A bifunctional xylanase encoded by the xynA gene of the rumen cellulolytic bacterium Ruminococcus flavefaciens 17 comprises two dissimilar domains linked by an asparagine/glutamine-rich sequence.
The nucleotide sequence of the xynA gene of Ruminococcus flavefaciens 17 was determined and found to consist of a 2862bp open reading frame beginning with a TTG start codon. The predicted product, XYLA, consisted of distinct amino-terminal (A) and carboxy terminal (C) domains (248 amino acids, including a putative signal sequence, and 332 amino acids, respectively) linked by a repetitive sequence (B, 374 amino acids) extraordinarily rich in asparagine (45%) and glutamine (26%) residues. Domains A and C were shown to be capable of expressing xylanase activity independently of each other when suitably truncated derivatives of the xynA coding region were expressed as lacZ fusions. The activities associated with the two domains were shown to differ with respect to the average size of hydrolysis products formed from oat-spelt xylan, with domain C releasing relatively more xylose and domain A more xylo-oligosaccharides. The amino acid sequence of domain A of XYLA closely resembled that of the Bacillus pumilus xynA enzyme (45% identical residues). On the other hand domain C showed significant similarity (33% to 40% identical residues) to a different group of bacterial xylanases and exoglucanases exemplified by the Caldocellum saccharolyticum xynA and celB products. The xynA product is, therefore, a bifunctional enzyme having two dissimilar catalytic domains capable of acting on xylan.
Zhang
J X
JX
Rowett Research Institute, Bucksburn, Aberdeen, UK.
Flint
H J
HJ
D000595:Amino Acid Sequence; D000818:Animals; D001216:Asparagine; D001421:Bacteria, Anaerobic / Q000201:enzymology / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D002417:Cattle; D002482:Cellulose / Q000378:metabolism; D003001:Cloning, Molecular; D003062:Codon; D004269:DNA, Bacterial; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005973:Glutamine; D006026:Glycoside Hydrolases / Q000235:genetics* / Q000378:metabolism; D006095:Gram-Positive Cocci / Q000201:enzymology / Q000235:genetics*; D008969:Molecular Sequence Data; D012091:Repetitive Sequences, Nucleic Acid; D043325:Xylan Endo-1,3-beta-Xylosidase
D001426:Bacterial Proteins; D003062:Codon; D004269:DNA, Bacterial; D005973:Glutamine; D001216:Asparagine; D002482:Cellulose; D006026:Glycoside Hydrolases; D043325:Xylan Endo-1,3-beta-Xylosidase