CELG FROM CLOSTRIDIUM-CELLULOLYTICUM - A MULTIDOMAIN ENDOGLUCANASE ACTING EFFICIENTLY ON CRYSTALLINE CELLULOSE

Citation
L. Gal et al., CELG FROM CLOSTRIDIUM-CELLULOLYTICUM - A MULTIDOMAIN ENDOGLUCANASE ACTING EFFICIENTLY ON CRYSTALLINE CELLULOSE, Journal of bacteriology, 179(21), 1997, pp. 6595-6601
Citations number
36
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Microbiology
Journal title
ISSN journal
0021-9193
Volume
179
Issue
21
Year of publication
1997
Pages
6595 - 6601
Database
ISI
SICI code
0021-9193(1997)179:21<6595:CFC-AM>2.0.ZU;2-8
Abstract
The gene coding for CelG, a family 9 cellulase from Clostridium cellul olyticum, was cloned and overexpressed in Escherichia coli. Four diffe rent forms of the protein were genetically engineered, purified, and s tudied: CelGL (the entire form of CelG), CelGcat1 (the catalytic domai n of CelG alone), CelGcat2 (CelGcat1 plus 91 amino acids at the beginn ing of the cellulose binding domain [CBD]), and GST-CBDCelG (the CBD o f CelG fused to glutathione S-transferase). The biochemical properties of CelG were compared with those of CelA, an endoglucanase from C. ce llulolyticum which was previously studied. CelG, like CelA, was found to have an endo cutting mode of activity on carboxymethyl cellulose (C MC) but exhibited greater activity on crystalline substrates (bacteria l microcrystalline cellulose and Avicel) than CelA. As observed with C elA, the presence of the nonhydrolytic miniscaffolding protein (miniCi pC(1)) enhanced the activity of CelG on phosphoric acid swollen cellul ose (PASC), but to a lesser extent. The absence of the CBD led to the complete inactivation of the enzyme. The abilities of CelG and GST-CBD CelG to bind various substrates were also studied. Although the entire enzyme is able to bind to crystalline cellulose at a limited number o f sites, the chimeric protein GST-CBDCelG does not bind to either of t he tested substrates (Avicel and PASC). The lack of independence betwe en the two domains and the weak binding to cellulose suggest that this CBD-like domain may play a special role and be either directly or ind irectly involved in the catalytic reaction.