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Literature summary for 3.1.1.14 extracted from

  • Lee, G.C.; Chepyshko, H.; Chen, H.H.; Chu, C.C.; Chou, Y.F.; Akoh, C.C.; Shaw, J.F.
    Genes and biochemical characterization of three novel chlorophyllase isozymes from Brassica oleracea (2010), J. Agric. Food Chem., 58, 8651-8657.
    View publication on PubMed
Search for more literature for 3.1.1.14

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli Rosseta-Gami B (DE3) cells Brassica oleracea

Protein Variants

Protein Variants Comment Organism
D170A inactive Brassica oleracea
H226A the mutation causes almost complete loss of Chlase activity Brassica oleracea
H239A the mutation causes almost complete loss of Chlase activity Brassica oleracea
H247A the mutation causes almost complete loss of Chlase activity Brassica oleracea
H66A the mutation causes almost complete loss of Chlase activity Brassica oleracea
S141A inactive Brassica oleracea

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.0764
-
 pheophorbide a methyl ester wild type isoform CLH1, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.143
-
 pheophorbide a methyl ester wild type isoform CLH2, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.308
-
 chlorophyll a wild type isoform CLH2, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.311
-
 chlorophyll b wild type isoform CLH2, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.466
-
 chlorophyll a wild type isoform CLH1, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.652
-
 chlorophyll b wild type isoform CLH1, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea

Localization

Localization Comment Organism GeneOntology No. Textmining
chloroplast isoform CLH2 Brassica oleracea 9507
-
chloroplast isoforms CLH1 Brassica oleracea 9507
-
plasma membrane isoform CLH3 Brassica oleracea 5886
-

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
23500
-
 x * 23500, isoform CLH3, calculated from amino acid sequence Brassica oleracea
34700
-
 x * 34700, isoform CLH1, calculated from amino acid sequence Brassica oleracea
35300
-
 x * 35300, isoform CLH2, calculated from amino acid sequence Brassica oleracea
66200
-
 x * 66200, isoform CLH3 as MBP-fusion protein, SDS-PAGE Brassica oleracea
77400
-
 x * 77400, isoform CLH1 as MBP-fusion protein, SDS-PAGE Brassica oleracea
78000
-
 x * 78000, isoform CLH2 as MBP-fusion protein, SDS-PAGE Brassica oleracea

Organism

Organism UniProt Comment Textmining
Brassica oleracea Q8GTM2 isoform CLH3; cultivar Green king
-
Brassica oleracea Q8GTM3 isoform CLH2; cultivar Green king
-
Brassica oleracea Q8GTM4 isoform CLH1; cultivar Green king
-

Purification (Commentary)

Purification (Comment) Organism
amylose resin affinity column chromatography and DEAE Sepharose column chromatography Brassica oleracea

Source Tissue

Source Tissue Comment Organism Textmining
floret
-
 Brassica oleracea
-

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
0.074
-
 mutant enzyme H247A, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.149
-
 mutant enzyme H239A, in 100mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.372
-
 mutant enzyme H66A, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.52
-
 mutant enzyme H226A, in 100mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
7.43
-
 wild type isoform CLH2, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
chlorophyll a + H2O
-
 Brassica oleracea ?
-
 ?
chlorophyll b + H2O
-
 Brassica oleracea ?
-
 ?
additional information isoform CLH1 preferably hydrolyzes Mg-free chlorophyll Brassica oleracea ?
-
 ?
additional information isoform CLH2 hydrolyzes both chlorophyll and Mg-free chlorophyll at a similar level Brassica oleracea ?
-
 ?
additional information isoform CLH3 shows very low Chlase activity Brassica oleracea ?
-
 ?
pheophorbide a methyl ester + H2O
-
 Brassica oleracea ?
-
 ?

Subunits

Subunits Comment Organism
? x * 23500, isoform CLH3, calculated from amino acid sequence Brassica oleracea
? x * 34700, isoform CLH1, calculated from amino acid sequence Brassica oleracea
? x * 35300, isoform CLH2, calculated from amino acid sequence Brassica oleracea
? x * 66200, isoform CLH3 as MBP-fusion protein, SDS-PAGE Brassica oleracea
? x * 77400, isoform CLH1 as MBP-fusion protein, SDS-PAGE Brassica oleracea
? x * 78000, isoform CLH2 as MBP-fusion protein, SDS-PAGE Brassica oleracea

Synonyms

Synonyms Comment Organism
Chlase
-
 Brassica oleracea
chlorophyll chlorophyllidohydrolase
-
 Brassica oleracea
CLH1 isoform Brassica oleracea
CLH2 isoform Brassica oleracea
CLH3 isoform Brassica oleracea

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
30
-
 isoform CLH1 Brassica oleracea
60
-
 isoform CLH2 Brassica oleracea

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
60
-
 after incubation at 60°C for 10 min, isoform CLH1 loses about 50% of activity Brassica oleracea
60
-
 after incubation at 60°C for 10 min, isoform CLH2 loses about 50% of activity Brassica oleracea

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.0024
-
 chlorophyll a wild type isoform CLH2, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.0037
-
 chlorophyll b wild type isoform CLH2, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.0039
-
 pheophorbide a methyl ester wild type isoform CLH2, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.072
-
 chlorophyll a wild type isoform CLH1, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.072
-
 chlorophyll b wild type isoform CLH1, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.259
-
 pheophorbide a methyl ester wild type isoform CLH1, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7
-
 isoform CLH1 Brassica oleracea
8
-
 isoform CLH2 Brassica oleracea

pH Stability

pH Stability pH Stability Maximum Comment Organism
5 8 isoform CLH1 Brassica oleracea
8 10 isoform CLH2 shows about 80% of activity at pH 9.0 and is stable between pH 8.0 and 10.0 Brassica oleracea

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.0078
-
 chlorophyll a wild type isoform CLH2, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.0119
-
 chlorophyll b wild type isoform CLH2, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.0272
-
 pheophorbide a methyl ester wild type isoform CLH2, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.1105
-
 chlorophyll b wild type isoform CLH1, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
0.1551
-
 chlorophyll a wild type isoform CLH1, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea
3.397
-
 pheophorbide a methyl ester wild type isoform CLH1, in 100 mM sodium phosphate, pH 7.4, at 40°C Brassica oleracea