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Thesis Advisor张宪孔 ; 李立人
Degree Grantor中国科学院水生生物研究所
Place of Conferral中国科学院水生生物研究所
Degree Discipline水生生物学
Abstract用~(35)S-Met标记法研究豌豆完整叶绿体内蛋白的合成,发现新合成的RubisCO大亚基与一高分子量的蛋白结合在一起形成复合物。经ATP处理后,复合物解离,释放出的大亚基参与了RubisCO的装配,这一高分子量的蛋白即为RubisCO亚基结合蛋白,该蛋白在RubisCO装配中起作用。用豌豆叶片提取液做初始材料提纯结合蛋白,经普通PAGE、SDS-PAGE、琼脂糖双扩散鉴定纯度达90%以上。测定纯化的结合蛋白表面疏基数为13 ± 1个,总疏基数为37 ± 1个。远紫外CD光谱分析结合蛋白的α-螺旋度为39%,具有典型的α-螺旋结构的光谱特性。50 ℃加热10分钟,结合蛋白既不解离也不变性,说明该蛋白是一热稳定性蛋白。用ATP处理蛋白浓度较低的叶片提取液,发现ATP能使结合蛋白解离,而ADP无此作用。在蛋白浓度高的情况下,ATP对结合蛋白无解离作用。结合蛋白的抗体与结合蛋白有高度专一的免疫反应,与豌豆RubisCO无免疫反应,与烟草RubisCO大、小亚基也无免疫交叉反应。
Other AbstractIt was discovered that newly synthesized RubisCO large subunits could bound to another high molecular weight protein that occurs as a complex, when the intact chloroplast was incubated with S-Met. The addition of ATP caused dissociation of the complex. The released large subunits then assembled into RubisCO. This high molecular weight protein is called RubisCO subunit binding protein, the protein plays an important role in the assembly of RubisCO. The purification of binding protein was directly started with extract of leaves. The purified binding protein was judged by ND-PAGE, SDS-PAGE and agarose double immunodiffusion. Finally it was obtained that 90% of then protein was the binding protein. It was measured that the surface SH groups of the purified binding protein were 13 ± 1 and the total SH groups were 37 ± 1. The measured and calculated results of the intrinsic region CD spectra showed that there was 39% a-helix and a typical character of a-helix spectra in the binding protein. The binding protein was not dissociated and denatured by heating at 50 ℃. This result shows that the binding protein is a stable protein against heat. The addition of ATP to extract of leaves, which had lower concentration of protein, caused dissociation of the binding protein dodecamer, and ADP did not cause its dissociation of the binding protein. Antibodies raised against the purified pea binding protein had highly specific cross-reaction with the binding protein, but it did not cross-react with the pea RubisCO, nor with the RubisCO large and small subunit from tobacco.
Document Type学位论文
Recommended Citation
GB/T 7714
赵若虹. RubisCO亚基结合蛋白的纯化及其性质、功能的研究[D]. 中国科学院水生生物研究所. 中国科学院水生生物研究所,1990.
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