Diselenide crosslinks for enhanced and simplified oxidative protein folding
Reem Mousa1, Taghreed Hidmi1, Sergei Pomyalov1
1Institute of Chemistry, The Hebrew University of Jerusalem, Jerusalem, Israel.
Communications Chemistry
|January 25, 2023
Summary
Diselenide bridges enhance protein folding rates and yields in vitro. This study shows diselenide substitutions in hirudin improve folding efficiency and structural integrity for disulfide-rich proteins.
Area of Science:
- Biochemistry
- Structural Biology
- Protein Chemistry
Background:
- Protein folding is crucial for biological function, with disulfide bonds playing a key role.
- Hirudin, a thrombin inhibitor, serves as a model for disulfide-rich proteins but exhibits complex folding pathways.
- Existing methods for in vitro oxidative folding of disulfide-rich proteins face challenges with efficiency and heterogeneity.
Purpose of the Study:
- To investigate the impact of diselenide bridges on the in vitro folding of hirudin.
- To assess how diselenide substitutions affect the folding rate, yield, structural integrity, and activity of hirudin analogues.
- To explore the potential of diselenide bridges in improving the preparation of disulfide-rich proteins.
Main Methods:
- Systematic substitution of native disulfide bonds in hirudin with diselenide bridges.
- In vitro oxidative folding experiments to assess folding kinetics and yields.
- Crystal structure analysis to determine the structural impact of diselenide substitutions.
- Functional assays to evaluate the activity of hirudin analogues.
Main Results:
- Diselenide crosslinks significantly enhanced the folding rate and yield of hirudin analogues.
- The complexity and heterogeneity of folding intermediates were reduced by diselenide substitutions.
- Crystal structures confirmed that diselenide substitutions preserved the overall three-dimensional protein structure.
- Functional activity of the diselenide-containing hirudin analogues remained largely unchanged.
Conclusions:
- Diselenide substitutions are a promising strategy to overcome limitations in the in vitro folding of disulfide-rich proteins like hirudin.
- This approach offers a method to improve the efficiency and homogeneity of protein folding processes.
- The findings have broad implications for the design, preparation, and characterization of complex disulfide-rich proteins.
More Related Videos
Related Concept Videos
01:26Protein Modifications in the RER
5.4K
Modification of secretory and transmembrane proteins entering the rough ER begins in the ER lumen. These modifications aid in protein folding and stabilize the acquired tertiary structure. Protein modifications in the rough ER co-occur at different stages of protein folding.
Broadly, these modifications can be categorized into four main categories — glycosylation, formation of disulfide bonds, assembly of protein subunits, and specific proteolytic cleavages like removal of signal...
Broadly, these modifications can be categorized into four main categories — glycosylation, formation of disulfide bonds, assembly of protein subunits, and specific proteolytic cleavages like removal of signal...
5.4K
01:25Protein Folding
8.3K
Proteins are chains of amino acids linked together by peptide bonds. Upon synthesis, a protein folds into a three-dimensional conformation, critical to its biological function. Interactions between its constituent amino acids guide protein folding, and hence the protein structure is primarily dependent on its amino acid sequence.
Protein Structure Is Critical to Its Biological Function
Proteins perform a wide range of biological functions such as catalyzing chemical reactions, providing...
Protein Structure Is Critical to Its Biological Function
Proteins perform a wide range of biological functions such as catalyzing chemical reactions, providing...
8.3K
02:15Protein and Protein Structure
80.0K
Proteins are one of the most abundant organic molecules in living systems and have the most diverse range of functions of all macromolecules. Proteins may be structural, regulatory, contractile, or protective. They may serve in transport, storage, or membranes; or they may be toxins or enzymes. Their structures, like their functions, vary greatly. They are all, however, amino acid polymers arranged in a linear sequence.
A protein's shape is critical to its function. For example, an enzyme...
A protein's shape is critical to its function. For example, an enzyme...
80.0K
02:44Oxidation of Alkenes: Syn Dihydroxylation with Osmium Tetraoxide
10.6K
Alkenes are converted to 1,2-diols or glycols through a process called dihydroxylation. It involves the addition of two hydroxyl groups across the double bond with two different stereochemical approaches, namely anti and syn. Dihydroxylation using osmium tetroxide progresses with syn stereochemistry.
10.6K
02:04Oxidation of Alkenes: Anti Dihydroxylation with Peroxy Acids
6.0K
Diols are compounds with two hydroxyl groups. In addition to syn dihydroxylation, diols can also be synthesized through the process of anti dihydroxylation. The process involves treating an alkene with a peroxycarboxylic acid to form an epoxide. Epoxides are highly strained three-membered rings with oxygen and two carbons occupying the corners of an equilateral triangle. This step is followed by ring-opening of the epoxide in the presence of an aqueous acid to give a trans diol.
6.0K
01:29Protein Folding Quality Check in the RER
3.8K
ER is the primary site for the maturation and folding of soluble and transmembrane secretory proteins. The calnexin cycle is a specific chaperone system that folds and assesses the confirmation of N-glycosylated proteins before they can exit the ER lumen. The primary players of this quality check pipeline are the lectins, ER-resident chaperones, and a glucosyl transferase enzyme. In case the calnexin system in the lumen fails to salvage a misfolded protein, it is transported to the cytoplasm...
3.8K