本文選題：重組人組蛋白　切入點：人血清白蛋白　出處：《中國生物化學與分子生物學報》2016年02期 　論文類型：期刊論文

【摘要】：細胞外組蛋白在膿毒癥、類風濕性關節(jié)炎、急性肺損傷等多種疾病的發(fā)生發(fā)展中起關鍵作用,但由于缺乏合適的標準品,至今無法對患者體內(nèi)的胞外組蛋白進行精確定量,導致在多種感染性疾病中無法根據(jù)血清組蛋白含量對疾病進行精確分級,也無法據(jù)此合理用藥。同時,對患者體內(nèi)胞外組蛋白精確定量也有助于確定細胞毒性機制研究的使用劑量。本研究用大腸桿菌表達單體變性組蛋白H3和H4,親和純化后用梯度稀釋和透析方法,可以得到復性的組蛋白單體H3、H4以及H3/H4復合物。通過對蛋白質在純化過程中穩(wěn)定性的比較,發(fā)現(xiàn)H3/H4復合物較單體更為穩(wěn)定。以該復合物(50μg/m L)處理HUVEC細胞,細胞存活率約為20%,與小牛胸腺組蛋白(200μg/m L)的毒性類似。該復合物引起的細胞毒性可被人血清白蛋白以濃度依賴的形式(0.625~10 mg/m L)緩解,提示其構象基本正確。因此,重組組蛋白H3/H4復合物可以作為精確定量組蛋白的標準品,對基于組蛋白含量的疾病分級和組蛋白毒性機制的研究均有應用價值。
[Abstract]:Extracellular histone plays a key role in the development of many diseases, such as sepsis, rheumatoid arthritis, acute lung injury and so on. In many infectious diseases, it is impossible to classify the disease accurately according to the content of serum histone, and it is impossible to use the medicine reasonably according to the level of serum histone. At the same time, Accurate quantification of extracellular histone was also helpful to determine the dose of cytotoxic mechanism. We used Escherichia coli to express denatured histone H3 and H4 and purified them by gradient dilution and dialysis. By comparing the stability of protein during purification, we found that H3 / H4 complex is more stable than monomers. The H3 / H4 complex was used to treat HUVEC cells with 50 渭 g / mL of H3 / H4 complex. The cell survival rate is about 20%, which is similar to the toxicity of calf thymus histone 200 渭 g / mL. The cytotoxicity induced by this complex can be alleviated by human serum albumin (HSA) in a concentration-dependent manner, suggesting that its conformation is basically correct. The recombinant histone H3 / H4 complex can be used as a standard for accurate quantification of histone. It is valuable for the study of histone classification and the mechanism of histone toxicity based on histone content.
【作者單位】： 上海交通大學醫(yī)學院附屬上海兒童醫(yī)學中心衛(wèi)生部兒童血液腫瘤重點實驗室;上海交通大學醫(yī)學院附屬上海兒童醫(yī)學中心兒科轉化醫(yī)學研究所感染科;
【基金】：國家自然科學青年基金(No.81201450) 上海市科委啟明星計劃(No.14QA1402900) 上海市博士后資助項目基金(No.2014M551418)~~
【分類號】：R329.2

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