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Related Concept Videos

Protein Kinases and Phosphatases02:54

Protein Kinases and Phosphatases

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Proteins undergo chemical modifications that trigger changes in the charge, structure, and conformation of the proteins. Phosphorylation, acetylation, glycosylation, nitrosylation, ubiquitination, lipidation, methylation, and proteolysis are various protein modifications that regulate protein activity. Such modifications are usually enzyme-driven.
Protein kinases
Many proteins in the cell are regulated by phosphorylation, the addition of a phosphate group. A family of enzymes called kinases...
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Phosphorylation01:02

Phosphorylation

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The addition or removal of phosphate groups from proteins is the most common chemical modification that regulates cellular processes. These modifications can affect the structure, activity, stability, and localization of proteins within cells as well as their interactions with other proteins.
During phosphorylation, protein kinases transfer the terminal phosphate group of ATP to specific amino acid side chains of substrate proteins. Serine, threonine, and tyrosine are the most commonly...
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Amplifying Signals via Enzymatic Cascade01:22

Amplifying Signals via Enzymatic Cascade

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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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Cooperative Allosteric Transitions01:58

Cooperative Allosteric Transitions

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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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Covalently Linked Protein Regulators02:04

Covalently Linked Protein Regulators

6.8K
Proteins can undergo many types of post-translational modifications, often in response to changes in their environment. These modifications play an important role in the function and stability of these proteins. Covalently linked molecules include functional groups, such as methyl, acetyl, and phosphate groups, and also small proteins, such as ubiquitin. There are around 200 different types of covalent regulators that have been identified.
These groups modify specific amino acids in a protein....
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Protein-protein Interfaces02:04

Protein-protein Interfaces

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Many proteins form complexes to carry out their functions, making protein-protein interactions (PPIs) essential for an organism's survival. Most PPIs are stabilized by numerous weak noncovalent chemical forces. The physical shape of the interfaces determines the way two proteins interact. Many globular proteins have closely-matching shapes on their surfaces, which form a large number of weak bonds. Additionally, many PPIs occur between two helices or between a surface cleft and a...
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Updated: Jun 24, 2025

Identification of Kinase-substrate Pairs Using High Throughput Screening
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Protein Interaction Kinetics Delimit the Performance of Phosphorylation-Driven Protein Switches.

Daniel L Winter1,2, Adelgisa R Wairara1, Jack L Bennett3

  • 1School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, NSW 2052, Australia.

ACS Synthetic Biology
|June 3, 2024
PubMed
Summary

We developed a computational framework and novel protein switches that respond to phosphorylation. These engineered protein switches offer fast, reversible control and can be used as biosensors for real-time kinase activity measurements.

Keywords:
binding kineticsbiosensorcoiled coilphosphorylationprotein kinaseprotein switch

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Systems Biology

Background:

  • Post-translational modifications (PTMs) rapidly alter protein function by changing surface chemistry and structure, controlling protein-protein interactions (PPIs) in signaling networks.
  • Engineered protein switches responsive to phosphorylation offer potential for studying PTM dynamics, creating nanodevices, and programming cellular behavior.
  • Understanding the physical and kinetic limitations of PTM-driven protein switches is crucial for their practical application.

Purpose of the Study:

  • To develop a framework for evaluating two-component, post-translational modification-driven protein switches.
  • To investigate the relationship between binding kinetics, phosphorylation kinetics, and switch concentration.
  • To design and validate novel phosphorylation-driven protein switches with controllable and reversible function.

Main Methods:

  • Computational modeling to analyze performance metrics: effective concentration, dynamic range, response time, and reversibility.
  • Design of novel protein switches using phosphorylation-sensitive coiled coils fused to fluorescent proteins.
  • In vitro evaluation of switch function using specific protein kinase and phosphatase to modulate phosphorylation states.

Main Results:

  • Computational models revealed intricate relationships governing switch sensitivity and reversibility based on binding and phosphorylation kinetics.
  • Engineered protein switches demonstrated fast, reversible transitions between 'on' and 'off' states modulated by kinase and phosphatase activity.
  • Switch response showed a linear correlation with kinase concentration, indicating potential for real-time kinase biosensing.

Conclusions:

  • The developed framework and computational models provide insights into the design principles of PTM-driven protein switches.
  • Novel phosphorylation-driven protein switches exhibit tunable and reversible control, enabling applications in biosensing and synthetic biology.
  • This work lays the foundation for designing PTM-driven switches with optimized performance for specific biological applications.