| LAMR1与PER1蛋白作用位点的筛选 |
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| 引用本文： | 常莉,李文辉,汪宇辉,刘延友,江舟,肖静,郭慧玲,王正荣.LAMR1与PER1蛋白作用位点的筛选[J].现代生物医学进展,2013,13(1):14-18. |
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| 作者姓名： | 常莉 李文辉 汪宇辉 刘延友 江舟 肖静 郭慧玲 王正荣 |
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| 作者单位： | 1. 云南省肿瘤医院昆明医科大学第三附属医院放疗中心 云南昆明650118
2. 四川大学华西基础医学与法医学院时间生物学卫生部重点实验室 四川成都610041 |
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| 基金项目： | 国家自然科学基金项目(81101693);云南省科技厅-昆明医科大学联合专项(2011FB206) |
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| 摘 要： | 目的：筛选出与PER1蛋白相互作用的LAMR1的核心位点。方法：采用盒式定点突变法,对重组目的质粒pGADT7-Rec/Lamr1201-295的插入片段Lamr1201-295羧基末端的205-216氨基酸序列RDPEEIEKEEQA进行定点突变,并以hPER1蛋白的bHLH-PAS结构域为诱饵蛋白,通过酵母双杂交系统确定这4种不同的突变LAMR1201-295片段与hPER1的相互作用。结果：营养缺陷培养基筛选显示4种转化菌在二缺SD/-Leu/-Trp、三缺SD/-His/-Leu/-Trp和四缺SD/-Ade/-His/-Leu/-Trp固体培养基上都可以生长;β-半乳糖苷酶印膜法检测结果显示4种转化菌均可显色。结论：突变片段LAMR1-RDP,LAMR1-EEI,LAMR1-EKE和LAMR1-EQA都能够与hPER1的bHLH-PAS结构域在酵母中发生相互作用。提示LAMR1的205-216Aa可能不是其与hPER1相互作用的核心氨基酸序列。
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| 关 键 词： | PER1 LAMR1 相互作用 酵母双杂交 定点突变 |
Screening for Key Region of Laminin Receptor 1 Interacted with PERIOD 1 |
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| CHANG Li,LI Wen-hui,WANG Yu-hui,LIU Yan-you,JIANG Zhou,XIAO Jing,GUO Hui-ling,WANG Zheng-rong.Screening for Key Region of Laminin Receptor 1 Interacted with PERIOD 1[J].Progress in Modern Biomedicine,2013,13(1):14-18. |
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| Authors: | CHANG Li LI Wen-hui WANG Yu-hui LIU Yan-you JIANG Zhou XIAO Jing GUO Hui-ling WANG Zheng-rong |
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| Institution: | 1 Department of Radiotherapy Oncology,Tumor Hospital of Yunnan Province/The Third Affliated Hospital of Kunming Medical University,Kunming,Yunnan,650118,China;2 Health Ministry Key Laboratory of Chronobiology,College of Basic Medicine and Forensic Medicine,Sichuan University,Chengdu,Sichuan,610041,China) |
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| Abstract: | Objective： To define the key region of LAMR1 to interact with hPER1. Methods： With cassette mutagenesis, pGADT7-Rec/hLamr1201-295 was mutated, targeting the sequence RDP, EEI, EKE and EQA at carboxyl terminal, respectively. And the interaction between PER1 and four kinds of mutated LAMR1201-295 were further identified using a yeast two-hybrid system. Results： Clones containing bHLH-PAS domain of Per1 and Lamrl could be seen in SD/-Leu/-Trp, SD/-His/-Leu/-Trp and SD-Ade/-His/-Leu/-Trp selection mediums. The blue stain was also obviously observed in colony-lift filter 13-galactosidase assay. Conclusions： hPER1 could interact with all mutant LAMR1, which suggested that the sequence RDPEEIEKEEQA at carboxyl terminal is not necessary for the interaction to occur. |
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| Keywords: | PER1 LAMR1 Interaction Yeast two-hybrid system Cassette mutagenesis |
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