Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary extracted from

  • Makhongela, H.S.; Glowacka, A.E.; Agarkar, V.B.; Sewell, B.T.; Weber, B.; Cameron, R.A.; Cowan, D.A.; Burton, S.G.
    A novel thermostable nitrilase superfamily amidase from Geobacillus pallidus showing acyl transfer activity (2007), Appl. Microbiol. Biotechnol., 75, 801-811.
    View publication on PubMed

Activating Compound

EC Number Activating Compound Comment Organism Structure
3.5.1.4 dithiothreitol activation to 150% original activity results from addition of dithiothreitol at millimolar concentration Aeribacillus pallidus

Application

EC Number Application Comment Organism
3.5.1.4 synthesis the amidase has potential for application under high temperature conditions as a biocatalyst for D-selective amide hydrolysis producing enantiomerically pure carboxylic acids and for production of novel amides by acyl transfer Aeribacillus pallidus

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.5.1.4 expressed in Escherichia coli Aeribacillus pallidus
3.5.1.4 expression in Escherichia coli Aeribacillus pallidus

General Stability

EC Number General Stability Organism
3.5.1.4 binding on Eupergit C beads at 4°C and on Amberlite-XAD57 results in low protein binding and low activity, but immobilization on Eupergit C beads at 25°C with cross-linking results in high protein binding yield and high immobilized specific activity (80% of non-immobilized activity) Aeribacillus pallidus

Inhibitors

EC Number Inhibitors Comment Organism Structure
3.5.1.4 2-mercaptoethanol complete inhibition at 5% (v/v) 2-mercaptoethanol Aeribacillus pallidus
3.5.1.4 acetamide substrate inhibition Aeribacillus pallidus
3.5.1.4 acetone 5% v/v, 86% inhibition; 86% decreased activity at 5% (v/v) Aeribacillus pallidus
3.5.1.4 acetonitrile 5% v/v, 93% inhibition; 93% decreased activity at 5% (v/v) Aeribacillus pallidus
3.5.1.4 Acrylamide substrate inhibition Aeribacillus pallidus
3.5.1.4 Co2+ 89-48% residual activity at 1 mM Aeribacillus pallidus
3.5.1.4 Cu2+ 89-48% residual activity at 1 mM Aeribacillus pallidus
3.5.1.4 Dimethyl sulfide 5% v/v, complete inhibition; complete inhibition at 5% (v/v) Aeribacillus pallidus
3.5.1.4 dimethyl sulfoxide 5% v/v, complete inhibition; complete inhibition at 5% (v/v) Aeribacillus pallidus
3.5.1.4 dithiothreitol 80% residual activity at 1 mM Aeribacillus pallidus
3.5.1.4 ethanol 58% decreased activity at 5% (v/v); 5% v/v, 58% inhibition Aeribacillus pallidus
3.5.1.4 Fe3+ 89-48% residual activity at 1 mM Aeribacillus pallidus
3.5.1.4 H2O2 5% v/v, complete inhibition; complete inhibition at 5% (v/v) H2O2 Aeribacillus pallidus
3.5.1.4 Isopropanol 5% v/v, 60% inhibition; 60% decreased activity at 5% (v/v) Aeribacillus pallidus
3.5.1.4 mercaptoethanol 5% v/v, complete inhibition Aeribacillus pallidus
3.5.1.4 methanol 50% decreased activity at 5% (v/v); 5% v/v, 50% inhibition Aeribacillus pallidus
3.5.1.4 n-butanol 48% decreased activity at 5% (v/v); 5% v/v, 48% inhibition Aeribacillus pallidus
3.5.1.4 Zn2+ 89-48% residual activity at 1 mM Aeribacillus pallidus

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.5.1.4 22.7
-
 acetamide pH 7.2 Aeribacillus pallidus
3.5.1.4 43.8
-
 Acrylamide pH 7.2 Aeribacillus pallidus

Molecular Weight [Da]

EC Number Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
3.5.1.4 38000
-
 6 * 38000, SDS-PAGE Aeribacillus pallidus
3.5.1.4 218000
-
 gel filtration Aeribacillus pallidus
3.5.1.4 228000
-
 SDS-PAGE Aeribacillus pallidus

Organism

EC Number Organism UniProt Comment Textmining
3.5.1.4 Aeribacillus pallidus
-
-
-
3.5.1.4 Aeribacillus pallidus RAPc8
-
-
-

Oxidation Stability

EC Number Oxidation Stability Organism
3.5.1.4 after incubation for 24 days at 20°C, buffered samples of amidase without reducing agents are completely inactivated. Samples incubated in the presence of 1 mM DTT show activities of approximately 80% of initial levels. At 20 mM DTT under otherwise identical conditions, acyl transfer activity is 200% higher than activities measured at time zero Aeribacillus pallidus

Purification (Commentary)

EC Number Purification (Comment) Organism
3.5.1.4
-
 Aeribacillus pallidus
3.5.1.4 heat precipitation and S-300 gel filtration Aeribacillus pallidus

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
3.5.1.4 37.5
-
 reombinant enzyme in crude extract, at pH 7 and 50°C Aeribacillus pallidus
3.5.1.4 221
-
-
 Aeribacillus pallidus
3.5.1.4 221
-
 recombinant enzyme after 6fold purification, at pH 7 and 50°C Aeribacillus pallidus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.5.1.4 acetamide + H2O
-
 Aeribacillus pallidus acetate + NH3
-
 ?
3.5.1.4 acetamide + H2O
-
 Aeribacillus pallidus RAPc8 acetate + NH3
-
 ?
3.5.1.4 acetamide + hydroxylamine
-
 Aeribacillus pallidus acetylhydroxamic acid + NH3
-
 ?
3.5.1.4 acrylamide + H2O 102% of the activity with acetamide Aeribacillus pallidus acrylate + NH3
-
 ?
3.5.1.4 acrylamide + hydroxylamine
-
 Aeribacillus pallidus acryloylhydroxamic acid + NH3
-
 ?
3.5.1.4 D-lactamide + H2O 17% of the activity with acetamide, D-selectivity towards lactamide Aeribacillus pallidus D-lactate + NH3
-
 ?
3.5.1.4 D-lactamide + hydroxylamine
-
 Aeribacillus pallidus ?
-
 ?
3.5.1.4 diacetamide + H2O 14% of the activity with acetamide Aeribacillus pallidus ?
-
 ?
3.5.1.4 fluoroacetamide + H2O 165% of the activity with acetamide Aeribacillus pallidus fluoroacetate + NH3
-
 ?
3.5.1.4 formamide + H2O 30% of the activity with acetamide Aeribacillus pallidus formate + NH3
-
 ?
3.5.1.4 formamide + H2O 30% of the activity with acetamide Aeribacillus pallidus RAPc8 formate + NH3
-
 ?
3.5.1.4 isobutyramide + H2O 16% of the activity with acetamide Aeribacillus pallidus isobutanoate + NH3
-
 ?
3.5.1.4 isobutyramide + hydroxylamine
-
 Aeribacillus pallidus isobutyrylhydroxamic acid + NH3
-
 ?
3.5.1.4 L-alaninamide + H2O 3% of the activity with acetamide Aeribacillus pallidus L-Ala + NH3
-
 ?
3.5.1.4 additional information acyl transfer reactions are demonstrated with acetamide, propionamide, isobutyramide, and acrylamide as substrates and hydroxylamine as the acyl acceptor. The highest reaction rate being with isobutyramide. Immobilization by entrapment in polyacrylamide gels, covalent binding on Eupergit C beads at 4°C and on Amberlite-XAD57 results in low protein binding and low activity, but immobilization on Eupergit C beads at 25°C with cross-linking resulted in high protein binding yield and high immobilized specific activity. No activity with: nicotinamide, isonicotinamide, L-asparagine, D-asparagine, DL-phenylalanine, L-prolinamide, hexanamide Aeribacillus pallidus ?
-
 ?
3.5.1.4 additional information not active on L-lactamide Aeribacillus pallidus ?
-
 ?
3.5.1.4 additional information acyl transfer reactions are demonstrated with acetamide, propionamide, isobutyramide, and acrylamide as substrates and hydroxylamine as the acyl acceptor. The highest reaction rate being with isobutyramide. Immobilization by entrapment in polyacrylamide gels, covalent binding on Eupergit C beads at 4°C and on Amberlite-XAD57 results in low protein binding and low activity, but immobilization on Eupergit C beads at 25°C with cross-linking resulted in high protein binding yield and high immobilized specific activity. No activity with: nicotinamide, isonicotinamide, L-asparagine, D-asparagine, DL-phenylalanine, L-prolinamide, hexanamide Aeribacillus pallidus RAPc8 ?
-
 ?
3.5.1.4 additional information not active on L-lactamide Aeribacillus pallidus RAPc8 ?
-
 ?
3.5.1.4 propionamide + H2O 67% of the activity with acetamide Aeribacillus pallidus propionate + NH3
-
 ?
3.5.1.4 propionamide + hydroxylamine
-
 Aeribacillus pallidus propionylhydroxamic acid + NH3
-
 ?
3.5.1.4 urea + H2O 1.5% of the activity with acetamide Aeribacillus pallidus ?
-
 ?
3.5.1.4 urea + H2O 1.5% of the activity with acetamide Aeribacillus pallidus RAPc8 ?
-
 ?

Subunits

EC Number Subunits Comment Organism
3.5.1.4 hexamer 6 * 38000, SDS-PAGE Aeribacillus pallidus
3.5.1.4 homohexamer 6 * 38000, SDS-PAGE Aeribacillus pallidus

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.5.1.4 50
-
-
 Aeribacillus pallidus
3.5.1.4 50
-
 soluble enzyme and enzyme immobilized on Eupergit C beads at 25°C with cross-linking Aeribacillus pallidus

Temperature Range [°C]

EC Number Temperature Minimum [°C] Temperature Maximum [°C] Comment Organism
3.5.1.4 40 80 40°C: about 70% of maximal activity, soluble enzyme. 40°C: about 30% of maximal activity, enzyme immobilized on Eupergit C beads. 80°C: about 50% of maximal activity, soluble enzyme. 80°C: about 55% of maximal activity, enzyme immobilized on Eupergit C beads Aeribacillus pallidus

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
3.5.1.4 50
-
 half-life: more than 5 h for soluble enzyme Aeribacillus pallidus
3.5.1.4 50 80 high thermal stability at 50°C and 60°C with half-lives greater than 5 h at both temperatures, the half-life values are 43 min and 10 min at 70°C and 80°C Aeribacillus pallidus
3.5.1.4 60
-
 half-life: more than 5 h for soluble enzyme Aeribacillus pallidus
3.5.1.4 70
-
 half-life: 43 min for soluble enzyme, 52 min for enzyme immobilized on Eupergit C beads at 25°C with cross-linking Aeribacillus pallidus
3.5.1.4 80
-
 half-life: 10 min for soluble enzyme, 30 min for enzyme immobilized on Eupergit C beads at 25°C with cross-linking Aeribacillus pallidus

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.5.1.4 38.7
-
 Acrylamide pH 7.2 Aeribacillus pallidus
3.5.1.4 2668
-
 acetamide pH 7.2 Aeribacillus pallidus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.5.1.4 6 8 enzyme immobilized on Eupergit C beads Aeribacillus pallidus
3.5.1.4 7
-
-
 Aeribacillus pallidus
3.5.1.4 7
-
 soluble enzyme Aeribacillus pallidus

pH Range

EC Number pH Minimum pH Maximum Comment Organism
3.5.1.4 4 9 pH 4.0: about 60% of maximal activity, soluble enzyme. pH 4.0: about 75% of maximal activity, enzyme immobilized on Eupergit C beads. and enzyme immobilized on Eupergit C beads. pH 9.0: about 40% of maximal activity, soluble enzyme. pH 9.0: about 90% of maximal activity, enzyme immobilized on Eupergit C beads Aeribacillus pallidus

Ki Value [mM]

EC Number Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
3.5.1.4 2.3
-
 Acrylamide pH 7.2 Aeribacillus pallidus
3.5.1.4 2.3
-
 Acrylamide at pH 7 and 50°C Aeribacillus pallidus
3.5.1.4 12.1
-
 acetamide pH 7.2 Aeribacillus pallidus
3.5.1.4 12.1
-
 acetamide at pH 7 and 50°C Aeribacillus pallidus