Sequential activation and deactivation of protein function using spectrally differentiated caged phosphoamino acids.
Sequential activation and deactivation of protein function using spectrally differentiated caged phosphoamino acids.
复制标题
DOI：
10.1021/ja2028074
复制
发表时间：
2011-07-27
影响因子：
15
通讯作者：
Imperiali, Barbara
中科院分区：
文献类型：
--
作者：
Goguen, Brenda N.;Aemissegger, Andreas;Imperiali, Barbara
包括1-（2-硝基苯基）乙基（NPE）基团在内的光耐性笼化基团已被用于探测许多生物过程，包括蛋白质磷酸化。尽管对npe笼养磷氨基酸的研究提供了有价值的信息，但这些研究仅限于在单个实验中使用一种笼养物种。为了扩大这些工具的范围，我们开发了一种方法，在一个系统中依次释放两种不同的磷酸肽，从而能够对多个磷酸化事件进行询问。我们提出了[7-（二乙基氨基）香豆素-4-基]甲基（DEACM）笼化磷酸化丝氨酸、苏氨酸和酪氨酸构建块的合成，用于基于fmoc的固相肽合成，以便于将这些残基结合到肽和蛋白质中。将DEACM-和npe -笼的磷酸肽暴露在420 nm光下，选择性地释放DEACM基团，而不影响npe -笼的肽。这使得随后的365 nm照射事件能够去除NPE基团并释放第二个磷酸肽。我们通过将其应用于两种波长的光来控制Wip1磷酸酶，证明了这种通用顺序解锁方法的多功能性。
Photolabile caging groups, including the 1-(2-nitrophenyl)ethyl (NPE) group, have been applied to probe many biological processes, including protein phosphorylation. Although studies with NPE-caged phosphoamino acids have provided valuable information, these investigations have been limited to the use of only one caged species in a single experiment. To expand the scope of these tools, we developed an approach to sequentially uncage two different phosphopeptides in one system, enabling interrogation of multiple phosphorylation events. We present the synthesis of [7-(diethylamino)coumarin-4-yl]methyl (DEACM)-caged phosphorylated serine, threonine, and tyrosine building blocks for Fmoc-based solid phase peptide synthesis to allow convenient incorporation of these residues into peptides and proteins. Exposure of DEACM- and NPE-caged phosphopeptides to 420 nm light selectively releases the DEACM group without affecting the NPE-caged peptide. This then enables a subsequent irradiation event at 365 nm to remove the NPE group and liberate a second phosphopeptide. We demonstrate the versatility of this general sequential uncaging approach by applying it to control the Wip1 phosphatase with two wavelengths of light.
登录
查看更多内容
影响因子：
16.6
作者：
Tarrant MK;Cole PA
通讯作者：
Cole PA
影响因子：
46.9
作者：
Nguyen, A;Rothman, DM;Yaffe, MB
通讯作者：
Yaffe, MB
影响因子：
15
作者：
Kotzur, Nico;Briand, Benoit;Hagen, Volker
通讯作者：
Hagen, Volker
影响因子：
15
作者：
Vázquez, ME;Nitz, M;Imperiali, B
通讯作者：
Imperiali, B
影响因子：
4.8
作者：
Mehta, D;Rahman, A;Malik, AB
通讯作者：
Malik, AB