title: Purification and characterization of a new endoglucanase from Clostridium thermocellum.
abstract: An endoglucanase (1,4-beta-D-glucan glucanohydrolase, EC 3.2.1.4) from the thermophilic anaerobe Clostridium thermocellum was purified to apparent homogeneity without the use of denaturants. No carbohydrate is associated with the endoglucanase. A molecular mass of 76,000 Da was determined by SDS/PAGE. The optimal pH is 7.0 and the enzyme is isoelectric at pH 5.05. The enzyme has a temperature optimum of 70 degrees C and retains approx. 50% of its activity after 48 h at 60 degrees C. Hydrolysis of CM-cellulose takes place with a rapid decrease in viscosity but a slow liberation of reducing sugars, indicating an endoglucanase type of activity. The endoglucanase shows little ability to hydrolyse highly ordered cellulose. Cellobiose inhibits whereas Mg2+ and Ca2+ stimulate the activity. The enzyme is completely inactivated by 1 mM-Hg2+ and is inhibited by a thiol-blocking reagent.
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
authors.forename: M P
authors.initials: MP
authors.lastname: Romaniec
authors.forename: U
authors.initials: U
authors.lastname: Fauth
authors.forename: T
authors.initials: T
authors.lastname: Kobayashi
authors.forename: N S
authors.initials: NS
authors.lastname: Huskisson
authors.forename: P J
authors.initials: PJ
authors.lastname: Barker
authors.forename: A L
authors.initials: AL
authors.lastname: Demain
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000187:drug effects / Q000302:isolation & purification* / Q000378:metabolism; D002480:Cellulase / Q000187:drug effects / Q000302:isolation & purification* / Q000378:metabolism; D002482:Cellulose / Q000378:metabolism; D003013:Clostridium / Q000201:enzymology*; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D008670:Metals / Q000494:pharmacology; D008969:Molecular Sequence Data; D013696:Temperature
chemical_list: D001426:Bacterial Proteins; D008670:Metals; D002482:Cellulose; D002480:Cellulase