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Related Concept Videos

Protein Diffusion in the Membrane01:24

Protein Diffusion in the Membrane

Proteins show rotational as well as lateral diffusion across the membrane. The lateral diffusion of proteins was confirmed through the cell fusion experiment where mouse and human cells were fused, resulting in hybrid cells. When the human and mouse cells fused, the specific membrane proteins on human and mouse cells were marked with the red and green-fluorescent markers, respectively. Initially, the red and green fluorescence was located on the respective hemisphere of the cell. As time...
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Neutron Spin Echo Spectroscopy as a Unique Probe for Lipid Membrane Dynamics and Membrane-Protein Interactions
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Evolving Accelerated Amidation by SpyTag/SpyCatcher to Analyze Membrane Dynamics.

Anthony H Keeble1, Anusuya Banerjee1, Matteo P Ferla1

  • 1Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, UK.

Angewandte Chemie (International Ed. in English)
|October 13, 2017
PubMed
Summary

Researchers enhanced SpyTag and SpyCatcher protein interactions using a phage-display platform. This accelerated covalent protein assembly for advanced molecular engineering and live cell imaging applications.

Keywords:
SpyTag/SpyCatchermembrane proteinsprotein engineeringprotein-protein interactionssynthetic biology

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Synthetic Biology

Background:

  • SpyTag and SpyCatcher form a covalent bond, acting as a protein superglue for molecular assembly.
  • Initial SpyTag/SpyCatcher system developed via rational design from a bacterial adhesin.

Purpose of the Study:

  • To accelerate the SpyTag-SpyCatcher interaction kinetics.
  • To develop an optimized protein ligation system for enhanced molecular assembly and biological applications.

Main Methods:

  • Utilized a phage-display platform to select for improved SpyTag/SpyCatcher variants.
  • Engineered SpyTag002 and SpyCatcher002 variants with significantly faster binding kinetics.
  • Fused SpyCatcher002 to bacterial intimin for targeted labeling.

Main Results:

  • Achieved an order of magnitude acceleration in SpyTag002/SpyCatcher002 interaction.
  • Demonstrated rapid and robust covalent bonding under diverse conditions and at protein termini.
  • Successfully visualized bacterial outer membrane dynamics using intimin-decorated SpyTag002 for live cell imaging.

Conclusions:

  • The optimized SpyTag002/SpyCatcher002 system offers rapid and versatile covalent protein ligation.
  • This enhanced system facilitates advanced molecular assembly and precise live cell imaging of bacterial processes.