Chapter title |
Site-Specific Protein Labeling Utilizing Lipoic Acid Ligase (LplA) and Bioorthogonal Inverse Electron Demand Diels-Alder Reaction
|
---|---|
Chapter number | 23 |
Book title |
Noncanonical Amino Acids
|
Published in |
Methods in molecular biology, January 2018
|
DOI | 10.1007/978-1-4939-7574-7_23 |
Pubmed ID | |
Book ISBNs |
978-1-4939-7573-0, 978-1-4939-7574-7
|
Authors |
Mathis Baalmann, Marcel Best, Richard Wombacher, Baalmann, Mathis, Best, Marcel, Wombacher, Richard |
Abstract |
Here, we describe a two-step protocol for selective protein labeling based on enzyme-mediated peptide labeling utilizing lipoic acid ligase (LplA) and bioorthogonal chemistry. The method can be applied to purified proteins, protein in cell lysates, as well as living cells. In a first step a W37V mutant of the lipoic acid ligase (LplAW37V) from Escherichia coli is utilized to ligate a synthetic chemical handle site-specifically to a lysine residue in a 13 amino acid peptide motif-a short sequence that can be genetically expressed as a fusion with any protein of interest. In a second step, a molecular probe can be attached to the chemical handle in a bioorthogonal Diels-Alder reaction with inverse electron demand (DAinv). This method is a complementary approach to protein labeling using genetic code expansion and circumvents larger protein tags while maintaining label specificity, providing experimental flexibility and straightforwardness. |
X Demographics
As of 1 July 2024, you may notice a temporary increase in the numbers of X profiles with Unknown location. Click here to learn more.
Geographical breakdown
Country | Count | As % |
---|---|---|
United States | 1 | 25% |
Unknown | 3 | 75% |
Demographic breakdown
Type | Count | As % |
---|---|---|
Members of the public | 2 | 50% |
Practitioners (doctors, other healthcare professionals) | 1 | 25% |
Scientists | 1 | 25% |
Mendeley readers
Geographical breakdown
Country | Count | As % |
---|---|---|
Unknown | 17 | 100% |
Demographic breakdown
Readers by professional status | Count | As % |
---|---|---|
Student > Ph. D. Student | 9 | 53% |
Student > Master | 2 | 12% |
Student > Doctoral Student | 2 | 12% |
Student > Bachelor | 1 | 6% |
Unknown | 3 | 18% |
Readers by discipline | Count | As % |
---|---|---|
Chemistry | 5 | 29% |
Agricultural and Biological Sciences | 3 | 18% |
Biochemistry, Genetics and Molecular Biology | 2 | 12% |
Chemical Engineering | 1 | 6% |
Philosophy | 1 | 6% |
Other | 2 | 12% |
Unknown | 3 | 18% |