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Structure-Guided Directed Evolution of Highly Selective P450-Based Magnetic Resonance Imaging Sensors for Dopamine and Serotonin

Brustad, Eric M. and Lelyveld, Victor S. and Snow, Christopher D. and Crook, Nathan C. and Jung, Sang Taek and Martinez, Francisco M. and Scholl, Tomothy J. and Jasanoff, Alan and Arnold, Frances H. (2012) Structure-Guided Directed Evolution of Highly Selective P450-Based Magnetic Resonance Imaging Sensors for Dopamine and Serotonin. Journal of Molecular Biology, 422 (2). pp. 245-262. ISSN 0022-2836. PMCID PMC3418479. https://resolver.caltech.edu/CaltechAUTHORS:20121030-144039360

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Abstract

New tools that allow dynamic visualization of molecular neural events are important for studying the basis of brain activity and disease. Sensors that permit ligand-sensitive magnetic resonance imaging (MRI) are useful reagents due to the noninvasive nature and good temporal and spatial resolution of MR methods. Paramagnetic metalloproteins can be effective MRI sensors due to the selectivity imparted by the protein active site and the ability to tune protein properties using techniques such as directed evolution. Here, we show that structure-guided directed evolution of the active site of the cytochrome P450‐BM3 heme domain produces highly selective MRI probes with submicromolar affinities for small molecules. We report a new, high‐affinity dopamine sensor as well as the first MRI reporter for serotonin, with which we demonstrate quantification of neurotransmitter release in vitro. We also present a detailed structural analysis of evolved cytochrome P450‐BM3 heme domain lineages to systematically dissect the molecular basis of neurotransmitter binding affinity, selectivity, and enhanced MRI contrast activity in these engineered proteins.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1016/j.jmb.2012.05.029DOIArticle
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3418479PubMed CentralArticle
ORCID:
AuthorORCID
Brustad, Eric M.0000-0002-7031-5433
Snow, Christopher D.0000-0002-7690-3519
Crook, Nathan C.0000-0001-6165-1972
Jasanoff, Alan0000-0002-2834-6359
Arnold, Frances H.0000-0002-4027-364X
Additional Information:© 2012 Elsevier Ltd. Received 29 February 2012; received in revised form 16 May 2012; accepted 18 May 2012. Available online 30 May 2012. Edited by D. Tawfik. We thank Fay Bi for technical assistance and P. Romero, M. F. Farrow, M. Smith, R. Lauchli, and P. Coehlo for helpful discussions during the preparation of the manuscript. We thank M. Shapiro for initiating work with BM3h-based contrast agents and for helpful discussions. We thank J. Kaiser, P. Nickolovski, J. Hoy, and the Molecular Observatory at the California Institute of Technology for assistance with high-throughput protein crystallography, X-ray data collection, and analysis. The Molecular Observatory is graciously supported by the Gordon and Betty Moore Foundation, the Beckman Institute, and the Sanofi-Aventis Bioengineering Research Program at Caltech. E.M.B. is supported by a Ruth M. Kirschstein National Institutes of Health (NIH) postdoctoral fellowship Award Number F32GM087102 from the National Institute of General Medical Sciences. We acknowledge support from the Jacobs Institute for Molecular Engineering for Medicine to F.H.A. This publication was made possible by grant 1R01DA028299‐01 from the NIH to A.J. and F.H.A. The content is solely the responsibility of the authors and does not necessarily represent the official view of the NIH.
Group:Jacobs Institute for Molecular Engineering for Medicine
Funders:
Funding AgencyGrant Number
Gordon and Betty Moore FoundationUNSPECIFIED
Caltech Beckman InstituteUNSPECIFIED
Caltech Sanofi-Aventis Bioengineering Research ProgramUNSPECIFIED
NIH Predocotral FellowshipF32GM087102
Jacobs Institute for Molecular Engineering for MedicineUNSPECIFIED
NIH1R01DA028299‐01
Subject Keywords: directed evolution; cytochrome P450; MRI contrast agents; neurotransmitter; biosensors
Issue or Number:2
PubMed Central ID:PMC3418479
Record Number:CaltechAUTHORS:20121030-144039360
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20121030-144039360
Official Citation:Eric M. Brustad, Victor S. Lelyveld, Christopher D. Snow, Nathan Crook, Sang Taek Jung, Francisco M. Martinez, Timothy J. Scholl, Alan Jasanoff, Frances H. Arnold, Structure-Guided Directed Evolution of Highly Selective P450-Based Magnetic Resonance Imaging Sensors for Dopamine and Serotonin, Journal of Molecular Biology, Volume 422, Issue 2, 14 September 2012, Pages 245-262, ISSN 0022-2836, 10.1016/j.jmb.2012.05.029. (http://www.sciencedirect.com/science/article/pii/S0022283612004275)
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:35185
Collection:CaltechAUTHORS
Deposited By: Ruth Sustaita
Deposited On:30 Oct 2012 22:11
Last Modified:15 Jun 2020 18:41

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