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Related Concept Videos

SNAREs and Membrane Fusion01:43

SNAREs and Membrane Fusion

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Once a transport vesicle has recognized its target organelle, the vesicular membrane needs to fuse with the target membrane to unload the cargo. Transmembrane proteins called SNAREs present on organelle membranes and their vesicles, mediate vesicle fusion.
SNAREs exist in pairs that symmetrically interact and catalyze the fusion of the lipid bilayers in vesicle and target organelle. v-SNARE in the vesicle membrane are single polypeptide chains that bind to a complementary t-SNARE, composed of 2...
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Fusion of Secretory Vesicles with the Plasma Membrane01:26

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Proteins and neurotransmitters in secretory vesicles can be released from a cell upon vesicle docking, priming, and fusion with the plasma membrane. Vesicles are docked and primed in preparation for the quick exocytosis of their contents in response to a stimulus. The fusion process is mainly carried out by a SNAP Receptor or SNARE complex, consisting of synaptobrevin, syntaxin-1, and SNAP-25.
In 1993, Jim Rothman proposed that the antiparallel pairing of vesicular and transmembrane SNAREs, or...
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Related Experiment Video

Updated: May 10, 2025

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Stabilizing Prefusion SARS-CoV-2 Spike by Destabilizing the Postfusion Conformation.

Debajyoti Chakraborty1, Randhir Singh2, Raju S Rajmani1

  • 1Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560012, India.

Vaccines
|April 23, 2025
PubMed
Summary

Introducing charged amino acids into SARS-CoV-2 Spike protein enhanced its expression and stability. This novel approach, destabilizing the postfusion form, offers a general strategy for improving viral surface protein yields for vaccines.

Keywords:
COVID-19HR1 and HR2SARS-CoV-2 Spikecharged amino acidsimmunogenicitystabilization

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Area of Science:

  • Structural biology
  • Vaccine development
  • Virology

Background:

  • The native SARS-CoV-2 Spike protein is metastable.
  • Current COVID-19 vaccines use stabilized Spike variants (Spike-2P, Spike-6P) with proline substitutions.
  • An alternative strategy involves destabilizing the postfusion conformation.

Purpose of the Study:

  • To investigate the effect of introducing two aspartic acid residues (2D) into the HR1 region of the Spike protein.
  • To enhance the yield and stability of the prefusion Spike conformation.
  • To evaluate the immunogenicity and protective efficacy of modified Spike variants.

Main Methods:

  • Recombinant protein expression in mammalian cell culture.
  • Characterization of protein yield and antigenicity.
  • Immunization of hamsters followed by challenge with live SARS-CoV-2 B.1.351 variant.

Main Results:

  • The 2D mutations increased protein expression six-fold.
  • Combining 2D with four proline mutations (Spike-4P-2D) further enhanced expression.
  • All tested Spike variants (2P, 2D, 6P, 4P-2D) protected hamsters against SARS-CoV-2 challenge and induced high neutralizing antibody titers.

Conclusions:

  • Destabilizing the postfusion conformation by introducing charged amino acids enhances viral surface protein yield and stability.
  • This strategy offers a general approach for improving viral protein expression for vaccine development.
  • The 2D mutation strategy shows promise for developing more effective vaccines.