Mapping the binding interface of ERK and transcriptional repressor Capicua using photocrosslinking.

TitleMapping the binding interface of ERK and transcriptional repressor Capicua using photocrosslinking.
Publication TypeJournal Article
Year of Publication2015
AuthorsFutran, AS, Kyin, S, Shvartsman, SY, A Link, J
JournalProc Natl Acad Sci U S A
Volume112
Issue28
Pagination8590-5
Date Published2015 Jul 14
ISSN1091-6490
KeywordsAmino 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>

DOI10.1073/pnas.1501373112
Alternate JournalProc. Natl. Acad. Sci. U.S.A.
PubMed ID26124095
PubMed Central IDPMC4507193
Grant ListR01 GM086537 / GM / NIGMS NIH HHS / United States
GM086537 / GM / NIGMS NIH HHS / United States