Purification and characterization of multiple forms of endochitinase from wheat leaves

Abstract The four major forms (I,II,III,VI) of chitinase were purified from leaves of wheat ( Triticum aestivum L.) seedlings by affinity chromatography of chitin, and high-performance gel-filtration and ion-exchange chromatography. The isozymes has similar m r s (33, 31.5, 34 and 33 kDa, respectively) as determined by SDS-polyacrylamide gel electrophoresis but widely differing isoelectric points of 5.8, 5.85, 6.7 and 9.1 respectively. Amino acid analyses and double immunodiffusion using specific polyclonal antisera raised in rabbits indicated that the enzymes were closely related. Analysis of the initial products from the digestion of [ 3 H]chitin indicated that all the isozymes were endo in action, with oligomers from the dimer up to at least the decamer being evident in the digests; minor differences were observed between the isozymes in the relative proportions of the oligomeric products. The results are discussed in relation to the release of lignification-elicting chitin oligosaccharides from fungal cell walls during attempted infections of wheat.