Substrate recognition and hydrolysis by a fungal xyloglucan-specific family 12 hydrolase.
Biochemical studies to elucidate the structural basis for xyloglucan specificity among GH12 xyloglucanases are lacking. Accordingly, the substrate specificity of a GH12 xyloglucanase from Aspergillus niger (AnXEG12A) was investigated using pea xyloglucan and 12 xylogluco-oligosaccharides, and data were compared to a structural model of the enzyme. The specific activity of AnXEG12A with pea xyloglucan was 113 micromol min(-1)mg(-1), and apparent k(cat) and K(m) values were 49 s(-1) and 0.54 mg mL(-1), respectively. These values are similar to previously published results using xyloglucan from tamarind seed, and suggest that substrate fucosylation does not affect the specific activity of this enzyme. AnXEG12A preferred xylogluco-oligosaccharides containing more than six glucose units, and with xylose substitution at the -3 and +1 subsites. The specific activities of AnXEG12A on 100 microM XXXGXXXG and 100 microM XLLGXLLG were 60+/-4 and 72+/-9 micromol min(-1)mg(-1), respectively. AnXEG12A did not hydrolyze XXXXXXXG, consistent with other data that demonstrate the requirement for an unbranched glucose residue for hydrolysis by this enzyme.
Department of Chemistry and Biochemistry, Concordia University, 7141 Sherbrooke St. West, Montreal, Quebec, Canada H4B 1R6.
Justin
J
Powlowski
Sonam
S
Mahajan
Matthieu
M
Schapira
Emma R
ER
Master
D001234:Aspergillus niger / Q000201:enzymology*; D002240:Carbohydrate Sequence; D005936:Glucans / Q000737:chemistry / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000378:metabolism*; D006868:Hydrolysis; D008958:Models, Molecular; D008969:Molecular Sequence Data; D018532:Pisum sativum / Q000737:chemistry; D011487:Protein Conformation; D013379:Substrate Specificity; D014990:Xylans / Q000737:chemistry / Q000378:metabolism*
D005936:Glucans; D014990:Xylans; C029353:xyloglucan; D006026:Glycoside Hydrolases; C072818:xyloglucan endo(1-4)-beta-D-glucanase