Cloned (Comment) | Organism |
---|---|
gene ChiW-SLHd, DNA and amino acid sequence determination and analysis, phylogenetic analysis, N-terminally His-tagged enzyme, the enzyme's N-terminus lacks the signal peptide and SLH domains , overexpression of active enzyme ChiW and mutants in Escherichia coli strain BL21 | Paenibacillus sp. |
Protein Variants | Comment | Organism |
---|---|---|
E1177Q | site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme | Paenibacillus sp. |
E691/E1177Q | site-directed mutagenesis, inactive mutant | Paenibacillus sp. |
E691Q | site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme | Paenibacillus sp. |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
2-mercaptoethanol | 15% inhibition | Paenibacillus sp. | |
4-nitrophenyl N,N'-diacetylchitobiose | substrate inhibition at the first catalytic domain with E691 | Paenibacillus sp. | |
4-O-N-acetyl-beta-D-glucosaminyl-N-acetyl-D-glucosamine | 21% inhibition | Paenibacillus sp. | |
Ca2+ | 12% inhibition at 5 mM | Paenibacillus sp. | |
Co2+ | 9% inhibition at 5 mM | Paenibacillus sp. | |
Cu2+ | 28% inhibition at 5 mM | Paenibacillus sp. | |
D-glucose | 18% inhibition | Paenibacillus sp. | |
DTT | 21% inhibition | Paenibacillus sp. | |
EDTA | 17% inhibition | Paenibacillus sp. | |
Fe3+ | 59% inhibition at 5 mM | Paenibacillus sp. | |
glutathione | GSH, 6% inhibition | Paenibacillus sp. | |
Mg2+ | 20% inhibition at 5 mM | Paenibacillus sp. | |
N-acetyl-D-glucosamine | 24% inhibition | Paenibacillus sp. |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | modified Michaelis-Menten kinetic with substrate inhibition | Paenibacillus sp. | |
0.14 | - |
4-nitrophenyl N,N'-diacetylchitobiose | recombinant enzyme, pH 5.5, 37°C | Paenibacillus sp. |
Localization | Comment | Organism | GeneOntology No. | Textmining |
---|---|---|---|---|
cell surface | - |
Paenibacillus sp. | 9986 | - |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
additional information | no effect by 5 mM Mn2+ | Paenibacillus sp. |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
130000 | - |
recombinant truncated His-tagged enzyme, gel filtration | Paenibacillus sp. |
150000 | - |
1 * 150000, about, chitinase ChiW, sequence calculation, 1 * 130000, recombinant truncated His-tagged enzyme, specifically cleaved between Asn282 and Ser283 during the purification step, the resulting two polypeptide fragments were non-covalently bound, SDS-PAGE | Paenibacillus sp. |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
chitin + H2O | Paenibacillus sp. | - |
? | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Paenibacillus sp. | K7ZLW6 | produces several different chitinases | - |
Purification (Comment) | Organism |
---|---|
recombinant soluble His-tagged enzyme ChiW, lacking the the signal peptide and SLH domains in the N-terminus, 1.6fold from Escherichia coli strain BL21 to homogeneity by nickel affinity and anion exchange chromatography | Paenibacillus sp. |
Specific Activity Minimum [µmol/min/mg] | Specific Activity Maximum [µmol/min/mg] | Comment | Organism |
---|---|---|---|
0.71 | - |
purified recombinant enzyme, pH 5.5, 37°C, substrate 4-nitrophenyl-N,N'-diacetyl-D-chitobiose | Paenibacillus sp. |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
4-nitrophenyl N,N'-diacetylchitobiose + H2O | synthetic substrate | Paenibacillus sp. | 4-nitrophenol + N,N'-diacetylchitobiose | - |
? | |
chitin + H2O | - |
Paenibacillus sp. | ? | - |
? | |
chitin + H2O | insoluble, powdered chitin and colloidal chitin, and chitin nanofiber | Paenibacillus sp. | N,N'-diacetylchitobiose + ? | final product | ? | |
ethylene glycol chitin + H2O | - |
Paenibacillus sp. | ? | - |
? | |
additional information | thin layer chromatography analysis of the reaction products, substrate specificity, overview. No activity with glycol chitosan | Paenibacillus sp. | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
monomer | 1 * 150000, about, chitinase ChiW, sequence calculation, 1 * 130000, recombinant truncated His-tagged enzyme, specifically cleaved between Asn282 and Ser283 during the purification step, the resulting two polypeptide fragments were non-covalently bound, SDS-PAGE | Paenibacillus sp. |
More | the enzyme contains an anchoring domains in the N-terminus and two catalytic domains at the C-terminal region, presence of a beta-stranded-rich structure in the N-terminus | Paenibacillus sp. |
Synonyms | Comment | Organism |
---|---|---|
ChiW | - |
Paenibacillus sp. |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
50 | - |
- |
Paenibacillus sp. |
Temperature Minimum [°C] | Temperature Maximum [°C] | Comment | Organism |
---|---|---|---|
20 | 70 | activity range, profile overview | Paenibacillus sp. |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
40 | - |
purified recombinant enzyme, stable up to | Paenibacillus sp. |
55 | - |
purified recombinant enzyme, 50% activity remaining | Paenibacillus sp. |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.99 | - |
4-nitrophenyl N,N'-diacetylchitobiose | recombinant enzyme, pH 5.5, 37°C | Paenibacillus sp. |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
5.5 | - |
assay at | Paenibacillus sp. |
pH Minimum | pH Maximum | Comment | Organism |
---|---|---|---|
4.5 | 8.5 | activity range, profile overview | Paenibacillus sp. |
Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
---|---|---|---|---|---|
0.089 | - |
4-nitrophenyl N,N'-diacetylchitobiose | recombinant enzyme, pH 5.5, 37°C | Paenibacillus sp. |
General Information | Comment | Organism |
---|---|---|
evolution | the enzyme is a typical glycoside hydrolase family 18 chitinase | Paenibacillus sp. |
additional information | Glu691 and Glu1177 in the two GH-18 domains are catalytic residues, two catalytic domains act synergistically to degrade chitin. The reaction mechanism of ChiW is suggested to be similar to the mechanism of GH-18 chitinases, in which one glutamate residue functions as a general acid/base catalyst and a GlcNAc C-2 acetamide group of the chitin substrate at subsite -1 carries out a nucleophilic attack on the GlcNAc C-1 carbon atom generating an oxazoline ion intermediate. Other important residues of the chitinase subfamily A enzymes for saccharide binding and catalytic reaction are also highly conserved in the ChiW GH-18 domains: Trp568 (Trp1055), Trp905 (Trp1396), Trp652 (Trp1138), and Trp772 (Trp1258) for saccharide binding, Tyr766 (Tyr1252), Asp687 (Asp1173), and Asp689 (Asp1175) for the catalytic reaction | Paenibacillus sp. |