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Simultaneous Affinity Enrichment of Two Post-Translational Modifications for Quantification and Site Localization
Published on: February 27, 2020
Site-specific protein modification with a dirhodium metallopeptide catalyst.
Zhen Chen1, Brian V Popp, Cara L Bovet
1Department of Chemistry, Rice University, Houston, Texas 77005, United States.
ACS Chemical Biology
|June 16, 2011
Summary
A novel dirhodium metallopeptide catalyst enables precise chemical protein modification. This method works at physiological pH and directly in bacterial lysate for efficient protein functionalization and affinity tagging.
Area of Science:
- Biochemistry
- Chemical Biology
- Catalysis
Background:
- Chemical protein modification is crucial for biological research and drug development.
- Existing methods often lack specificity or require harsh conditions.
- Metallopeptide catalysts offer a promising avenue for targeted biomolecule functionalization.
Purpose of the Study:
- To develop a new method for site-specific chemical protein modification.
- To expand the utility of dirhodium-catalyzed reactions for biomolecules.
- To demonstrate efficient protein functionalization in complex biological environments.
Main Methods:
- Utilizing a dirhodium metallopeptide catalyst for chemical protein modification.
- Employing peptide-based molecular recognition for site-specificity.
- Testing the catalyst's activity at physiological pH and in biologically relevant buffers.
- Assessing modification efficiency directly in E. coli lysate.
- Synthesizing and utilizing a novel biotin-diazo conjugate for affinity tagging.
Main Results:
- Achieved site-specific protein functionalization using the dirhodium metallopeptide catalyst.
- Demonstrated broad side-chain scope for the catalyst.
- Expanded the utility of dirhodium-catalyzed biomolecule modification to physiological conditions.
- Confirmed specific protein modification directly in E. coli lysate.
- Successfully used a new biotin-diazo conjugate for affinity tagging of target proteins.
Conclusions:
- The designed dirhodium metallopeptide catalyst enables efficient and specific chemical protein modification.
- The method is applicable under physiological conditions and in complex biological samples like bacterial lysate.
- This work expands the capabilities of metallopeptide catalysts for protein functionalization and labeling.
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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 sequences.
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02:04Covalently Linked Protein Regulators
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.
These groups modify specific amino acids in a protein.
01:34Directing Proteins to the Rough Endoplasmic Reticulum
The organelle-specific signaling sequences direct proteins synthesized in the cytosol to their final destination like ER, mitochondria, peroxisomes, etc. Some of the proteins directed to ER are then trafficked via vesicles to other organelles within the cell or the extracellular environment through the Golgi complex. For example, the rough ER synthesizes soluble proteins for transportation to the lysosomes or secretion out of the cell. It can also synthesize transmembrane proteins that can...