植物の自己防御タンパク質、キチナーゼの構造と機能に関する研究

对山药地上块茎几丁质酶进行了纯化，并对其蛋白质结构和在植物体内的定位进行了研究。为了纯化，开发了使用季铵离子洗涤剂的新方法。对纯化的几丁质酶进行氨基酸序列测定，结果表明高分子量和低分子量的几丁质酶分别为I类和III类几丁质酶。进一步确定了Ⅰ类几丁质酶的S-S键的位置。测定的位置与大麦Ⅱ类几丁质酶相同，但与黑麦Ⅱ类几丁质酶C-末端的位置不同。这表明C-末端区域的柔性。对植物几丁质酶的结构关系进行了系统遗传学研究。所建立的蛋白质谱系显示I类和II类几丁质酶中的两个亚类。估计高分子量亚类是祖先分子。在III类几丁质酶中没有检测到亚类。对山药Ⅰ类和Ⅲ类几丁质酶进行了酶化学定位研究。染色结果表明，Ⅰ类几丁质酶定位于所有幼嫩组织，Ⅲ类几丁质酶仅定位于块茎。这一结果表明，这些几丁质酶在植物体内具有不同的防御作用。

Yam aerial tuber chitinase, one of the inherent self defense protein, was purified and determined their protein structure and their localization in plant. For the purification, new method using quaternary ammonium ion detergent was developed. The purified chitinases were determined their amino acid sequences, which revealed that the high and low molecular weight isoforms were class I and class III chitinases, respectively. Further, the positions of S-S bonds of class I chitinase were determined. The determined positions were same as barley class II chitinase, but different from those of rye class II chitinase at C-terminal region. This indicates the flexibility at the C-terminal region. The structural relationships of plant chitinases were phylogenetically studied. The established protein genealogy showed the two subclasses in class I and class II chitinases. The high molecular weight subclass was estimated to be an ancestral molecule. No subclass was detected in class III chitinases. The localizations of yam class I and class III chitinases were immunohistochemically studied. The staining showed that the class I chitinase locates every young tissue and class III chitinase locates only tubers. This result indicates that these chitinases possess different defense role in plant.