Exploring Protein’s Conformations in Vitro and in Cell with Gd(III) Spin Labeling - Joint EaStCHEM/BSRC Colloquium

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Abstract

Daniella Goldfarb
Weizmann Institute of Science
Distance measurements between two spin labels attached at well -defined sites in proteins by double electron -electron resonance (DEER) spectroscopy have become an established structural tool in Structural Biology. Such measurements are carried out on frozen solutions and provide information on conformational distribution s and conformational changes induced by ligand /substrate binding. The standard spin labels used for this purpose are based on nitroxide radicals. In recent years we, and others, have demonstr ated the utility of high frequency W-band, 95 GHz) DEER distance measurements in proteins using a new family of spin labels based on Gd(III) chelates. These are particularly useful due to the high sensitivity they have at high magnetic fields. The DEER per formance depends on the width of the EPR spectru m, which is determined by the Gd (III) zero field splitting (ZFS) that is a function of the chelate structure, and on the flexibility of the tether used to attach the Gd(III) chelate to the protein. We will p resent Gd(III) - Gd(III) DEER distance measurements on different protein systems using a variety of Gd(III) tags and will discuss their pros and cons. In addition , we will show that by combining different types of spin labels , like Gd(III), Mn(II) and a nit roxide in the same samples, we can significantly extend the structure information content derived from one sample. Finally we show that owing to the high sensitivity of Gd(III) -Gd(III) distances measurements and the stability of Gd 3+ chelates, in -cell distance measurements are within our reach. Examples of in -cell Gd(III) -Gd(III distance measurements on Gd Gd(III) -Gd(III labeled ubiquitin and calmodulin delivered into human HeLa cells using osmotic shock and electroporation will be described. Download PDF

  • Venue

    Theatre C, Purdie, University of St. Andrews, St. Andrews, Fife, Scotland, UK

  • Venue Info

  • Date

    May 2, 2018

  • Time

    From: 15h30 To: 16h30

  • Sponsor

    University of St Andrews
    The oldest university in Scotland, with international renown for both research and education of undergraduates and postgraduates.

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