水稻条斑病细菌类Harpin蛋白的纯化与特性研究

 用硫酸铵沉淀、制备等电聚焦电泳、阴离子交换层析等方法从水稻条斑病细菌(Xanthomonas oryzae pv. oryzicola,Xooc) RS105菌株突变体M51菌体破碎液中纯化出可激发烟草产生过敏性反应(hypersensitive response,HR)的蛋白类物质,分子量约为25.5 kDa。该物质与梨火疫病菌(Erwinia amylovora))的HarpinEa和水稻白叶枯病菌(Xanthomonas oryzae pv. oryzae,Xoo)的Harpin_Xoo具有相似的生物活性和理化特性:可激发烟草产生典型的HR反应;对热稳定,对蛋白酶K敏感;RNA转录抑制剂放线菌素D、蛋白质合成抑制剂环己酰亚铵和钙离子通道阻断剂氯化镧能够抑制该物质激发烟草产生HR;该物质具有诱导烟草抗TMV的功能。据此,将该物质命名为类Harpin蛋白(Harpin-like protein,HLP_Xooc)。

Purification and characteristics of a harpin-like protein from Xanthomonas oryzae pv.oryzicola

A proteinaceous elicitor which could induce hypersensitive response (HR) reaction on tobacco was purified by (NH4)2SO4 precipitation, ultra speed centrifugation, preparative IEF and ion exchange techniques from cells of strain M51, an hrp mutant of the wild type RS105 (Xanthomonas oryzae pv. oryzicola, Xooc). The molecular weight of this protein was 25. 5 kDa determined by 15% SDS-PAGE. It possessed many similarities with HarpinEa(from Erwinia arnylovora) and HarpinXoo(from Xanthomonas oryzae pv. oryzae, Xoo) in physical and biological properties. This protein was heat-stable and sensitive to protease K. It also could elicit a typical HR reaction on tobacco leaves, which was dispelled by eu-karyotic metabolic inhibitors: actinomycin D, cycloheximide and LaCl3. We also found that the protein had the ability to induce disease-resistance to TMV when sprayed on tobacco. Thus, the protein is designated as HLPxooc(Harpin-like protein).