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Total Chemical Synthesis of an Integral Membrane Enzyme: Diacylglycerol Kinase from Escherichia coli

Authors

  • Dr. Sunanda Lahiri,

    1. Max Planck Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund (Germany)
    2. Current address: Universität Freiburg, Institute of Physiology 2, Engesserstrasse 4, 79108 Freiburg (Germany)
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  • Manuel Brehs,

    1. Center of Integrated Protein Science and Technische Universität München, Protein Chemistry, Lichtenbergstrasse 4, 85747 Garching (Germany), Fax: (+49) 89-289-13345 http://www.ch.tum.de/proteinchemie
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  • Dr. Diana Olschewski,

    1. Max Planck Institute of Molecular Physiology, Otto-Hahn-Strasse 11, 44227 Dortmund (Germany)
    2. Current address: Lonza Ltd., 3930 Visp (Switzerland)
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  • Prof. Dr. Christian F. W. Becker

    Corresponding author
    1. Center of Integrated Protein Science and Technische Universität München, Protein Chemistry, Lichtenbergstrasse 4, 85747 Garching (Germany), Fax: (+49) 89-289-13345 http://www.ch.tum.de/proteinchemie
    • Center of Integrated Protein Science and Technische Universität München, Protein Chemistry, Lichtenbergstrasse 4, 85747 Garching (Germany), Fax: (+49) 89-289-13345 http://www.ch.tum.de/proteinchemie===

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  • We thank Katja Bäuml and Sascha Gentz for excellent technical assistance with peptide synthesis. We also thank Francis Lau for technical advice on establishing the DAGK activity assay. Financial support from the DFG is gratefully acknowledged.

Abstract

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From the ground up: Chemical synthesis provided direct access to a catalytically active membrane-embedded kinase with three transmembrane domains (TMs). Three key segments formed from the individual amino acids (represented by red and gray balls) by solid-phase peptide synthesis were connected by native chemical ligation (NCL; see picture); the synthetic protein underwent spontaneous folding in detergent micelles. ATP=adenosine 5′-triphosphate.

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