Related Experiment Video

Updated: May 27, 2025

Combining Non-reducing SDS-PAGE Analysis and Chemical Crosslinking to Detect Multimeric Complexes Stabilized by Disulfide Linkages in Mammalian Cells in Culture
09:37

Combining Non-reducing SDS-PAGE Analysis and Chemical Crosslinking to Detect Multimeric Complexes Stabilized by Disulfide Linkages in Mammalian Cells in Culture

Published on: May 2, 2019

10.0K

Disulfide bonds are required for cell division, cell envelope biogenesis and antibiotic resistance proteins in

Adrian Mejia-Santana1, Rebecca Collins1, Emma H Doud2,3

  • 1Department of Biology. Indiana University. Bloomington, IN. U.S.A.

Biorxiv : the Preprint Server for Biology
|February 20, 2025
PubMed

Insights

Targeting disulfide bond (DSB) formation in mycobacteria, crucial for cell envelope integrity, offers a novel strategy against tuberculosis. Inhibiting the DsbA-VKOR pathway disrupts cell division and mycomembrane stability, showing promise for combating drug-resistant strains.

Area of Science:

  • Microbiology
  • Molecular Biology
  • Biochemistry

Background:

  • Mycobacteria possess a unique cell envelope essential for survival and resistance.
  • Disulfide bond (DSB) formation is critical for proper folding of cell envelope proteins.
  • The DSB pathway, involving DsbA and VKOR enzymes, is vital for mycobacterial survival.

Purpose of the Study:

  • To identify cell envelope proteins reliant on DSB formation in mycobacteria.
  • To investigate the role of the DsbA-VKOR pathway in mycobacterial cell envelope maintenance.
  • To explore the therapeutic potential of inhibiting the DSB pathway against mycobacterial infections.

Main Methods:

  • Bioinformatics analysis and cysteine profiling proteomics to identify DSB-dependent proteins.
  • In vivo alkylation assays to validate protein stability.
  • Chemical inhibition of VKOR to assess phenotypic consequences.

Main Results:

  • Identified key cell envelope proteins, including LamA (MmpS3), PstP, LpqW, and EmbB, that depend on DSBs for stability.
  • Demonstrated that chemical inhibition of VKOR mimics the effects of its genetic deletion.
  • Showed that targeting the DsbA-VKOR system impacts both cell division and mycomembrane integrity.

Conclusions:

  • The DsbA-VKOR pathway is essential for mycobacterial cell envelope integrity and function.
  • Inhibition of DSB formation presents a promising novel therapeutic strategy against mycobacterial infections, including multidrug-resistant tuberculosis.
  • Targeting this pathway could compromise essential cellular processes, leading to bacterial death.
Keywords:
AftBAftDDsbAEccB3EmbBLamALpqWMmpS3MycP3PP2CPstPRv2507Ser/Thr phosphataseVKORactinobacteriadisulfide bondsessential proteinsmycobacteriamycomembraneoxidative protein foldingsubstrates

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