Production by Clostridium acetobutylicum ATCC 824 of CelG, a cellulosomal glycoside hydrolase belonging to family 9.
The genome sequence of Clostridium acetobutylicum ATCC 824, a noncellulolytic solvent-producing strain, predicts the production of various proteins with domains typical for cellulosomal subunits. Most of the genes coding for these proteins are grouped in a cluster similar to that found in cellulolytic clostridial species, such as Clostridium cellulovorans. CAC0916, one of the open reading frames present in the putative cellulosome gene cluster, codes for CelG, a putative endoglucanase belonging to family 9, and it was cloned and overexpressed in Escherichia coli. The overproduced CelG protein was purified by making use of its high affinity for cellulose and was characterized. The biochemical properties of the purified CelG were comparable to those of other known enzymes belonging to the same family. Expression of CelG by C. acetobutylicum grown on different substrates was studied by Western blotting by using antibodies raised against the purified E. coli-produced protein. Whereas the antibodies cross-reacted with CelG-like proteins secreted by cellobiose- or cellulose-grown C. cellulovorans cultures, CelG was not detectable in extracellular medium from C. acetobutylicum grown on cellobiose or glucose. However, notably, when lichenan-grown cultures were used, several bands corresponding to CelG or CelG-like proteins were present, and there was significantly increased extracellular endoglucanase activity.
López-Contreras
Ana M
AM
Laboratory of Microbiology and Agrotechnological Research Institute, Wageningen University and Research Centre, Wageningen, The Netherlands. a.m.lopezcontreras@ato.wag-ur.nl
Martens
Aernout A
AA
Szijarto
Nora
N
Mooibroek
Hans
H
Claassen
Pieternel A M
PA
van der Oost
John
J
de Vos
Willem M
WM
D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D002480:Cellulase / Q000096:biosynthesis* / Q000235:genetics; D002482:Cellulose / Q000378:metabolism; D003013:Clostridium / Q000201:enzymology* / Q000235:genetics; D003470:Culture Media; D005936:Glucans / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000096:biosynthesis / Q000235:genetics; D008969:Molecular Sequence Data; D005810:Multigene Family; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014994:Xylose / Q000378:metabolism
D001426:Bacterial Proteins; D003470:Culture Media; D005936:Glucans; D011994:Recombinant Proteins; D002482:Cellulose; D014994:Xylose; D006026:Glycoside Hydrolases; C081472:celG protein, Bacteria; D002480:Cellulase; C015968:lichenin