Short peptides containing an N-terminal primary or secondary amino group, an amino acid with a thiol group in the side chain and either a Gly-Gly-Trp or a Gly-Trp appendix at the C terminus, were used. The tryptophan residues facilitated HPLC analysis of the peptides and reaction products due to the strong absorption at 280 nm. Using a 0.5-ml reaction tube, the following reagents were added sequentially and incubated for 1 hour at 37°C: 78 μl of 60 mM NH4HCO3 (pH 8), 2 μl of peptide in H2O (5 mM), and 20 μl of linker reagent in ACN (4 mM). The final concentrations of peptide and linker reagent were 100 and 800 μM, respectively. The reactions were analyzed by LC-MS (LCMS-2020, Shimadzu) [injection volume of 4 μl; Kinetix C18 column, 2.6 μm, 100 Å, 50 mm by 2.1 mm (Phenomenex); gradient: solvent A (99.95% H2O and 0.05% formic acid) and 0 to 40% solvent B (99.95% ACN and 0.05% formic acid) for 5 min; flow, 1 ml/min; MS analysis, positive mode].

Note: The content above has been extracted from a research article, so it may not display correctly.



Q&A
Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.



We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.