Ohtsuki T, Taniguchi Y, Kohno K, Fukuda S, Usui M, Kurimoto M
Fujisaki Institute, Hayashibara Biochemical Laboratories, Inc, Okayama, Japan.
Allergy. 1995 Jun;50(6):483-8. doi: 10.1111/j.1398-9995.1995.tb01183.x.
We examined Cry j 2, a major allergen of Japanese cedar (Cryptomeria japonica) pollen, for polygalacturonase enzyme activity, since a nucleotide sequence of cDNA of Cry j 2 showed a significant homology with that of tomato polygalacturonase. Polygalacturonase is well known to depolymerize preferentially polygalacturonic acid (PGA) by hydrolysis. However, Cry j 2 did not act on PGA, but was found to depolymerize pectin and methylesterified PGA in a dose-dependent manner. The substrate specificity of Cry j 2 was different from that of polygalacturonase derived from Aspergillus niger. The depolymerizing activity of Cry j 2 reached a maximum at 50%-60% of methylesterification of PGA. In contrast, polygalacturonase showed its maximum activity of PGA, and the activity decreased as the degree of methylesterification increased. Interestingly, the pectin-depolymerizing activity of Cry j 2 was due to a hydrolysis, but not a lyase, activity which splits the glycosidic bonds by beta-elimination, since no unsaturated uronides were found by measurement of absorbance at 235 nm in the reaction mixture. The enzyme activity was markedly inhibited by anti-Cry j 2 antibodies. These results indicate that Cry j 2 probably has polymethylgalacturonase enzyme activity, as postulated by von Neukom in 1963, although existence of this activity has not yet been proven.
我们检测了日本柳杉花粉的主要过敏原Cry j 2的多聚半乳糖醛酸酶活性，因为Cry j 2的cDNA核苷酸序列与番茄多聚半乳糖醛酸酶的序列有显著同源性。众所周知，多聚半乳糖醛酸酶通过水解优先使聚半乳糖醛酸（PGA）解聚。然而，Cry j 2对PGA无作用，但被发现能以剂量依赖的方式使果胶和甲基酯化的PGA解聚。Cry j 2的底物特异性与黑曲霉来源的多聚半乳糖醛酸酶不同。Cry j 2的解聚活性在PGA甲基酯化程度为50%-60%时达到最大值。相比之下，多聚半乳糖醛酸酶对PGA表现出最大活性，且随着甲基酯化程度的增加活性降低。有趣的是，Cry j 2的果胶解聚活性是由于水解活性，而非通过β-消除作用裂解糖苷键的裂解酶活性，因为在反应混合物中通过235nm处的吸光度测量未发现不饱和糖醛酸。该酶活性被抗Cry j 2抗体显著抑制。这些结果表明，尽管这种活性尚未得到证实，但Cry j 2可能具有1963年冯·诺伊科姆所假设的聚甲基半乳糖醛酸酶活性。
