Physiological role of the pyrenoids in the green alga Bryopsis maxima.

本文采用以下几种方法研究了类果核在海洋藻类Bryopsis Maxima中的生理作用：一、免疫细胞化学研究。(1)用免疫组织化学方法研究了核酮糖1，5-二磷酸羧化酶/加氧酶(Rubisco)在叶绿体中的定位。制备了3株抗Rubisco大亚基的单抗。免疫荧光和免疫酶学研究表明，Rubisco的大亚基主要集中在类蛋白核和类蛋白核周围的淀粉颗粒表面。(2)当藻类在黑暗中保存或成熟以排出配子体时，类蛋白核中的Rubisco浓度降低。(2)生化研究(1)通过乙醚处理和1.8M磷酸盐缓冲液蔗糖密度梯度离心法分离得到完整的类蛋白核基质。纯化后的蛋白核含有Rubisco和10多个小分子多肽。它们在溶解时也表现出Rubisco活性(比活性为每毫克蛋白每分钟固定0.62微克二氧化碳)。(2)类果核具有NADH-硝酸还原酶活性，并含有亚硝酸盐。比活力为每毫克蛋白每分钟生成0.024微克分子亚硝酸根，是叶绿体粗提物的80倍。III.分子生物学研究克隆了Rubisco大亚基基因，并进行了序列测定。该基因的内含子很大(2.3kbp)，产物没有经过翻译后加工。

Physiological role of pyrenoids in the marine alga, Bryopsis maxima was studies using several methods as follows.I. Immunocytochemical researches. (1) The localization of ribulose 1,5-bisphosphate carboxylase/oxygenase(RuBisCO) in chloroplasts was examined by immunological techniques. Three strains of monoclonal antibodies against the large subunit of RuBisCO were made for the study. Immunofluorescence and immunoenzymatic studies showed that the large subunit of RuBisCO was concentrated in pyrenoids and on the surface of starch grains surrounding the pyrenoids.(2) The concentration of RuBisCO in Pyrenoids decreased when the alga was kept in the dark or matured to eject the gametes.II Biochemical studies (1) Intact pyrenoid core matrix was isolated by diethyl ether treatment and sucrose density gradient centrifugation using 1.8 M phosphate buffer. The purified pyrenoids contained RuBisCO and more than 10 minor polypeptides. They also showed RuBisCO activity when solubilized(specific activity was 0.62 umol CO2 fixed per mg protein per min). (2) The pyrenoids showed NADH-nitrate reductase activity and contained nitrite. The specific activity was 0.024 umols NO2 formed per mg protein per min, which was 80 times greater than that in the crude chloroplasts extract.III. Molecular biological research The gene of the large subunit of RuBisCO was cloned and sequenced. The gene has a large (2.3 kbp) intron and the product was not subjected to the posttranslational processing.