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Related Concept Videos

Allosteric Regulation01:08

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Allosteric regulation of enzymes occurs when the binding of an effector molecule to a site that is different from the active site causes a change in the enzymatic activity. This alternate site is called an allosteric site, and an enzyme can contain more than one of these sites. Allosteric regulation can either be positive or negative, resulting in an increase or decrease in enzyme activity. Most enzymes that display allosteric regulation are metabolic enzymes involved in the degradation or...
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Cooperative allosteric transitions can occur in multimeric proteins, where each subunit of the protein has its own ligand-binding site. When a ligand binds to any of these subunits, it triggers a conformational change that affects the binding sites in the other subunits; this can change the affinity of the other sites for their respective ligands. The ability of the protein to change the shape of its binding site is attributed to the presence of a mix of flexible and stable segments in the...
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Binding sites linkages can regulate a protein's function.  For example, enzyme activity is often regulated through a feedback mechanism where the end product of the biochemical process serves as an inhibitor.
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When a ligand binds to a cell-surface receptor, the receptor's intracellular domain changes shape, which may either activate its enzyme function or allow its binding to other molecules. The initial signal is amplified by most signal transduction pathways. This means that a single ligand molecule can activate multiple molecules of a downstream target. Proteins that relay a signal are most commonly phosphorylated at one or more sites, activating or inactivating the protein. Kinases catalyze...
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Using Microtiter Dish Radiolabeling for Multiple In Vivo Measurements Of Escherichia coli pppGpp Followed by Thin Layer Chromatography
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(p)ppGpp controls stringent factors by exploiting antagonistic allosteric coupling between catalytic domains.

Mohammad Roghanian1, Katleen Van Nerom2, Hiraku Takada1

  • 1Department of Molecular Biology, Umeå University, 901 87, Umeå, Sweden; Laboratory for Molecular Infection Medicine Sweden, Umeå University, 901 87, Umeå, Sweden.

Molecular Cell
|August 20, 2021
PubMed
Summary

Bacterial cells sense amino acid starvation using RelA/Rel enzymes. The alarmone guanosine tetraphosphate (ppGpp) activates these enzymes by relieving autoinhibition, crucial for the starvation response.

Keywords:
(p)ppGppHydrogen-Deuterium exchange Mass SpectrometryRelRelARelA-SpoT HomologSpoTallosterypositive feedback regulationribosomestringent response

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Area of Science:

  • Molecular biology
  • Microbial stress response

Background:

  • RelA/Rel enzymes in E. coli and B. subtilis sense amino acid starvation via ribosomal A-site tRNA.
  • These enzymes synthesize the alarmone (p)ppGpp, which positively regulates their activity through an unknown mechanism.
  • Enzymatic activity is auto-inhibited by the N-terminal domain's hydrolase/pseudo-hydrolase domain.

Purpose of the Study:

  • To elucidate the mechanism of (p)ppGpp-mediated activation of RelA/Rel enzymes.
  • To identify the allosteric binding site for (p)ppGpp.
  • To understand the role of (p)ppGpp in coordinating the starvation response.

Main Methods:

  • Enzyme kinetics assays
  • Biochemical analysis of enzyme-ribosome interactions
  • In vivo functional studies in E. coli

Main Results:

  • The allosteric (p)ppGpp binding site was localized to the interface between the SYNTH and pseudo-HD/HD domains.
  • (p)ppGpp stimulates RelA/Rel activity by exploiting dynamics within the N-terminal domain's autoinhibition.
  • Starved ribosomes require (p)ppGpp for efficient activation of RelA/Rel.
  • SpoT, a related enzyme in E. coli, lacks (p)ppGpp control, explaining its weak activity.

Conclusions:

  • Activation of RelA/Rel by (p)ppGpp is essential for the acute starvation response in bacteria.
  • The second messenger (p)ppGpp coordinates enzymatic activity for effective stress adaptation.
  • (p)ppGpp regulation is absent in SpoT, differentiating its function from RelA/Rel.