Characterizing the pH-dependent stability and catalytic mechanism of the family 11 xylanase from the alkalophilic Bacillus agaradhaerens.
The xylanase, BadX, from the alkalophilic Bacillus agaradhaerens was cloned, expressed and studied in comparison to a related family 11 xylanase, BcX, from B. circulans. Despite the alkaline versus neutral conditions under which these bacteria grow, BadX and BcX both exhibit optimal activity near pH 5.6 using the substrate o-nitrophenyl beta-xylobioside. Analysis of the bell-shaped activity profile of BadX yielded apparent pK(a) values of 4.2 and 7.1, assignable to its nucleophile Glu94 and general acid Glu184, respectively. In addition to having an approximately 10-fold higher k(cat)/K(m) value with this substrate at pH 6 and 40 degrees C, BadX has significantly higher thermal stability than BcX under neutral and alkaline conditions. This enhanced stability, rather than a shift in its pH-optimum, may allow BadX to hydrolyze xylan under conditions of elevated temperature and pH.
Poon
David K Y
DK
The Department of Chemistry, University of British Columbia, BC, V6T 1Z1, Vancouver, Canada.
Webster
Philip
P
Withers
Stephen G
SG
McIntosh
Lawrence P
LP
D001407:Bacillus / Q000201:enzymology*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D011489:Protein Denaturation; D043325:Xylan Endo-1,3-beta-Xylosidase; D014995:Xylosidases / Q000378:metabolism*
D014995:Xylosidases; D043325:Xylan Endo-1,3-beta-Xylosidase