Title | Mapping the binding interface of ERK and transcriptional repressor Capicua using photocrosslinking. |
Publication Type | Journal Article |
Year of Publication | 2015 |
Authors | Futran, AS, Kyin, S, Shvartsman, SY, A Link, J |
Journal | Proc Natl Acad Sci U S A |
Volume | 112 |
Issue | 28 |
Pagination | 8590-5 |
Date Published | 2015 Jul 14 |
ISSN | 1091-6490 |
Keywords | Amino Acid Sequence, Animals, Binding Sites, Drosophila, Drosophila Proteins, Extracellular Signal-Regulated MAP Kinases, HMGB Proteins, Humans, Mass Spectrometry, Molecular Sequence Data, Photochemical Processes, Repressor Proteins |
Abstract | <p>Extracellular signal-regulated kinase (ERK) coordinates cellular responses to a range of stimuli by phosphorylating its numerous substrates. One of these substrates, Capicua (Cic), is a transcriptional repressor that was first identified in Drosophila and has been implicated in a number of human diseases. Here we use a chemical biology approach to map the binding interface of ERK and Cic. The noncanonical amino acid p-azidophenylalanine (AzF) was introduced into the ERK-binding region of Drosophila Cic, and photocrosslinking and tandem mass spectrometry were used to pinpoint its binding site on ERK. We also identified the ERK-binding region of human Cic and showed that it binds to the same site on ERK despite lacking conservation with the Drosophila Cic binding region. Finally, we mapped the amino acids involved in human Cic binding to ERK using AzF-labeled ERK. These results reveal the molecular details of the ERK-Cic interaction and demonstrate that the photocrosslinking approach is complementary to existing methods for mapping kinase-substrate binding interfaces. </p> |
DOI | 10.1073/pnas.1501373112 |
Alternate Journal | Proc Natl Acad Sci U S A |
PubMed ID | 26124095 |
PubMed Central ID | PMC4507193 |
Grant List | R01 GM086537 / GM / NIGMS NIH HHS / United States GM086537 / GM / NIGMS NIH HHS / United States |