Identification of cellular substrate for CaMKII using unnatural amino acid incorporation

使用非天然氨基酸掺入鉴定 CaMKII 的细胞底物

• 批准号: 24650204
• 负责人: SANEYOSHI Takeo
• 金额: $ 2.5万
• 依托单位: Institute of Physical and Chemical Research
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Challenging Exploratory Research
• 财政年份: 2012
• 资助国家: 日本
• 起止时间: 2012-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-24650204/
• 关键词: シナプス; CaMKII; 酵素; 非天然アミノ酸; 基質; リン酸化; シナプス可塑性; 酵素反応; プロテオミクス

It is widely accepted that Calmodulin-kinase II (CaMKII) plays important roles in learning and memory. Although plasticity of synapse regulated by CaMKII, direct targets of CaMKII has poorly identified. To obtain cellular CaMKII substrates, we tried to apply unnatural amino acid incorporation method. Several phenylalanine residues in catalytic domain of CaMKII were made for subsequent incorporation of an unnatural amino acid pBpa. The pBpa were successfully incorporated on CaMKII in HEK293 cells. The covalent bonds were formed upon UV irradiation in cells expressing CaMKII mutants, especially residues, F16, F157, F171. This mutants are useful molecules to identify cellular substrate for CaMKII.

钙调素激酶II（CaMKII）在学习记忆中起重要作用。虽然突触的可塑性受CaMK Ⅱ的调节，但CaMK Ⅱ的直接靶点还不清楚。为了获得细胞CaMKII底物，我们尝试应用非天然氨基酸掺入法。在CaMKII的催化结构域中的几个苯丙氨酸残基用于随后掺入非天然氨基酸pBpa。在HEK293细胞中，pBpa成功地掺入到CaMK II上。在表达CaMKII突变体（特别是残基F16、F157、F171）的细胞中，在紫外线照射后形成共价键。这些突变体是鉴定CaMK II的细胞底物的有用分子。

项目成果

Structural and molecular remodeling of dendritic spine substructures during long-term potentiation.

• DOI: 10.1016/j.neuron.2014.03.021
• 发表时间: 2014-04-16
• 期刊: Neuron
• 影响因子: 16.2
• 作者: Bosch M;Castro J;Saneyoshi T;Matsuno H;Sur M;Hayashi Y
• 通讯作者: Hayashi Y

Synapse reorganization-a new partnership revealed.

Synapse 重组——新的合作伙伴关系揭晓。

• DOI: 10.1002/embj.201488619
• 发表时间: 2014
• 期刊: EMBO J.
• 作者: Saneyoshi T;Hayashi Y.
• 通讯作者: Hayashi Y.

Persistent Interaction between CaMKII and TIAM1 durign long-term potentiation

CaMKII 和 TIAM1 之间的持续相互作用在长时程增强过程中

• 发表时间: 2013
• 作者: Saneyoshi;Takeo. Hayashi;Yasunori
• 通讯作者: Yasunori

光活性化蛋白質によるシナプス構造可塑性制御

光激活蛋白控制突触结构可塑性

• 发表时间: 2013
• 作者: Saneyoshi T.;Hayashi Y.
• 通讯作者: Hayashi Y.