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Related Concept Videos

Conserved Binding Sites01:49

Conserved Binding Sites

Many proteins’ biological role depends on their interactions with their ligands, small molecules that bind to specific locations on the protein known as ligand-binding sites. Ligand-binding sites are often conserved among homologous proteins as these sites are critical for protein function.
Binding sites are often located in large pockets, and if their location on a protein’s surface is unknown, it can be predicted using various approaches. The energetic method computationally analyses the...
Protein-protein Interfaces02:04

Protein-protein Interfaces

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 polypeptide...
Assembly of Signaling Complexes01:30

Assembly of Signaling Complexes

Multiprotein signaling complexes are formed in a dynamic process involving protein-protein interactions at the cytoplasmic domain of transmembrane receptors or enzymatic and non-enzymatic proteins associated with the receptor. These complexes ensure the activation and propagation of intracellular signals that regulate cell functions.
Interaction domains in cell signaling
Interaction domains recognize exposed features of their binding partners containing post-translationally modified sequences,...
Ligand Binding Sites02:40

Ligand Binding Sites

Proteins are dynamic macromolecules that carry out a wide variety of essential processes; however, the activities of most proteins depend on their interactions with other molecules or ions, known as ligands.
Protein-ligand interactions are quite specific; even though numerous potential ligands surround a cellular protein at any given time, only a particular ligand can bind to that protein. Moreover, a ligand binds only to a dedicated area on the surface of the protein, known as the...
The Equilibrium Binding Constant and Binding Strength02:18

The Equilibrium Binding Constant and Binding Strength

The equilibrium binding constant (Kb) quantifies the strength of a protein-ligand interaction. Kb can be calculated as follows when the reaction is at equilibrium:
Ligand Binding and Linkage00:49

Ligand Binding and Linkage

Allosteric proteins have more than one ligand binding site; the binding of a ligand to any of these sites influences the binding of ligands to the other sites. When a protein is allosteric, its binding sites are called coupled or linked.  In the case of enzymes, the site that binds to the substrate is known as the active site and the other site is known as the regulatory site. When a ligand binds to the regulatory site, this leads to conformational changes in the protein that can influence the...

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Protein WISDOM: A Workbench for In silico De novo Design of BioMolecules
10:58

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Published on: July 25, 2013

Computational design of a PAK1 binding protein.

Ramesh K Jha1, Andrew Leaver-Fay, Shuangye Yin

  • 1Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7260, USA.

Journal of Molecular Biology
|May 13, 2010
PubMed
Summary

We developed DDMI, a computational protocol for protein redesign, enabling scaffolds to bind specific target protein regions. This method successfully created a novel protein binder, Spider Roll, with a 100 muM affinity for PAK1 kinase.

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Published on: January 26, 2024

Area of Science:

  • Computational biology
  • Protein engineering
  • Structural biology

Background:

  • Designing novel protein-protein interactions is crucial for developing therapeutics and molecular probes.
  • Existing methods for designing specific protein binders are limited.

Purpose of the Study:

  • To develop and validate a computational protocol (DDMI) for redesigning scaffold proteins to bind specific target protein regions.
  • To experimentally test the DDMI protocol's efficacy by designing a binder for p21-activated kinase 1 (PAK1).

Main Methods:

  • DDMI protocol implemented in Rosetta molecular modeling software.
  • Utilized rigid-body docking, sequence design, and gradient-based minimization.
  • Iterative rounds of sequence design and conformational optimization, including molecular dynamics simulations.
  • Experimental characterization using binding affinity assays, NMR-based structure prediction, and mutagenesis studies.

Main Results:

  • The DDMI protocol generated models with calculated binding energies comparable to native complexes.
  • Three out of four experimentally tested designs bound PAK1 with affinities tighter than 350 muM.
  • The tightest binder, Spider Roll, exhibited a 100 muM affinity for PAK1.
  • Spider Roll demonstrated specific binding to the activated form of PAK1 and preserved the scaffold's architecture.

Conclusions:

  • The DDMI protocol is effective for rationally designing novel protein-protein interactions.
  • Designed proteins can achieve significant binding affinities to target proteins.
  • This approach holds promise for creating new protein-based therapeutics and research tools.