Nucleotide sequence of the Ruminococcus albus SY3 endoglucanase genes celA and celB.
The complete nucleotide sequences of Ruminococcus albus genes celA and celB coding for endoglucanase A (EGA) and endoglucanase B (EGB), respectively, have been determined. The celA structural gene consists of an open reading frame of 1095 bp. Confirmation of the nucleotide sequence was obtained by comparing the predicted amino acid sequence with that derived by N-terminal analysis of purified EGA. The celB structural gene consists of an open reading frame of 1227 bp; 7 bp upstream of the translational start codon of celB is a typical gram-positive Shine-Dalgarno sequence. The deduced N-terminal region of EGB conforms to the general pattern for the signal peptides of secreted prokaryotic proteins. The complete celB gene, cloned into pUC vectors, caused lethality in Escherichia coli. In contrast, celA cloned in pUC18, under the control of lacZp, directed high-level synthesis of EGA in E. coli JM83. EGA in cell-free extract, purified to near homogeneity by ion-exchange chromatography, had a Mr of 44.5 kDa. Gene deletion and subcloning studies with celA revealed that EGA hydrolysed both CMC and xylan, and did not contain discrete functional domains. EGA and EGB showed considerable homology with each other, in addition to exhibiting similarity with Eg1 (R. albus), EGE (Clostridium thermocellum) and End (Butyrivibrio fibrisolvens).
Poole
D M
DM
Department of Agricultural Biochemistry and Nutrition, University of Newcastle upon Tyne, UK.
Hazlewood
G P
GP
Laurie
J I
JI
Barker
P J
PJ
Gilbert
H J
HJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D002480:Cellulase / Q000235:genetics* / Q000378:metabolism; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010459:Peptococcaceae / Q000201:enzymology / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid; D013379:Substrate Specificity
D002480:Cellulase