Highly Efficient Separation of Methylated Peptides Utilizing Selective Complexation between Lysine and 18-Crown-6.
Anal Chem
; 92(23): 15663-15670, 2020 12 01.
Article
em En
| MEDLINE
| ID: mdl-33169968
ABSTRACT
Protein methylation is one of the most common and important post-translational modifications, and it plays vital roles in epigenetic regulation, signal transduction, and chromatin metabolism. However, due to the diversity of methylation forms, slight difference between methylated sites and nonmodified ones, and ultralow abundance, it is extraordinarily challenging to capture and separate methylated peptides from biological samples. Here, we introduce a simple and highly efficient method to separate methylated and nonmethylated peptides using 18-crown-6 as a mobile phase additive in high-performance liquid chromatography. Selective complexation between lysine and 18-crown-6 remarkably increases the retention of the peptides on a C18 stationary phase, leading to an excellent baseline separation between the lysine methylated and nonmethylated peptides. A possible binding mechanism is verified by nuclear magnetic resonance titration, biolayer interferometry technology, and quantum chemistry calculation. Through establishment of a simple enrichment methodology, a good selectivity is achieved and four methylated peptides with greatly improved signal-to-noise (S/N) ratios are successfully separated from a complex peptide sample containing 10-fold bovine serum albumin tryptic digests. By selecting rLys N as an enzyme to digest histone, methylation information in the histone could be well identified based on our enrichment method. This study will open an avenue and provide a novel insight for selective enrichment of lysine methylated peptides in post-translational modification proteomics.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Peptídeos
/
Éteres de Coroa
/
Lisina
Limite:
Animals
Idioma:
En
Ano de publicação:
2020
Tipo de documento:
Article