427170 Genetically Encoded ATP Sensor Based on Magnetic Resonance Imaging

Thursday, November 12, 2015: 1:50 PM
151A/B (Salt Palace Convention Center)
Arnab Mukherjee1, George Sun2, Xiaowei Zhang3, David V. Schaffer4 and Mikhail Shapiro3, (1)Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA, (2)University of California, Berkeley, Berkeley, CA, (3)California Institute of Technology, Pasadena, CA, (4)Department of Chemical and Biomolecular Engineering, University of California, Berkeley, CA

In this work, we report the development of the first genetically encoded sensor for noninvasive detection of adenosine triphosphate (ATP) using magnetic resonance imaging. Despite the critical relevance of ATP in cellular metabolism, neurotransmission, and as a clinical biomarker for neurodegenerative diseases and cerebrovascular injury, there is currently no method for noninvasive imaging of ATP in vivo. In particular, existing sensors for ATP imaging are based on fluorescence [1], which renders them unsuitable for imaging in preclinical animal models owing to the limited tissue penetration that is achievable using visible light. With a view towards enabling truly noninvasive molecular imaging of ATP, we developed an ATP sensor that is based on a bacterial glutamine synthetase. Glutamine synthetase produces potent MRI contrast owing to the presence of high spin manganese ions in the enzyme [2]. Here, we demonstrate that the contrast can be modulated by the binding of ATP to the enzyme, which serves as a molecular framework to develop an MRI-based contrast agent for ATP. We engineered glutamine synthetase to abrogate enzymatic activity while retaining ATP-dependent MRI contrast, thereby developing an enzymatically silent variant (known as GSATP) as a sensor for quantitative detection of ATP. We characterized relaxivities of GSATP in vitro at magnetic field strengths of 1 T and 7 T and  demonstrate an approximately 2-fold difference in relaxivities between the ATP-bound and ATP-free states of the sensor that results in appreciable ATP-dependent contrast on T1 and T2-weighted MR images. Finally, we demonstrated dynamic imaging of ATP using GSATP to monitor enzymatic degradation of ATP by apyrase. Overall, GSATP produces significantly stronger ligand-dependent contrast compared to existing genetically encoded MRI biosensors [3] and provides a platform for the development of other noninvasive MR sensors based on high spin manganese binding metalloproteins.       

References:

  1. Imamura, H. et al. PNAS, 106:15651-15656 (2009)
  2. Villafranca, J.J. et al. Biochemistry, 15:536-543 (1976)
  3. Shapiro, M.G. et al. Nature Biotechnology, 28: 264-270 (2010)

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