稻瘟菌糖蛋白激发子(CSBI)的纯化及其鉴定

Purification and Identification of a Glycoprotein Elicitor(CSBI) from Magnaporthe grisea

稻瘟菌(Magnaporthegrisea)ZC13菌株97-151a菌丝经离心、超滤、SephacrylS-100凝胶柱、DEAE-SepharoseFF阴离子交换柱层析,纯化获得糖蛋白激发子CSBI。CSBI经SDS-PAGE后银染显示单一条带,糖/蛋白比例约为9.32。CSBI对非亲和性互作水稻叶片中过氧化物酶的诱导显著高于亲和性互作水稻(P<0.05)。经N端氨基酸同源序列比对表明,CSBI与MG07877.4推测蛋白的同源性最高。经基质辅助激光解析电离飞行时间质谱鉴定也表明CSBI是该推测蛋白。

A glycoprotein elicitor, CSBI, isolated from hyphal cell walls of the strain 97-151a of M. grisea race ZC13 was purified by centrifugation, ultra-filtration, gel filtration and anion exchange chromatography （Fig. 1）. CSBI appeared as a single band on silver-stained SDS-PAGE （Fig.2）. Anthrone-colorimetric assay and Coomassie blue G- 250 staining showed that the carbohydrate-to-protein ratio was 9.32 （Table 1）. The induction of peroxidase activity in incompatible interactions was stronger than in compatible interactions （P〈0.05） after treatment with CSBI on rice leaves （Fig.3）. The N-terminal sequence of CSBI was determined to be ITPEAMLSANCCSD, which showed high homology to a 78.671-kD hypothetical protein MG07877.4 （accession No. gi38107424） from M. grisea in NCBI databases. CSBI was either identified as hypothetical protein MG07877.4 by matrix-assisted laser desorption/ionization time of flight mass spectrometry （MALDI-TOF-MS） with 9 matching peptides （Fig.4, Table 2）.