Screening libraries for improved solubility: using E. coli dihydrofolate reductase as a reporter.
1CSIRO Ecosystem Sciences, Canberra, Australia.
Methods in Molecular Biology (Clifton, N.J.)
|February 21, 2013
Summary
Evolving proteins for better solubility is challenging. This study introduces a novel method using dihydrofolate reductase (DHFR) fusions to select more soluble mutant proteins based on altered resistance to Trimethoprim (TMP).
Area of Science:
- Protein Science
- Molecular Biology
- Biotechnology
Background:
- Low protein solubility hinders protein science applications.
- Current chemical solubilization methods are often ineffective and costly.
- Protein evolution offers a promising alternative for enhancing solubility.
Purpose of the Study:
- To develop a method for selecting mutant proteins with improved solubility from large libraries.
- To overcome the hurdle of identifying soluble variants in protein engineering.
Main Methods:
- Utilized a selection strategy based on dihydrofolate reductase (DHFR) expression in Escherichia coli.
- Created protein chimeras by fusing target proteins to DHFR.
- Leveraged altered resistance to Trimethoprim (TMP) as a readout for protein solubility.
Main Results:
- Demonstrated that DHFR fusion chimeras exhibit varying levels of TMP resistance.
- Showed that TMP resistance correlates with the solubility of the fused protein.
Conclusions:
- The described method provides an effective way to select for soluble mutant proteins.
- This approach facilitates protein engineering efforts aimed at improving protein solubility.
Related Concept Videos
04:01Factors Affecting Solubility
37.2K
Compared with pure water, the solubility of an ionic compound is less in aqueous solutions containing a common ion (one also produced by dissolution of the ionic compound). This is an example of a phenomenon known as the common ion effect, which is a consequence of the law of mass action that may be explained using Le Chȃtelier’s principle. Consider the dissolution of silver iodide:
37.2K
03:07Solubility Equilibria
57.6K
Solubility equilibria are established when the dissolution and precipitation of a solute species occur at equal rates. These equilibria underlie many natural and technological processes, ranging from tooth decay to water purification. An understanding of the factors affecting compound solubility is, therefore, essential to the effective management of these processes. This section applies previously introduced equilibrium concepts and tools to systems involving dissolution and precipitation.
The...
The...
57.6K
02:55Solubility of Ionic Compounds
68.3K
Solubility is the measure of the maximum amount of solute that can be dissolved in a given quantity of solvent at a given temperature and pressure. Solubility is usually measured in molarity (M) or moles per liter (mol/L). A compound is termed soluble if it dissolves in water.
68.3K
02:19Physical Properties Affecting Solubility
26.8K
Solutions of Gases in Liquids
As for any solution, the solubility of a gas in a liquid is affected by the attractive intermolecular forces between solute and solvent species. Unlike solid and liquid solutes, however, there is no solute-solute intermolecular attraction to overcome when a gaseous solute dissolves in a liquid solvent since the atoms or molecules comprising a gas are far separated and experience negligible interactions. Consequently, solute-solvent interactions are the sole...
As for any solution, the solubility of a gas in a liquid is affected by the attractive intermolecular forces between solute and solvent species. Unlike solid and liquid solutes, however, there is no solute-solute intermolecular attraction to overcome when a gaseous solute dissolves in a liquid solvent since the atoms or molecules comprising a gas are far separated and experience negligible interactions. Consequently, solute-solvent interactions are the sole...
26.8K
03:00Solubility
21.2K
Solution, Solubility, and Solubility Equilibrium
A solution is a homogeneous mixture composed of a solvent, the major component, and a solute, the minor component. The physical state of a solution—solid, liquid, or gas—is typically the same as that of the solvent. Solute concentrations are often described with qualitative terms such as dilute (of relatively low concentration) and concentrated (of relatively high concentration).
In a solution, the solute particles (molecules,...
A solution is a homogeneous mixture composed of a solvent, the major component, and a solute, the minor component. The physical state of a solution—solid, liquid, or gas—is typically the same as that of the solvent. Solute concentrations are often described with qualitative terms such as dilute (of relatively low concentration) and concentrated (of relatively high concentration).
In a solution, the solute particles (molecules,...
21.2K
01:24Data Reporting and Recording
5.5K
Reporting and recording are crucial in data documentation. The timely, thorough, and accurate documentation of facts is essential when recording patient data. Failure to record findings during an assessment or interpretation of a problem will result in loss of information and make the patient document unreliable. The reader is left with general impressions if the information is not specific. A recording is documenting data of the individual's health information in a traceable, secure, and...
5.5K