A Decoupled Automation Platform for Hydrogen/Deuterium Exchange Mass Spectrometry Experiments
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J. Am. Soc. Mass Spectrom. (2019) DOI: 10.1007/s13361-019-02331-2
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A Decoupled Automation Platform for Hydrogen/Deuterium Exchange Mass Spectrometry Experiments Alfonso Espada,1 Ruben Haro,1 Jesus Castañon,1 Cristina Sayago,1 Francisco Perez-Cozar,1 Leticia Cano,1 Pablo Redero,1 Manuel Molina-Martin,1 Howard Broughton,1 Ryan E. Stites,2 Bruce D. Pascal,3 Patrick R. Griffin,4 Jeffrey A. Dodge,2 Michael J. Chalmers2 1
Centro de Investigación Lilly, SA, Avenida de la Industria 30, 28108, Alcobendas, Spain Eli Lilly and Company, Lilly Corporate Center, Lilly Research Laboratories, Indianapolis, IN 46285, USA 3 Omics Informatics LLC, 1050 Bishop Street #517, Honolulu, HI 96813, USA 4 Department of Molecular Medicine, The Scripps Research Institute, 130 Scripps Way, Jupiter, FL 33458, USA 2
Abstract. Hydrogen/deuterium exchange mass spectrometry (HDX-MS) is a biophysical technique well suited to the characterization of protein dynamics and protein–ligand interactions. In order to accurately define the rate of exchange, HDX experiments require the repeated measure of deuterium incorporation into the target protein across a range of time points. Accordingly, the HDX-MS experiment is well suited to automation, and a number of automated systems for HDXMS have been developed. The most widely utilized platforms all operate an integrated design, where robotic liquid handling is interfaced directly with a mass spectrometer. With integrated designs, the exchange samples are prepared and injected into the LC-MS following a “real-time” serial workflow. Here we describe a new HDX-MS platform that is comprised of two complementary pieces of automation that disconnect the sample preparation from the LC-MS analysis. For preparation, a plate-based automation system is used to prepare samples in parallel, followed by immediate freezing and storage. A second piece of automation has been constructed to perform the thawing and LC-MS analysis of frozen samples in a serial mode and has been optimized to maximize the duty cycle of the mass spectrometer. The decoupled configuration described here reduces experiment time, significantly improves capacity, and improves the flexibility of the platform when compared with a fully integrated system. Keywords: HDX, Hydrogen deuterium exchange, Automation, Protein–ligand interaction Received: 10 June 2019/Revised: 19 August 2019/Accepted: 21 August 2019
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Alfonso Espada and Ruben Haro contributed equally to this work. Electronic supplementary material The online version of this article (https:// doi.org/10.1007/s13361-019-02331-2) contains supplementary material, which is available to authorized users. Correspondence to: Michael Chalmers; e-mail: [email protected]
mall molecule drug discovery projects often rely heavily on the application of biophysical techniques to characterize protein–ligand interactions [1]. It is frequently the case that multiple methods are required to adequately profile the interaction for any single target of interest, with many of t
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