved DXDD motif of SHCs is essential for this protonation reaction. In Alicyclobacillus acidocaldarius SHC, Asp376 of this motif is hydrogen bonded to His451, and an additional hydrogen bond exists to an ordered water molecule, which connects D376 to the hydroxyl group of the Y495 side chain and thus further enhances its acidity. The carboxyl groups of Asp374 and Asp377 accommodate the positive charge of the D376-H451 pair prior to proton transfer. After proton transfer to the 2,3-double bond of squalene, the D376-H451 pair loses its charge, leaving the remaining negative charge on the D374-D377 pair for stabilization of the initial cationic intermediates (24, 101). Reprotonation of D376 occurs through a water molecule bound to Y495-OH, which can transfer protons from disordered water in the solvent-accessible upper cavity of SHC Alicyclobacillus acidocaldarius
squalene = hop-22(29)-ene overall mechanism of the polycyclization reaction of SHCs and structures of squalene cyclization products, overview Bradyrhizobium japonicum
squalene = hop-22(29)-ene overall mechanism of the polycyclization reaction of SHCs and structures of squalene cyclization products, overview Alicyclobacillus acidocaldarius

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
additional information substrate specificity, overview Alicyclobacillus acidocaldarius ?
-
?
additional information substrate specificity, detailed overview Streptomyces peucetius ?
-
?
additional information substrate specificity, detailed overview Zymomonas mobilis ?
-
?
additional information the enzyme also catalyzes 2,3-oxidosqualene cyclization, but no tetrahymanol formation. Substrate specificity, detailed overview Bradyrhizobium japonicum ?
-
?
additional information the enzyme also catalyzes 2,3-oxidosqualene cyclization, but no tetrahymanol formation. Substrate specificity, detailed overview Alicyclobacillus acidocaldarius ?
-
?
additional information the enzyme also catalyzes 2,3-oxidosqualene cyclization, substrate specificity, detailed overview Tetrahymena thermophila ?
-
?
additional information the enzyme does not catalyze 2,3-oxidosqualene cyclization nor tetrahymanol formation. Substrate specificity, detailed overview Methylococcus capsulatus ?
-
?
additional information the enzyme does not catalyze tetrahymanol formation. Substrate specificity, detailed overview Rhodopseudomonas palustris ?
-
?
additional information product pattern of alternative substrates, overview Alicyclobacillus acidocaldarius ?
-
?
squalene
-
Bradyrhizobium japonicum hop-22(29)-ene
-
?
squalene
-
Rhodopseudomonas palustris hop-22(29)-ene
-
?
squalene
-
Streptomyces peucetius hop-22(29)-ene
-
?
squalene
-
Zymomonas mobilis hop-22(29)-ene
-
?
squalene
-
Methylococcus capsulatus hop-22(29)-ene
-
?
squalene
-
Tetrahymena thermophila hop-22(29)-ene
-
?
squalene
-
Alicyclobacillus acidocaldarius hop-22(29)-ene
-
?

Subunits

Subunits Comment Organism
? x * 72000, SDS-PAGE Tetrahymena thermophila
? x * 71600, SDS-PAGE Alicyclobacillus acidocaldarius
? x * 72300, SDS-PAGE Rhodopseudomonas palustris
? x * 74100, SDS-PAGE Streptomyces peucetius
? x * 74100, SDS-PAGE Zymomonas mobilis
? x * 74100, SDS-PAGE Methylococcus capsulatus
? x * 76300, SDS-PAGE Bradyrhizobium japonicum
? x * 76300, about, sequence calculation Bradyrhizobium japonicum
homodimer 2 * 71600, about, sequence calculation Alicyclobacillus acidocaldarius
More dumbbell-shaped structure of chain A with a more structured beta-barrel structure in domain 1, active site structure, structure analysis, overview Alicyclobacillus acidocaldarius
More each subunit consists of alpha-helical domains that build up a dumbbell-shaped structure. The first domain consists of a regular (alpha/alpha)6 barrel structure, whereas the second domain shows an alpha-barrel structure in a less periodic manner Alicyclobacillus acidocaldarius

Synonyms

Synonyms Comment Organism
SHC
-
Bradyrhizobium japonicum
SHC
-
Rhodopseudomonas palustris
SHC
-
Streptomyces peucetius
SHC
-
Zymomonas mobilis
SHC
-
Methylococcus capsulatus
SHC
-
Tetrahymena thermophila
SHC
-
Alicyclobacillus acidocaldarius
squalene-hopene cyclase
-
Bradyrhizobium japonicum
squalene-hopene cyclase
-
Alicyclobacillus acidocaldarius

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
28
-
-
Bradyrhizobium japonicum
30
-
-
Rhodopseudomonas palustris
30
-
-
Zymomonas mobilis
30
-
-
Tetrahymena thermophila
35
-
-
Streptomyces peucetius
40
-
-
Methylococcus capsulatus
60
-
-
Alicyclobacillus acidocaldarius

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6
-
-
Zymomonas mobilis
6
-
-
Alicyclobacillus acidocaldarius
6.5
-
-
Bradyrhizobium japonicum
6.5
-
-
Rhodopseudomonas palustris
6.8
-
-
Streptomyces peucetius
6.8
-
-
Methylococcus capsulatus
7
-
-
Tetrahymena thermophila

pH Range

pH Minimum pH Maximum Comment Organism
5 8 activity range Rhodopseudomonas palustris

General Information

General Information Comment Organism
evolution structure-function relationships of squalene-hopene cyclases, the DXDD motif, which is typical for all squalene-hopene cyclases, overview Bradyrhizobium japonicum
evolution structure-function relationships of squalene-hopene cyclases, the DXDD motif, which is typical for all squalene-hopene cyclases, overview Rhodopseudomonas palustris
evolution structure-function relationships of squalene-hopene cyclases, the DXDD motif, which is typical for all squalene-hopene cyclases, overview Streptomyces peucetius
evolution structure-function relationships of squalene-hopene cyclases, the DXDD motif, which is typical for all squalene-hopene cyclases, overview Zymomonas mobilis
evolution structure-function relationships of squalene-hopene cyclases, the DXDD motif, which is typical for all squalene-hopene cyclases, overview Methylococcus capsulatus
evolution structure-function relationships of squalene-hopene cyclases, the DXDD motif, which is typical for all squalene-hopene cyclases, overview Tetrahymena thermophila
evolution structure-function relationships of squalene-hopene cyclases, the DXDD motif, which is typical for all squalene-hopene cyclases, overview Alicyclobacillus acidocaldarius
evolution enzyme distribution in the different taxa, overview Bradyrhizobium japonicum
evolution enzyme distribution in the different taxa, overview Alicyclobacillus acidocaldarius
metabolism the enzyme is involved in biosynthesis of hopanoids, members of a large group of cyclic triterpenoic compounds that have important functions in many prokaryotic and eukaryotic organisms Bradyrhizobium japonicum
metabolism the enzyme is involved in biosynthesis of hopanoids, members of a large group of cyclic triterpenoic compounds that have important functions in many prokaryotic and eukaryotic organisms Rhodopseudomonas palustris
metabolism the enzyme is involved in biosynthesis of hopanoids, members of a large group of cyclic triterpenoic compounds that have important functions in many prokaryotic and eukaryotic organisms Streptomyces peucetius
metabolism the enzyme is involved in biosynthesis of hopanoids, members of a large group of cyclic triterpenoic compounds that have important functions in many prokaryotic and eukaryotic organisms Zymomonas mobilis
metabolism the enzyme is involved in biosynthesis of hopanoids, members of a large group of cyclic triterpenoic compounds that have important functions in many prokaryotic and eukaryotic organisms Methylococcus capsulatus
metabolism the enzyme is involved in biosynthesis of hopanoids, members of a large group of cyclic triterpenoic compounds that have important functions in many prokaryotic and eukaryotic organisms Alicyclobacillus acidocaldarius
metabolism the nezym eis involved in biosynthesis of hopanoids, members of a large group of cyclic triterpenoic compounds that have important functions in many prokaryotic and eukaryotic organisms Tetrahymena thermophila
metabolism the enzyme converts squalene to hopanol, EC 4.2.1.129, and to hopene, EC 5.4.99.17, but not to tetrahymanol, EC 4.2.1.123, pathway overview Alicyclobacillus acidocaldarius
metabolism the enzyme converts squalene to tetrahymanol, EC 4.2.1.123, to hopene, EC 5.4.99.17, and to hopanol, EC 4.2.1.129, pathway overview Bradyrhizobium japonicum
additional information structure-function relationships of squalene-hopene cyclases, overview Bradyrhizobium japonicum
additional information structure-function relationships of squalene-hopene cyclases, overview Rhodopseudomonas palustris
additional information structure-function relationships of squalene-hopene cyclases, overview Streptomyces peucetius
additional information structure-function relationships of squalene-hopene cyclases, overview Zymomonas mobilis
additional information structure-function relationships of squalene-hopene cyclases, overview Methylococcus capsulatus
additional information structure-function relationships of squalene-hopene cyclases, overview Tetrahymena thermophila
additional information structure-function relationships of squalene-hopene cyclases, overview. A large central cavity represents the catalytic site in Alicyclobacillus acidocaldarius enzyme that takes up and orientates the squalene molecule. The channel and active-site cavity inside the protein are separated by a narrow constriction buildup of four amino acids, D376, F166, C435, and F434, that appear to block access to the active site Alicyclobacillus acidocaldarius
additional information structure-function relationships of SHCs, active site structure, overview Bradyrhizobium japonicum
additional information structure-function relationships of SHCs, active site structure, overview. A protruding part in the center of the nonpolar region contains a lipophilic channel and directs the substrate to the active-site cavity inside the protein. The channel and cavity are separated by a narrow constriction buildup of four amino acids, D376, F166, C435, and F434, that appear to block access to the active site. Residues C435 and F434 are part of a loop that seems to be flexible enough to permit passage of the substrate and the product Alicyclobacillus acidocaldarius