The N-terminal region of an endoglucanase from Pseudomonas fluorescens subspecies cellulosa constitutes a cellulose-binding domain that is distinct from the catalytic centre.
The substrate specificity of an endoglucanase (EGB) from Pseudomonas fluorescens subspecies cellulosa was determined. The enzyme was most active against barley beta-glucan, but showed significant activity against amorphous and crystalline cellulose. EGB was purified to homogeneity by affinity chromatography with crystalline cellulose (Avicel). The Mr of the purified enzyme was 50,000, which is in good agreement with the size of EGB deduced from the nucleotide sequence of the celB gene, coding for EGB. The N-terminal region of the mature form of EGB showed strong homology to another endoglucanase and to a xylanase expressed by the same organism; homologous sequences included highly conserved serine-rich regions. Truncated forms of celB, in which the gene sequence encoding the conserved domain had been deleted, directed the synthesis of a functional endoglucanase that did not bind to crystalline cellulose. This indicates that the conserved region of endoglucanases and xylanases expressed by P. fluorescens subsp. cellulosa constitutes a cellulose-binding domain, which is distinct from the active centre. The possible role of this substrate-binding region is discussed.
Gilbert
H J
HJ
Department of Agricultural Biochemistry and Nutrition, University of Newcastle upon Tyne, UK.
Hall
J
J
Hazlewood
G P
GP
Ferreira
L M
LM
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D002480:Cellulase / Q000235:genetics*; D002482:Cellulose / Q000378:metabolism*; D002872:Chromosome Deletion; D004926:Escherichia coli / Q000201:enzymology; D006026:Glycoside Hydrolases / Q000096:biosynthesis / Q000235:genetics; D008969:Molecular Sequence Data; D011549:Pseudomonas / Q000235:genetics*; D011551:Pseudomonas fluorescens / Q000235:genetics*; D015183:Restriction Mapping; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D043325:Xylan Endo-1,3-beta-Xylosidase
D002482:Cellulose; D006026:Glycoside Hydrolases; D043325:Xylan Endo-1,3-beta-Xylosidase; D002480:Cellulase