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Literature summary extracted from

  • Perugino, G.; Trincone, A.; Giordano, A.; van der Oost, J.; Kaper, T.; Rossi, M.; Moracci, M.
    Activity of hyperthermophilic glycosynthases is significantly enhanced at acidic pH (2003), Biochemistry, 42, 8484-8493.
    View publication on PubMed

Protein Variants

EC Number Protein Variants Comment Organism
3.2.1.21 N62Y/Q63K/L231F/V303A/A412V mutations observed in all the clones isolated Thermosphaera aggregans
3.2.1.21 N62Y/Q63K/L231F/V303A/E386G/A412V additional mutation in nucleophile residue E386, mutation completely abolishes activity. The addition of 2 M sodium formate as an external nucleophile leads to the recovery of 4.60% activity with accumulation of oligosaccharides. At pH 3.0 and low concentrations of sodium formate buffer, the hyperthermophilic glycosynthase shows kcat values similar to those of the wild-type and 17fold higher than those observed at the usual reactivation conditions in 2 M sodium formate at pH 6.5 Thermosphaera aggregans
3.2.1.B26 E386G mutation in nucleophile residue E387, mutation completely abolishes activity under statndard conditions. The addition of 2 M sodium formate as an external nucleophile leads to the recovery of 8.40% activity with accumulation of oligosaccharides. At pH 3.0 and low concentrations of sodium formate buffer, the hyperthermophilic glycosynthase shows kcat values similar to those of the wild-type and 17fold higher than those observed at the usual reactivation conditions in 2 M sodium formate at pH 6.5 Saccharolobus solfataricus
3.2.1.B28 E386G mutation in nucleophile residue E387, mutation completely abolishes activity under statndard conditions. The addition of 2 M sodium formate as an external nucleophile leads to the recovery of 8.40% activity with accumulation of oligosaccharides. At pH 3.0 and low concentrations of sodium formate buffer, the hyperthermophilic glycosynthase shows kcat values similar to those of the wild-type and 17fold higher than those observed at the usual reactivation conditions in 2 M sodium formate at pH 6.5 Pyrococcus furiosus

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.2.1.21 0.2
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C Thermosphaera aggregans
3.2.1.21 0.38
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C, presence of 2 M formate Thermosphaera aggregans
3.2.1.21 1.01
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 6.5, 65°C, presence of 2 M formate Thermosphaera aggregans
3.2.1.21 2.8
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 6.0, 65°C Thermosphaera aggregans
3.2.1.21 3.1
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 5.0, 65°C Thermosphaera aggregans
3.2.1.21 3.3
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 4.0, 65°C Thermosphaera aggregans
3.2.1.21 6.6
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.B26 0.5
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C, presence of 2 M formate Saccharolobus solfataricus
3.2.1.B26 1.01
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C Saccharolobus solfataricus
3.2.1.B26 1.1
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 6.0, 65°C Saccharolobus solfataricus
3.2.1.B26 1.17
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 6.5, 65°C, presence of 2 M formate Saccharolobus solfataricus
3.2.1.B26 2.2
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 5.0, 65°C Saccharolobus solfataricus
3.2.1.B26 4.9
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 4.0, 65°C Saccharolobus solfataricus
3.2.1.B26 16.4
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B28 0.22
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C, presence of 2 M formate Pyrococcus furiosus
3.2.1.B28 0.28
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C Pyrococcus furiosus
3.2.1.B28 1.3
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 5.0, 65°C Pyrococcus furiosus
3.2.1.B28 1.49
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 6.5, 65°C, presence of 2 M formate Pyrococcus furiosus
3.2.1.B28 2.1
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 4.0, 65°C Pyrococcus furiosus
3.2.1.B28 4.3
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 3.0, 65°C Pyrococcus furiosus

Organism

EC Number Organism UniProt Comment Textmining
3.2.1.21 Thermosphaera aggregans Q9YGA8
-
-
3.2.1.B26 Saccharolobus solfataricus
-
-
-
3.2.1.B28 Pyrococcus furiosus Q51723
-
-

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
3.2.1.21 0.39
-
 mutant N62Y/Q63K/L231F/V303A/E386G/A412V, substrate cellobiose, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 6.3
-
 mutant N62Y/Q63K/L231F/V303A/E386G/A412V, substrate 2-nitrophenyl beta-D-galactopyranoside, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 6.4
-
 mutant N62Y/Q63K/L231F/V303A/E386G/A412V, substrate 2-nitrophenyl beta-D-xylopyranoside, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 10.4
-
 wild-type, substrate 2-nitrophenyl beta-D-xylopyranoside, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 12
-
 mutant N62Y/Q63K/L231F/V303A/E386G/A412V, substrate 4-nitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 33.8
-
 wild-type, substrate cellobiose, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 61.7
-
 wild-type, substrate 4-nitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 72.2
-
 wild-type, substrate 2-nitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 75
-
 wild-type, substrate 2,4-dinitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 129.8
-
 mutant N62Y/Q63K/L231F/V303A/E386G/A412V, substrate 2-nitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 140.9
-
 wild-type, substrate 2-nitrophenyl beta-D-galactopyranoside, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 207.1
-
 mutant N62Y/Q63K/L231F/V303A/E386G/A412V, substrate 2,4-dinitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.B26 1.2
-
 mutant E387G, substrate cellobiose, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 2.5
-
 mutant E387G, substrate 2-nitrophenyl beta-D-xylopyranoside, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 2.7
-
 mutant E387G, substrate 2-nitrophenyl beta-D-galactopyranoside, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 15
-
 mutant E387G, substrate 4-nitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 19.1
-
 wild-type, substrate 2-nitrophenyl beta-D-xylopyranoside, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 61.9
-
 wild-type, substrate cellobiose, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 63.3
-
 mutant E387G, substrate 2-nitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 127
-
 mutant E387G, substrate 2,4-dinitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 150
-
 wild-type, substrate 4-nitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 196
-
 wild-type, substrate 2,4-dinitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 219
-
 wild-type, substrate 2-nitrophenyl beta-D-galactopyranoside, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 253
-
 wild-type, substrate 2-nitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B28 0.08
-
 mutant E372A, substrate 2-nitrophenyl beta-D-galactopyranoside, pH 3.0, 65°C Pyrococcus furiosus
3.2.1.B28 0.28
-
 mutant E372A, substrate 2-nitrophenyl beta-D-xylopyranoside, pH 3.0, 65°C Pyrococcus furiosus
3.2.1.B28 4.7
-
 mutant E372A, substrate 2-nitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Pyrococcus furiosus
3.2.1.B28 8.3
-
 mutant E372A, substrate 2,4-dinitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Pyrococcus furiosus
3.2.1.B28 66
-
 wild-type, substrate cellobiose, pH 3.0, 65°C Pyrococcus furiosus
3.2.1.B28 75
-
 wild-type, substrate 2-nitrophenyl beta-D-xylopyranoside, pH 3.0, 65°C Pyrococcus furiosus
3.2.1.B28 291
-
 wild-type, substrate 4-nitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Pyrococcus furiosus
3.2.1.B28 320
-
 wild-type, substrate 2,4-dinitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Pyrococcus furiosus
3.2.1.B28 534
-
 wild-type, substrate 2-nitrophenyl beta-D-glucopyranoside, pH 3.0, 65°C Pyrococcus furiosus
3.2.1.B28 1049
-
 wild-type, substrate 2-nitrophenyl beta-D-galactopyranoside, pH 3.0, 65°C Pyrococcus furiosus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.2.1.21 2,4-dinitrophenyl beta-D-glucopyranoside + H2O
-
 Thermosphaera aggregans 2,4-dinitrophenol + D-glucopyranose
-
 ?
3.2.1.21 2-nitrophenyl beta-D-galactopyranoside + H2O
-
 Thermosphaera aggregans 2-nitrophenol + D-galactopyranose
-
 ?
3.2.1.21 2-nitrophenyl beta-D-glucopyranoside + H2O
-
 Thermosphaera aggregans 2-nitrophenol + D-glucopyranose
-
 ?
3.2.1.21 2-nitrophenyl beta-D-xylopyranoside + H2O
-
 Thermosphaera aggregans 2-nitrophenol + D-xylopyranose
-
 ?
3.2.1.21 4-nitrophenyl beta-D-glucopyranoside + H2O
-
 Thermosphaera aggregans 4-nitrophenol + D-glucopyranose
-
 ?
3.2.1.21 cellobiose + H2O
-
 Thermosphaera aggregans 2 D-glucose
-
 ?
3.2.1.B26 2,4-dinitrophenyl beta-D-glucopyranoside + H2O
-
 Saccharolobus solfataricus 2,4-dinitrophenol + D-glucopyranose
-
 ?
3.2.1.B26 2-nitrophenyl beta-D-galactopyranoside + H2O
-
 Saccharolobus solfataricus 2-nitrophenol + D-galactopyranose
-
 ?
3.2.1.B26 2-nitrophenyl beta-D-glucopyranoside + H2O
-
 Saccharolobus solfataricus 2-nitrophenol + D-glucopyranose
-
 ?
3.2.1.B26 2-nitrophenyl beta-D-xylopyranoside + H2O
-
 Saccharolobus solfataricus 2-nitrophenol + D-xylopyranose
-
 ?
3.2.1.B26 4-nitrophenyl beta-D-glucopyranoside + H2O
-
 Saccharolobus solfataricus 4-nitrophenol + D-glucopyranose
-
 ?
3.2.1.B26 cellobiose + H2O
-
 Saccharolobus solfataricus 2 D-glucose
-
 ?
3.2.1.B28 2,4-dinitrophenyl beta-D-glucopyranoside + H2O
-
 Pyrococcus furiosus 2,4-dinitrophenol + D-glucopyranose
-
 ?
3.2.1.B28 2-nitrophenyl beta-D-galactopyranoside + H2O
-
 Pyrococcus furiosus 2-nitrophenol + D-galactopyranose
-
 ?
3.2.1.B28 2-nitrophenyl beta-D-glucopyranoside + H2O
-
 Pyrococcus furiosus 2-nitrophenol + D-glucopyranose
-
 ?
3.2.1.B28 2-nitrophenyl beta-D-xylopyranoside + H2O
-
 Pyrococcus furiosus 2-nitrophenol + D-xylopyranose
-
 ?
3.2.1.B28 4-nitrophenyl beta-D-glucopyranoside + H2O
-
 Pyrococcus furiosus 4-nitrophenol + D-glucopyranose
-
 ?
3.2.1.B28 cellobiose + H2O
-
 Pyrococcus furiosus 2 D-glucose
-
 ?

Synonyms

EC Number Synonyms Comment Organism
3.2.1.B26 beta-Gly
-
 Saccharolobus solfataricus
3.2.1.B28 CelB
-
 Pyrococcus furiosus

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.2.1.21 11.6
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 6.0, 65°C Thermosphaera aggregans
3.2.1.21 53.9
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 5.0, 65°C Thermosphaera aggregans
3.2.1.21 72.9
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 6.5, 65°C, presence of 2 M formate Thermosphaera aggregans
3.2.1.21 305
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 4.0, 65°C Thermosphaera aggregans
3.2.1.21 309
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C Thermosphaera aggregans
3.2.1.21 439.8
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C, presence of 2 M formate Thermosphaera aggregans
3.2.1.21 970
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.B26 20.7
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 6.0, 65°C Saccharolobus solfataricus
3.2.1.B26 52.9
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 5.0, 65°C Saccharolobus solfataricus
3.2.1.B26 72.9
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 6.5, 65°C, presence of 2 M formate Saccharolobus solfataricus
3.2.1.B26 309
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C Saccharolobus solfataricus
3.2.1.B26 321.9
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 4.0, 65°C Saccharolobus solfataricus
3.2.1.B26 439.8
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C, presence of 2 M formate Saccharolobus solfataricus
3.2.1.B26 901.4
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B28 0.45
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 5.0, 65°C Pyrococcus furiosus
3.2.1.B28 3.9
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 6.5, 65°C, presence of 2 M formate Pyrococcus furiosus
3.2.1.B28 6.7
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 4.0, 65°C Pyrococcus furiosus
3.2.1.B28 47.6
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 3.0, 65°C Pyrococcus furiosus
3.2.1.B28 1189
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C, presence of 2 M formate Pyrococcus furiosus
3.2.1.B28 1797
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C Pyrococcus furiosus

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
3.2.1.21 4.2
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 6.0, 65°C Thermosphaera aggregans
3.2.1.21 17.2
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 5.0, 65°C Thermosphaera aggregans
3.2.1.21 72.9
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 6.5, 65°C, presence of 2 M formate Thermosphaera aggregans
3.2.1.21 91
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 4.0, 65°C Thermosphaera aggregans
3.2.1.21 147.4
-
 2-nitrophenyl beta-D-glucopyranoside mutant N62Y/Q63K/L231F/V303A/E386G/A412V, pH 3.0, 65°C Thermosphaera aggregans
3.2.1.21 1164
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C, presence of 2 M formate Thermosphaera aggregans
3.2.1.21 1575
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C Thermosphaera aggregans
3.2.1.B26 18.5
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 6.0, 65°C Saccharolobus solfataricus
3.2.1.B26 23.8
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 5.0, 65°C Saccharolobus solfataricus
3.2.1.B26 45
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 6.5, 65°C, presence of 2 M formate Saccharolobus solfataricus
3.2.1.B26 55
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 3.0, 65°C Saccharolobus solfataricus
3.2.1.B26 65.9
-
 2-nitrophenyl beta-D-glucopyranoside mutant E387G, pH 4.0, 65°C Saccharolobus solfataricus
3.2.1.B26 533
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C Saccharolobus solfataricus
3.2.1.B26 850
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C, presence of 2 M formate Saccharolobus solfataricus
3.2.1.B28 0.3
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 5.0, 65°C Pyrococcus furiosus
3.2.1.B28 2.6
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 6.5, 65°C, presence of 2 M formate Pyrococcus furiosus
3.2.1.B28 3.1
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 4.0, 65°C Pyrococcus furiosus
3.2.1.B28 10.9
-
 2-nitrophenyl beta-D-glucopyranoside mutant E372A, pH 3.0, 65°C Pyrococcus furiosus
3.2.1.B28 5350
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C, presence of 2 M formate Pyrococcus furiosus
3.2.1.B28 6480
-
 2-nitrophenyl beta-D-glucopyranoside wild-type, pH 6.5, 65°C Pyrococcus furiosus