Cloning and characterization of two endoxylanases from the cereal phytopathogen Fusarium graminearum and their inhibition profile against endoxylanase inhibitors from wheat.
Two genes encoding family 11 endo-beta-1,4-xylanases (XylA, XylB) from Fusarium graminearum were cloned and expressed in Escherichia coli. The amount of active endoxylanase in the cytoplasmic soluble fraction was considerably improved by varying different expression parameters, including host strain and temperature during induction. Both recombinant endoxylanases showed a temperature optimum around 35 degrees C and neutral pH optima (around pH 7 and 8 for XylB and XylA, respectively). For the first time this allowed one to test endoxylanases of a phytopathogenic organism for inhibition by proteinaceous endoxylanase inhibitors TAXI and XIP. Whereas XylA and XylB were inhibited by TAXI-I, no inhibition activity could be detected upon incubation with XIP-I. The insensitivity of both F. graminearum endoxylanases towards XIP is surprising, since the latter is typically active against endoxylanases produced by (aerobic) fungi. As F. graminearum is an important phytopathogen, these findings have implications for the role of endoxylanase inhibitors in plant defence.
Beliën
Tim
T
Laboratory of Gene Technology, Katholieke Universiteit Leuven, Kasteelpark Arenberg 21, B-3001 Leuven, Belgium.
Van Campenhout
Steven
S
Van Acker
Maarten
M
Volckaert
Guido
G
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D002523:Edible Grain / Q000382:microbiology*; D043364:Endo-1,4-beta Xylanases / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005670:Fusarium / Q000201:enzymology* / Q000235:genetics / Q000472:pathogenicity; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D013696:Temperature; D014908:Triticum / Q000737:chemistry*
D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D043364:Endo-1,4-beta Xylanases