Fluorescent Indicators for Cyclic GMP Based on Cyclic GMP-Dependent Protein Kinase I and Green Fluorescent Proteins
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- 作者：Moritoshi Sato ; Naoki Hida ; Takeaki Ozawa ; and Yoshio Umezawa
- 刊名：Analytical Chemistry
- 出版年：2000
- 出版时间：December 15, 2000
- 年：2000
- 卷：72
- 期：24
- 页码：5918 - 5924
- 全文大小：460K
- 年卷期：v.72,no.24(December 15, 2000)
- ISSN：1520-6882
文摘
We describe herein fluorescent indicators for cyclic GMP(cGMP) in single living cells. cGMP-dependent proteinkinase I
(PKG I), a receptor for cGMP, was fused withblue- and red-shifted green fluorescent proteins (GFPs)to its N- and C-termini, respectively. Using PKG I
1-47,in which the dimerization domain was deleted, fluorescence resonance energy transfer between the GFPs wasfound to increase upon cGMP-induced conformationalchange in PKG I1-47. We demonstrated that thus-developed fluorescent indicators reversibly responded tocGMP that was produced in nitric oxide-stimulatedHEK293 cells. The present genetically encoded fluorescent indicators open a way not only for understanding thedynamics of cGMP signaling in single cells and organismsbut also for discovering pharmaceuticals such as isoform-specific inhibitors for phosphodiesterases.
(PKG I), a receptor for cGMP, was fused withblue- and red-shifted green fluorescent proteins (GFPs)to its N- and C-termini, respectively. Using PKG I1-47,in which the dimerization domain was deleted, fluorescence resonance energy transfer between the GFPs wasfound to increase upon cGMP-induced conformationalchange in PKG I1-47. We demonstrated that thus-developed fluorescent indicators reversibly responded tocGMP that was produced in nitric oxide-stimulatedHEK293 cells. The present genetically encoded fluorescent indicators open a way not only for understanding thedynamics of cGMP signaling in single cells and organismsbut also for discovering pharmaceuticals such as isoform-specific inhibitors for phosphodiesterases.