Related Experiment Video
Updated: Jul 19, 2026
13:30
A New Screening Method for the Directed Evolution of Thermostable Bacteriolytic Enzymes
Published on: November 7, 2012
Energetic landscape of alpha-lytic protease optimizes longevity through kinetic stability
Sheila S Jaswal1, Julie L Sohl, Jonathan H Davis
1Department of Biochemistry and Biophysics, University of California at San Francisco, 94143-0448, USA.
Nature
|January 18, 2002
Summary
Alpha-lytic protease (alphaLP) evolution decoupled protein folding and stability. A pro-region facilitates folding, creating a stable native state with reduced fluctuations, extending functional lifetime.
Area of Science:
- Biochemistry
- Protein Folding
- Evolutionary Biology
Background:
- Protein folding is crucial for biological activity, typically yielding stable native states.
- Alpha-lytic protease (alphaLP) exhibits decoupled folding and stability, with an unstable native state.
- alphaLP refolding takes millennia without a catalyst, while its native state is kinetically stable.
Purpose of the Study:
- To investigate the evolutionary decoupling of folding and stability in alphaLP.
- To understand how the pro-region influences alphaLP folding and stability.
- To explore the impact of separated folding/unfolding pathways on protein dynamics and function.
Main Methods:
- Comparative analysis of alphaLP folding landscapes with and without the pro-region.
- Characterization of kinetic and thermodynamic properties of alphaLP folding and unfolding.
- Assessment of dynamic fluctuations in the native state of alphaLP.
Main Results:
- The pro-region acts as a folding catalyst, enabling the formation of a thermodynamically unstable native state.
- Separation of folding and unfolding pathways allows for reduced dynamic fluctuations in the alphaLP native state.
- This reduction in dynamics enhances alphaLP's functional lifetime by minimizing proteolytic degradation.
Conclusions:
- Decoupling folding from thermodynamic stability in alphaLP enabled evolutionary optimization of its native state.
- The pro-region's role in facilitating folding and subsequent degradation is key to alphaLP's unique properties.
- Reduced protein dynamics in alphaLP's native state contribute to its extended functional longevity and resistance to proteolysis.
Related Concept Videos
01:30Lytic Cycle of Bacteriophages
Bacteriophages, also known as phages, are specialized viruses that infect bacteria. A key characteristic of phages is their distinctive “head-tail” morphology. A phage begins the infection process (i.e., lytic cycle) by attaching to the outside of a bacterial cell. Attachment is accomplished via proteins in the phage tail that bind to specific receptor proteins on the outer surface of the bacterium. The tail injects the phage’s DNA genome into the bacterial cytoplasm. In the lytic replication...
01:19Enzyme Kinetics
Enzymes speed up reactions by lowering the activation energy of the reactants. The speed at which the enzyme turns reactants into products is called the rate of reaction. Several factors impact the rate of reaction, including the number of available reactants. Enzyme kinetics is the study of how an enzyme changes the rate of a reaction.
Scientists typically study enzyme kinetics with a fixed amount of enzyme in the controlled environment of a test tube. When more reactant, or substrate, is...
Scientists typically study enzyme kinetics with a fixed amount of enzyme in the controlled environment of a test tube. When more reactant, or substrate, is...
02:18The Proteasome
Eukaryotic cells can degrade proteins through several pathways. One of the most important amongst these is the ubiquitin-proteasome pathway. It helps the cell eliminate the misfolded, damaged, or unwarranted cytoplasmic proteins in a highly specific manner.
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. A series of enzymes carry out the ubiquitination of the target proteins - E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3...
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. A series of enzymes carry out the ubiquitination of the target proteins - E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3...
02:18The Proteasome
Eukaryotic cells can degrade proteins through several pathways. One of the most important amongst these is the ubiquitin-proteasome pathway. It helps the cell eliminate the misfolded, damaged, or unwarranted cytoplasmic proteins in a highly specific manner.
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. A series of enzymes carry out the ubiquitination of the target proteins - E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3...
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. A series of enzymes carry out the ubiquitination of the target proteins - E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3...
01:19Protein Dynamics in Living Cells
Different fluorescence-based techniques are used to study the protein dynamics in living cells. These techniques include FRAP, FRET, and PET.
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...
01:13The Proteasome
Eukaryotic cells can degrade proteins through several pathways. One of the most important among these is the ubiquitin-proteasome pathway. It helps the cell eliminate the misfolded, damaged, or unwarranted cytoplasmic proteins in a highly specific manner.
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. This involves participation of a series of enzymes including— E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3 (ubiquitin...
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. This involves participation of a series of enzymes including— E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3 (ubiquitin...