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Literature summary for 1.1.1.195 extracted from

  • Bhattacharyya, D.; Hazra, S.; Banerjee, A.; Datta, R.; Kumar, D.; Chakrabarti, S.; Chattopadhyay, S.
    Transcriptome-wide identification and characterization of CAD isoforms specific for podophyllotoxin biosynthesis from Podophyllum hexandrum (2016), Plant Mol. Biol., 92, 1-23.
    View publication on PubMed

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21(DE3) cells Sinopodophyllum hexandrum
gene PhCAD1, DNA and amino acid sequence determination and analysis, genotyping and phylogenetic analysis and transcriptome analysis, semi-quantitative RT-PCR enzyme expression analysis, overview. Recombinant expression of His-tagged enzyme Sinopodophyllum hexandrum
gene PhCAD2, DNA and amino acid sequence determination and analysis, genotyping and phylogenetic analysis and transcriptome analysis, semi-quantitative RT-PCR enzyme expression analysis, overview. Recombinant expression of His-tagged enzyme Sinopodophyllum hexandrum
gene PhCAD3, DNA and amino acid sequence determination and analysis, genotyping and phylogenetic analysis and transcriptome analysis, semi-quantitative RT-PCR enzyme expression analysis, overview. Recombinant expression of His-tagged enzyme Sinopodophyllum hexandrum
gene PhCAD4, DNA and amino acid sequence determination and analysis, genotyping and phylogenetic analysis and transcriptome analysis, semi-quantitative RT-PCR enzyme expression analysis, overview. Recombinant expression of His-tagged enzyme Sinopodophyllum hexandrum

Protein Variants

Protein Variants Comment Organism
additional information in PhCAD1 the active site residue G302 from the sinapyl alcohol dehydrogenase, SAD, is mutated to C302 and active site residue A293 is mutated to M293. Accumulation of ptox and lignin in PhCAD1-4 isoforms overexpressing transgenic lines, overview Sinopodophyllum hexandrum
additional information in PhCAD2 the active site residue G302 from the sinapyl alcohol dehydrogenase, SAD, is mutated to A300 and active site residue A293 is mutated to D290 Sinopodophyllum hexandrum
additional information in PhCAD3 the active site residue G302 from the sinapyl alcohol dehydrogenase, SAD, is conserved, but active site residue A293 is mutated to T290. Accumulation of ptox and lignin in PhCAD1-4 isoforms overexpressing transgenic lines, overview Sinopodophyllum hexandrum
additional information in PhCAD4 the active site residue G302 from the sinapyl alcohol dehydrogenase, SAD, is conserved, but active site residue A293 is mutated to M296 Sinopodophyllum hexandrum

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.0369
-
coniferaldehyde isoform CAD1, at pH 7.5 and 30°C Sinopodophyllum hexandrum
0.04792
-
sinapaldehyde isoform CAD1, at pH 7.5 and 30°C Sinopodophyllum hexandrum
0.0676
-
coniferaldehyde isoform CAD3, at pH 7.5 and 30°C Sinopodophyllum hexandrum
0.0797
-
coniferaldehyde isoform CAD4, at pH 7.5 and 30°C Sinopodophyllum hexandrum
0.1136
-
coniferaldehyde isoform CAD2, at pH 7.5 and 30°C Sinopodophyllum hexandrum
0.1378
-
sinapaldehyde isoform CAD2, at pH 7.5 and 30°C Sinopodophyllum hexandrum
0.1699
-
sinapaldehyde isoform CAD4, at pH 7.5 and 30°C Sinopodophyllum hexandrum
0.2082
-
sinapaldehyde isoform CAD3, at pH 7.5 and 30°C Sinopodophyllum hexandrum

Metals/Ions

Metals/Ions Comment Organism Structure
Zn2+ required Sinopodophyllum hexandrum

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
38700
-
-
Sinopodophyllum hexandrum
38800
-
-
Sinopodophyllum hexandrum
38900
-
-
Sinopodophyllum hexandrum
42100
-
-
Sinopodophyllum hexandrum

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
cinnamaldehyde + NADPH + H+ Sinopodophyllum hexandrum
-
cinnamyl alcohol + NADP+
-
?
coniferaldehyde + NADPH + H+ Sinopodophyllum hexandrum isoform CAD3 has a higher binding preference with coniferaldehyde over sinapaldehyde, followed by isoforms CAD4, CAD2, and CAD1, respectively coniferyl alcohol + NADP+
-
?
coniferyl aldehyde + NADPH + H+ Sinopodophyllum hexandrum
-
coniferyl alcohol + NADP+
-
r
sinapaldehyde + NADPH + H+ Sinopodophyllum hexandrum
-
sinapyl alcohol + NADP+
-
?
sinapyl aldehyde + NADPH + H+ Sinopodophyllum hexandrum
-
sinapyl alcohol + NADP+
-
r

Organism

Organism UniProt Comment Textmining
Sinopodophyllum hexandrum
-
-
-

Purification (Commentary)

Purification (Comment) Organism
Ni-NTA column chromatography Sinopodophyllum hexandrum

Source Tissue

Source Tissue Comment Organism Textmining
callus from leaves Sinopodophyllum hexandrum
-
cell suspension culture
-
Sinopodophyllum hexandrum
-
leaf
-
Sinopodophyllum hexandrum
-

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
cinnamaldehyde + NADPH + H+
-
Sinopodophyllum hexandrum cinnamyl alcohol + NADP+
-
?
coniferaldehyde + NADPH + H+ isoform CAD3 has a higher binding preference with coniferaldehyde over sinapaldehyde, followed by isoforms CAD4, CAD2, and CAD1, respectively Sinopodophyllum hexandrum coniferyl alcohol + NADP+
-
?
coniferyl aldehyde + NADPH + H+
-
Sinopodophyllum hexandrum coniferyl alcohol + NADP+
-
r
sinapaldehyde + NADPH + H+
-
Sinopodophyllum hexandrum sinapyl alcohol + NADP+
-
?
sinapyl aldehyde + NADPH + H+
-
Sinopodophyllum hexandrum sinapyl alcohol + NADP+
-
r

Subunits

Subunits Comment Organism
? x * 38700, isoform CAD4, calculated from amino acid sequence Sinopodophyllum hexandrum
? x * 38800, isoform CAD4, calculated from amino acid sequence Sinopodophyllum hexandrum
? x * 38900, isoform CAD4, calculated from amino acid sequence Sinopodophyllum hexandrum
? x * 42100, isoform CAD4, calculated from amino acid sequence Sinopodophyllum hexandrum

Synonyms

Synonyms Comment Organism
CAD
-
Sinopodophyllum hexandrum
CAD1 isoform Sinopodophyllum hexandrum
CAD2 isoform Sinopodophyllum hexandrum
CAD3 isoform Sinopodophyllum hexandrum
CAD4 isoform Sinopodophyllum hexandrum
cinnamyl alcohol dehydrogenase
-
Sinopodophyllum hexandrum
PhCAD1
-
Sinopodophyllum hexandrum
PhCAD2
-
Sinopodophyllum hexandrum
PhCAD3
-
Sinopodophyllum hexandrum
PhCAD4
-
Sinopodophyllum hexandrum

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
30
-
-
Sinopodophyllum hexandrum

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.4
-
sinapaldehyde isoform CAD3, at pH 7.5 and 30°C Sinopodophyllum hexandrum
2.76
-
sinapaldehyde isoform CAD4, at pH 7.5 and 30°C Sinopodophyllum hexandrum
3.4
-
sinapaldehyde isoform CAD2, at pH 7.5 and 30°C Sinopodophyllum hexandrum
7.9
-
coniferaldehyde isoform CAD1, at pH 7.5 and 30°C Sinopodophyllum hexandrum
8.3
-
sinapaldehyde isoform CAD1, at pH 7.5 and 30°C Sinopodophyllum hexandrum
9
-
coniferaldehyde isoform CAD2, at pH 7.5 and 30°C Sinopodophyllum hexandrum
10.8
-
coniferaldehyde isoform CAD4, at pH 7.5 and 30°C Sinopodophyllum hexandrum
12
-
coniferaldehyde isoform CAD3, at pH 7.5 and 30°C Sinopodophyllum hexandrum

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
-
Sinopodophyllum hexandrum

Cofactor

Cofactor Comment Organism Structure
NADP+
-
Sinopodophyllum hexandrum
NADPH
-
Sinopodophyllum hexandrum

pI Value

Organism Comment pI Value Maximum pI Value
Sinopodophyllum hexandrum isoform CAD4, calculated from amino acid sequence
-
5.9
Sinopodophyllum hexandrum isoform CAD3, calculated from amino acid sequence
-
6
Sinopodophyllum hexandrum isoform CAD2, calculated from amino acid sequence
-
6.2
Sinopodophyllum hexandrum isoform CAD1, calculated from amino acid sequence
-
6.5

Expression

Organism Comment Expression
Sinopodophyllum hexandrum the CAD genes are strongly upregulated by methyljasmonate, transcriptome analysis, overview up
Sinopodophyllum hexandrum the CAD genes are strongly upregulated by methyljasmonate, transcriptome analysis, overview. Accumulation of ptox and lignin in PhCAD1-4 isoforms overexpressing transgenic lines, overview up

General Information

General Information Comment Organism
additional information enzyme molecular docking analysis with substrates, active site structure, structure comparisons between isozymes, overview Sinopodophyllum hexandrum
physiological function CAD catalyzes the synthesis of coniferyl alcohol and sinapyl alcohol from coniferaldehyde (CAld) and sinapaldehyde respectively. Coniferyl alcohol can produce both lignin and lignan while sinapyl alcohol produces only lignin. Accumulation of ptox and lignin in PhCAD1-4 isoforms overexpressing transgenic lines, overview. Podophyllotoxin (ptox) is a therapeutically important lignan with economic importance derived from Podophyllum hexandrum and is used as a precursor for the synthesis of anticancer drugs etoposide, teniposide and etopophose Sinopodophyllum hexandrum

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
11
-
sinapaldehyde isoform CAD3, at pH 7.5 and 30°C Sinopodophyllum hexandrum
16
-
sinapaldehyde isoform CAD4, at pH 7.5 and 30°C Sinopodophyllum hexandrum
24
-
sinapaldehyde isoform CAD2, at pH 7.5 and 30°C Sinopodophyllum hexandrum
70
-
coniferaldehyde isoform CAD2, at pH 7.5 and 30°C Sinopodophyllum hexandrum
130
-
coniferaldehyde isoform CAD4, at pH 7.5 and 30°C Sinopodophyllum hexandrum
170
-
sinapaldehyde isoform CAD1, at pH 7.5 and 30°C Sinopodophyllum hexandrum
180
-
coniferaldehyde isoform CAD3, at pH 7.5 and 30°C Sinopodophyllum hexandrum
210
-
coniferaldehyde isoform CAD1, at pH 7.5 and 30°C Sinopodophyllum hexandrum