Cross-inhibitory activity of cereal protein inhibitors against alpha-amylases and xylanases.
The purification and characterisation of a xylanase inhibitor (XIP-I) from wheat was reported previously. In our current work, XIP-I is also demonstrated to have the capacity to inhibit the two barley alpha-amylase isozymes (AMY1 and AMY2). XIP-I completely inhibited the activity of AMY1 and AMY2 towards insoluble Blue Starch and a soluble hepta-oligosaccharide derivative. A ternary complex was formed between insoluble starch, a catalytically inactive mutant of AMY1 (D180A), and XIP-I, suggesting that the substrate-XIP-I interaction is necessary for inhibition of barley alpha-amylases. K(i) values for alpha-amylase inhibition, however, could not be calculated due to the nonlinear nature of the inhibition pattern. Furthermore, surface plasmon resonance and gel electrophoresis did not indicate interaction between XIP-I and the alpha-amylases. The inhibition was abolished by CaCl(2), indicating that the driving force for the interaction is different from that of complexation between the barley alpha-amylase/subtilisin inhibitor (BASI) and AMY2. This is the first report of a proteinaceous inhibitor of AMY1. BASI, in addition, was demonstrated to partially inhibit the endo-1,4-beta-D-xylanase from Aspergillus niger (XylA) of glycoside hydrolase family 11. Taken together, the data demonstrate for the first time the dual target enzyme specificity of BASI and XIP-I inhibitors for xylanase and alpha-amylase.
Sancho
Ana I
AI
Institute of Food Research, Norwich Research Park, Colney, Norwich NR4 7UA, UK.
Faulds
Craig B
CB
Svensson
Birte
B
Bartolomé
Begoña
B
Williamson
Gary
G
Juge
Nathalie
N
D000818:Animals; D002122:Calcium Chloride / Q000378:metabolism; D002417:Cattle; D004591:Electrophoresis, Polyacrylamide Gel; D001467:Hordeum / Q000201:enzymology; D007527:Isoenzymes / Q000378:metabolism; D012710:Serum Albumin, Bovine / Q000378:metabolism; D014908:Triticum / Q000378:metabolism; D014360:Trypsin Inhibitor, Kunitz Soybean / Q000302:isolation & purification / Q000378:metabolism*; D043325:Xylan Endo-1,3-beta-Xylosidase; D014995:Xylosidases / Q000037:antagonists & inhibitors*; D000516:alpha-Amylases / Q000037:antagonists & inhibitors*
D007527:Isoenzymes; C092772:barley alpha-amylase-subtilisin inhibitor; D012710:Serum Albumin, Bovine; D014360:Trypsin Inhibitor, Kunitz Soybean; D014995:Xylosidases; D000516:alpha-Amylases; D043325:Xylan Endo-1,3-beta-Xylosidase; D002122:Calcium Chloride