Purification and properties of a 1,3-1,4-β-d -glucanase (lichenase, 1,3-1,4-β-d -glucan 4-glucanohydrolase, EC 3.2.1.73) from Bacteroides succinogenes cloned in Escherichia coli
| Autor: | J E Irvin, R.M. Teather, J.D. Erfle, P J Wood |
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| Rok vydání: | 1988 |
| Předmět: |
Glycoside Hydrolases
Lichenin Bacillus subtilis Biology Biochemistry Chromatography Affinity Substrate Specificity Laminarin chemistry.chemical_compound Affinity chromatography Escherichia coli Bacteroides Cloning Molecular Glucans Molecular Biology chemistry.chemical_classification Chromatography Chromatofocusing Substrate (chemistry) Cell Biology Hydrogen-Ion Concentration Glucanase Chromatography Ion Exchange biology.organism_classification Xylan Endo-1 3-beta-Xylosidase Enzyme chemistry Research Article |
| Zdroj: | Biochemical Journal. 255:833-841 |
| ISSN: | 1470-8728 0264-6021 |
| Popis: | A 1,3-1,4-beta-D-glucanase (lichenase, 1,3-1,4-beta-D-glucan 4-glucanohydrolase, EC 3.2.1.73) from Bacteroides succinogenes cloned in Escherichia coli was purified 600-fold by chromatography on Q-Sepharose and hydroxyapatite. The cloned enzyme hydrolysed lichenin and oat beta-D-glucan but not starch, CM(carboxymethyl)-cellulose, CM-pachyman, laminarin or xylan. The enzyme had a broad pH optimum with maximum activity at approx. pH 6.0 and a temperature optimum of 50 degrees C. The pH of elution from a chromatofocusing column for the cloned enzyme was 4.7 (purified) and 4.9 (crude) compared with 4.8 for the mixed-linkage beta-D-glucanase activity in B. succinogenes. The Mr of the cloned enzyme was estimated to be 37,200 by gel filtration and 35,200 by electrophoresis. The Km values estimated for lichenin and oat beta-D-glucan were 0.35 and 0.71 mg/ml respectively. The major hydrolytic products with lichenin as substrate were a trisaccharide (82%) and a pentasaccharide (9.5%). Hydrolysis of oat beta-D-glucan yielded a trisaccharide (63.5%) and a tetrasaccharide (29.6%) as the major products. The chromatographic patterns of the products from the cloned enzyme appear to be similar to those reported for the mixed-linkage beta-D-glucanase isolated from Bacillus subtilis. The data presented illustrate the similarity in properties of the cloned mixed-linkage enzyme and the 1,3-1,4-beta-D-glucanase from B. subtilis and the similarity with the 1,4-beta-glucanase in B. succinogenes. |
| Databáze: | OpenAIRE |
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