IQ 基序突变对AtIQM1的钙调素结合活性的影响

doi:10.13560/j.cnki.biotech.bull.1985.2014.12.021

生物技术通报 ›› 2014, Vol. 0 ›› Issue (12): 128-132.doi: 10.13560/j.cnki.biotech.bull.1985.2014.12.021

IQ 基序突变对AtIQM1的钙调素结合活性的影响

黄章科 ,张艺能, 莫忠蓁 ,周玉萍, 黄小玲 ,田长恩

广州大学生命科学学院植物抗逆基因功能研究广州市重点实验室，广州 510006

收稿日期:2014-05-26 出版日期:2014-12-08 发布日期:2014-12-12
作者简介:黄章科，男，硕士研究生，研究方向:植物分子遗传学;E-mail:huang.zhangke@163.com
基金资助:
国家自然科学基金项目(31170204)，羊城学者科研项目(12A001G)

Effects of Mutations in IQ Motif of AtIQM1 on Its Calmodulin Binding

Huang Zhangke, Zhang Yi’neng, Mo Zhongzhen, Zhou Yuping, Huang Xiaoling, Tian Chang’en

(Guangzhou Key Laboratory for Functional Study on Stress-Resistant Genes in Plants, School of Life Sciences, Guangzhou University, Guangzhou 510006)

Received:2014-05-26 Published:2014-12-08 Online:2014-12-12

摘要/Abstract

摘要： 旨在确定 IQ 基序中关键氨基酸残基突变对 IQM1 的 CaM 结合活性的影响，利用基因体外定点突变技术将 LQ 缺失或将 L 突变成 S，并通过酵母双杂交分析突变蛋白 iqm1 的 CaM 结合活性。结果显示，用重组质粒 pGADT7-IQM1CDS、pGADT7- IQM1Del113-114、pGADT7-IQM1L113S 或 pGBKT7-CaM5 分别转化酵母 AH109，未出现自激活现象；用 pGADT7-IQM1CDS 和 pGBKT7-CaM5 共转化酵母能在选择性培养基(SD/-Ade/-His/-Leu/-Trp/X-α-Gal)上形成蓝色菌落；而用 pGADT7-IQM1Del113-114 和 pGBKT7-CaM5 或 pGADT7-IQM1L113S 和 pGBKT7-CaM5 共转化酵母则不能在选择性培养基(SD/-Ade/-His/-Leu/-Trp)上形成菌落。结果表明，IQ 基序中 LQ 或者 L 是 IQM1 与 CaM5 结合的关键氨基酸残基。

关键词: AtIQM1, IQ, 基序突变, 酵母双杂交, CaM, 结合

Abstract: To confirm the effects of mutation of key amino acid residues in IQ motif of AtIQM1 on its calmodulin binding, the LQ was deleted or L was mutated to S through the mutagenesis technology in vitro. Then calmodulin binding of the mutant protein iqm1 was analyzed via yeast two-hybrid assay. The self-activation was not observed when the recombinant plasmid pGADT7-IQM1CDS, pGADT7-IQM1Del113-114, pGBKT7- CaM5 or pGADT7-IQM1L113S was transformed into the yeast AH109 strain, respectively. Blue colonies were achieved on the selective agar plates (SD/-Ade/-His/-Leu/-Trp/X-α-Gal)when pGADT7-IQM1CDS and pGBKT7-CaM5 were co-transformed into AH109 yeast strain. However, the yeasts which were co-transformed with pGADT7-IQM1Del113-114 and pGBKT7-CaM5 or pGADT7-IQM1L113S and pGBKT7-CaM5 were not able to grow on the selective medium(SD/-Ade/-His/-Leu/-Trp). The results suggest that the LQ or L in IQ motif is required for AtIQM1 to combine with CaM5.

Key words: AtIQM1, Mutantion in IQ motif, Yeast two hybrid, Calmodulin binding

黄章科 ,张艺能, 莫忠蓁 ,周玉萍, 黄小玲 ,田长恩. IQ 基序突变对AtIQM1的钙调素结合活性的影响[J]. 生物技术通报, 2014, 0(12): 128-132.

Huang Zhangke, Zhang Yi’neng, Mo Zhongzhen, Zhou Yuping, Huang Xiaoling, Tian Chang’en. Effects of Mutations in IQ Motif of AtIQM1 on Its Calmodulin Binding[J]. Biotechnology Bulletin, 2014, 0(12): 128-132.

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IQ 基序突变对AtIQM1的钙调素结合活性的影响

Effects of Mutations in IQ Motif of AtIQM1 on Its Calmodulin Binding

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