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Literature summary for 3.1.1.72 extracted from

  • Yoshida, S.; Mackie, R.I.; Cann, I.K.
    Biochemical and domain analyses of FSUAxe6B, a modular acetyl xylan esterase, identify a unique carbohydrate binding module in Fibrobacter succinogenes S85 (2010), J. Bacteriol., 192, 483-493.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
genes of wild-type and its truncational mutant proteins (TM1, TM2, TM3, and TM4) amplified and cloned into the pGEM-T vector and subcloned into a modified pET-28a expression vector. Truncated mutant protein TM5 amplified and annealed to the pET-46 Ek/LIC vector. Expressed in Escherichia coli BL21(DE3) CodonPlus RIPL-competent cells Fibrobacter succinogenes subsp. succinogenes S85

Protein Variants

Protein Variants Comment Organism
D270A displays catalytic activity against tetraacetyl-xylopyranoside Fibrobacter succinogenes subsp. succinogenes S85
D270N displays catalytic activity against tetraacetyl-xylopyranoside Fibrobacter succinogenes subsp. succinogenes S85
E194A displays some catalytic activity Fibrobacter succinogenes subsp. succinogenes S85
E194A/D270A activity is completely abolished Fibrobacter succinogenes subsp. succinogenes S85
E194N exhibits detectable catalytic activity Fibrobacter succinogenes subsp. succinogenes S85
H273Q activity is completely abolished Fibrobacter succinogenes subsp. succinogenes S85
additional information truncational mutant proteins TM1 (esterase domain plus carbohydrate-binding module), TM2 (esterase domain), TM3 (carbohydrate-binding module plus FPm-1 domain), TM4 (carbohydrate-binding module) and TM5 (FPm-1 domain). Truncation of FPm-1 leads to a dramatic reduction of binding activity for the TM1 mutant. TM1 and TM2 have no discernible affinity for insoluble oat spelt xylan, whereas TM3, TM4, and TM5 bind to this substrate Fibrobacter succinogenes subsp. succinogenes S85
S44G activity is completely abolished Fibrobacter succinogenes subsp. succinogenes S85

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.07
-
tetraacetyl-xylopyranoside mutant TM2, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
0.08
-
tetraacetyl-xylopyranoside wild-type, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
0.09
-
tetraacetyl-xylopyranoside mutant TM1, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
0.2
-
tetraacetyl-xylopyranoside mutants E194A, D270N and D270A, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
7
-
tetraacetyl-xylopyranoside mutant E194N, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85

Organism

Organism UniProt Comment Textmining
Fibrobacter succinogenes subsp. succinogenes S85 Q9F106
-
-

Purification (Commentary)

Purification (Comment) Organism
truncational mutant proteins purified to near homogeneity Fibrobacter succinogenes subsp. succinogenes S85

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
insoluble oat spelt xylan
-
Fibrobacter succinogenes subsp. succinogenes S85 ?
-
?
additional information wild-type protein does not show any binding activity to Avicel. The enzyme is a modular protein comprised of an esterase domain, a carbohydrate-binding module, and a region of unknown function (Fibrobacter succinogenes-specific paralogous module 1, FPm-1). The N-terminal half of Axe6B harbors the activity that cleaves side chain acetyl groups from xylan-like substrates, and the binding of insoluble xylan is determined to originate from FPm-1. Esterase activity is derived from a tetrad composed of Ser44, His273, Glu194, and Asp270, with both Glu194 and Asp270 functioning as helper acids, instead of a single carboxylate residue proposed to initiate catalysis Fibrobacter succinogenes subsp. succinogenes S85 ?
-
?
tetraacetyl-xylopyranoside + H2O
-
Fibrobacter succinogenes subsp. succinogenes S85 ? + acetate
-
?

Synonyms

Synonyms Comment Organism
acetyl xylan esterase
-
Fibrobacter succinogenes subsp. succinogenes S85
Axe6B
-
Fibrobacter succinogenes subsp. succinogenes S85

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
1.8
-
tetraacetyl-xylopyranoside mutant D270A, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
2
-
tetraacetyl-xylopyranoside mutant D270N, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
2.8
-
tetraacetyl-xylopyranoside mutant E194N, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
2.9
-
tetraacetyl-xylopyranoside mutant E194A, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
13
-
tetraacetyl-xylopyranoside mutant TM2, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
15
-
tetraacetyl-xylopyranoside wild-type and mutant TM1, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85

pI Value

Organism Comment pI Value Maximum pI Value
Fibrobacter succinogenes subsp. succinogenes S85 mutant TM4, calculated from sequence
-
4.6
Fibrobacter succinogenes subsp. succinogenes S85 mutant TM2, calculated from sequence
-
5.2
Fibrobacter succinogenes subsp. succinogenes S85 mutant TM5, calculated from sequence
-
10.1

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.4
-
tetraacetyl-xylopyranoside mutant E194N, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
9
-
tetraacetyl-xylopyranoside mutant D270A, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
10
-
tetraacetyl-xylopyranoside mutant D270N, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
14
-
tetraacetyl-xylopyranoside mutant E194A, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
170
-
tetraacetyl-xylopyranoside mutant TM1, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85
190
-
tetraacetyl-xylopyranoside wild-type and mutant TM2, at 37°C, in 100 mM Tris-HCl buffer, pH 7.4, and 3 mM MgCl2 Fibrobacter succinogenes subsp. succinogenes S85