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Literature summary for 5.4.99.17 extracted from

  • Sato, T.; Kouda, M.; Hoshino, T.
    Site-directed mutagenesis experiments on the putative deprotonation site of squalene-hopene cyclase from Alicyclobacillus acidocaldarius (2004), Biosci. Biotechnol. Biochem., 68, 728-738.
    View publication on PubMed
Search for more literature for 5.4.99.17

Cloned(Commentary)

Cloned (Comment) Organism
expression in Escherichia coli BL21 Alicyclobacillus acidocaldarius

Protein Variants

Protein Variants Comment Organism
E45A mutation located around the "back waters" Alicyclobacillus acidocaldarius
E45A production of hop-22(29)-ene is less throughout the entire temperature range than that by the wild-type. Hop-21(22)ene is not produced Alicyclobacillus acidocaldarius
E93A mutation located around the "back waters" Alicyclobacillus acidocaldarius
E93A production of hop-22(29)-ene is less throughout the entire temperature range than that by the wild-type. Hop-21(22)ene is not produced Alicyclobacillus acidocaldarius
F434A mutation near the substrat channel Alicyclobacillus acidocaldarius
F434A production of hop-22(29)-ene is decreased, production of hopanol is markedly increased at lower temperatures Alicyclobacillus acidocaldarius
F437A mutation near the substrat channel Alicyclobacillus acidocaldarius
F437A production of hop-22(29)-ene is decreased, production of hopanol is markedly increased at lower temperatures Alicyclobacillus acidocaldarius
G262A the mutant produces hopanol as the main product instead of hop-22(29)-ene. The mutant also produces hop-21(22)ene Alicyclobacillus acidocaldarius
P263A mutation located between C29 of the hopanyl cation and the "front water" Alicyclobacillus acidocaldarius
P263A the mutant produces hopanol as the main product instead of hop-22(29)-ene. The mutant also produces hop-21(22)ene Alicyclobacillus acidocaldarius
P263G mutation located between C29 of the hopanyl cation and the "front water" Alicyclobacillus acidocaldarius
P263G the mutant produces hopanol as the main product instead of hop-22(29)-ene. The mutant also produces hop-21(22)ene Alicyclobacillus acidocaldarius
Q262A mutation located between C29 of the hopanyl cation and the "front water" Alicyclobacillus acidocaldarius
Q262G mutation located between C29 of the hopanyl cation and the "front water" Alicyclobacillus acidocaldarius
Q262G the mutant produces hopanol as the main product instead of hop-22(29)-ene. The mutant also produces hop-21(22)ene Alicyclobacillus acidocaldarius
R127Q mutation located around the "back waters" Alicyclobacillus acidocaldarius
R127Q production of hop-22(29)-ene is less throughout the entire temperature range than that by the wild-type. Hop-21(22)ene is not produced Alicyclobacillus acidocaldarius
T41A mutation located around the "back waters" Alicyclobacillus acidocaldarius
T41A production of hop-22(29)-ene is less throughout the entire temperature range than that by the wild-type. Hop-21(22)ene is not produced Alicyclobacillus acidocaldarius
W133A mutation located around the "back waters" Alicyclobacillus acidocaldarius
W133A production of hop-22(29)-ene is less throughout the entire temperature range than that by the wild-type. Hop-21(22)ene is not produced Alicyclobacillus acidocaldarius
Y267A mutation near the substrat channel Alicyclobacillus acidocaldarius
Y267A production of hop-22(29)-ene is decreased, production of hopanol is markedly increased at lower temperatures Alicyclobacillus acidocaldarius

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.0162
-
 squalene wild-type, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.0162
-
 squalene pH 6.0, 45°C, wild-type enzyme Alicyclobacillus acidocaldarius
0.0167
-
 squalene mutant T41A, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.0167
-
 squalene pH 6.0, 45°C, mutant T41A Alicyclobacillus acidocaldarius
0.0169
-
 squalene mutant W133A, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.0169
-
 squalene pH 6.0, 45°C, mutant W133A Alicyclobacillus acidocaldarius
0.0189
-
 squalene mutant E93A, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.0189
-
 squalene pH 6.0, 45°C, mutant E93A Alicyclobacillus acidocaldarius
0.0213
-
 squalene mutant E45A, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.0213
-
 squalene pH 6.0, 45°C, mutant E45A Alicyclobacillus acidocaldarius
0.0255
-
 squalene mutant R127Q, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.0255
-
 squalene pH 6.0, 45°C, mutant R127Q Alicyclobacillus acidocaldarius
0.0816
-
 squalene pH 6.0, 45°C, mutant F434A Alicyclobacillus acidocaldarius
0.0955
-
 squalene pH 6.0, 45°C, mutant F437A Alicyclobacillus acidocaldarius
0.102
-
 squalene mutant Q262A, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.102
-
 squalene pH 6.0, 45°C, mutant Q262A Alicyclobacillus acidocaldarius
0.156
-
 squalene mutant Q262G, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.156
-
 squalene pH 6.0, 45°C, mutant Q262G Alicyclobacillus acidocaldarius
0.185
-
 squalene mutant P263A, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.185
-
 squalene pH 6.0, 45°C, mutant P263A Alicyclobacillus acidocaldarius
0.197
-
 squalene mutantY267A, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.197
-
 squalene pH 6.0, 45°C, mutant Y267A Alicyclobacillus acidocaldarius
0.237
-
 squalene mutant P263G, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.237
-
 squalene pH 6.0, 45°C, mutant P263G Alicyclobacillus acidocaldarius
0.816
-
 squalene mutant F434A, 45°C, pH 6.0 Alicyclobacillus acidocaldarius
0.955
-
 squalene mutant F437A, 45°C, pH 6.0 Alicyclobacillus acidocaldarius

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
squalene Alicyclobacillus acidocaldarius
-
 hop-22(29)-ene
-
 ?

Organism

Organism UniProt Comment Textmining
Alicyclobacillus acidocaldarius
-
-
-

Purification (Commentary)

Purification (Comment) Organism
-
 Alicyclobacillus acidocaldarius

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
squalene
-
 Alicyclobacillus acidocaldarius hop-22(29)-ene
-
 ?
squalene two types of water molecules ("front water" and "back waters") are involved around the deprotonation site. The two residues of Gln262 and Pro263 probably work to keep away the isopropyl group of the hopanyl cation intermediate from the "front water molecule", that is, to place the "front water" in a favorable position, leading to the minimal production of by-products, i.e., hopanol and hop-21(22)-ene. The five residues of Thr41, Glu45, Glu93, Arg127 and Trp133, by which the hydrogen-bonded network incorporating the "back waters" is constructed, increase the polarization of the "front water" to facilitate proton elimination from the isopropyl moiety of the hopanyl cation, leading to the normal product, hop-22(29)-ene Alicyclobacillus acidocaldarius hop-22(29)-ene
-
 ?

Synonyms

Synonyms Comment Organism
SHC
-
 Alicyclobacillus acidocaldarius
squalene-hopene cyclase
-
 Alicyclobacillus acidocaldarius

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
35 50 mutant Q262G Alicyclobacillus acidocaldarius
45
-
 mutant F437A0 Alicyclobacillus acidocaldarius
45
-
 mutant F437A Alicyclobacillus acidocaldarius
45 55 mutant Q262A Alicyclobacillus acidocaldarius
50
-
 mutant E45A Alicyclobacillus acidocaldarius
50
-
 mutant P263A Alicyclobacillus acidocaldarius
50
-
 mutant P263G Alicyclobacillus acidocaldarius
50
-
 mutant R127Q Alicyclobacillus acidocaldarius
50
-
 mutant P263G, mutant P263A, mutant E45A, mutant R127Q Alicyclobacillus acidocaldarius
50 60 mutantY267A Alicyclobacillus acidocaldarius
50 60 mutant Y267A Alicyclobacillus acidocaldarius
55
-
 mutant E93A Alicyclobacillus acidocaldarius
55
-
 mutant W133A Alicyclobacillus acidocaldarius
55
-
 mutant E93A, mutant W133A Alicyclobacillus acidocaldarius
60
-
 wild-type Alicyclobacillus acidocaldarius
60
-
 mutant F434A Alicyclobacillus acidocaldarius
60
-
 mutant T41A Alicyclobacillus acidocaldarius
60
-
 wild-type, mutant T41A, mutant F434A Alicyclobacillus acidocaldarius

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.345
-
 squalene pH 6.0, 45°C, mutant E45A Alicyclobacillus acidocaldarius
0.353
-
 squalene pH 6.0, 45°C, mutant E93A Alicyclobacillus acidocaldarius
0.448
-
 squalene pH 6.0, 45°C, mutant R127Q Alicyclobacillus acidocaldarius
0.54
-
 squalene pH 6.0, 45°C, mutant F434A Alicyclobacillus acidocaldarius
0.62
-
 squalene pH 6.0, 45°C, mutant W133A Alicyclobacillus acidocaldarius
0.64
-
 squalene pH 6.0, 45°C, mutant Y267A Alicyclobacillus acidocaldarius
0.735
-
 squalene pH 6.0, 45°C, mutant T41A Alicyclobacillus acidocaldarius
0.86
-
 squalene pH 6.0, 45°C, wild-type enzyme Alicyclobacillus acidocaldarius
1.03
-
 squalene pH 6.0, 45°C, mutant F437A Alicyclobacillus acidocaldarius
1.06
-
 squalene pH 6.0, 45°C, mutant P263G Alicyclobacillus acidocaldarius
1.09
-
 squalene pH 6.0, 45°C, mutant P263A Alicyclobacillus acidocaldarius
1.21
-
 squalene pH 6.0, 45°C, mutant Q262A Alicyclobacillus acidocaldarius
1.37
-
 squalene pH 6.0, 45°C, mutant Q262G Alicyclobacillus acidocaldarius

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6
-
 assay at Alicyclobacillus acidocaldarius

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
3.2
-
 squalene pH 6.0, 45°C, mutant Y267A Alicyclobacillus acidocaldarius
4.5
-
 squalene pH 6.0, 45°C, mutant P263G Alicyclobacillus acidocaldarius
5.9
-
 squalene pH 6.0, 45°C, mutant P263A Alicyclobacillus acidocaldarius
6.6
-
 squalene pH 6.0, 45°C, mutant F434A Alicyclobacillus acidocaldarius
8.8
-
 squalene pH 6.0, 45°C, mutant Q262G Alicyclobacillus acidocaldarius
10.8
-
 squalene pH 6.0, 45°C, mutant F437A Alicyclobacillus acidocaldarius
11.9
-
 squalene pH 6.0, 45°C, mutant Q262A Alicyclobacillus acidocaldarius
16.1
-
 squalene pH 6.0, 45°C, mutant E45A Alicyclobacillus acidocaldarius
17.6
-
 squalene pH 6.0, 45°C, mutant R127Q Alicyclobacillus acidocaldarius
18.7
-
 squalene pH 6.0, 45°C, mutant E93A Alicyclobacillus acidocaldarius
36.7
-
 squalene pH 6.0, 45°C, mutant W133A Alicyclobacillus acidocaldarius
44
-
 squalene pH 6.0, 45°C, mutant T41A Alicyclobacillus acidocaldarius
53.09
-
 squalene pH 6.0, 45°C, wild-type enzyme Alicyclobacillus acidocaldarius