High-throughput selection of transmembrane sequences that enhance receptor tyrosine kinase activation

Lijuan He, Andrew R. Hoffmann, Christopher Serrano, Kalina A Hristova, William C. Wimley

Research output: Contribution to journalArticle

Abstract

Dimerization is a critical requirement for the activation of the intracellular kinase domains of receptor tyrosine kinases (RTKs). The single transmembrane (TM) helices of RTKs contribute to dimerization, but the details are not well understood. Work with TM helices in various model systems has revealed a small number of specific dimerization sequence motifs, and it has been suggested that RTK dimerization is modulated by such motifs. Yet questions remain about the universality of these sequence motifs for RTK dimerization and about how TM domain dimerization in model systems relates to RTK activation in mammalian membranes. To investigate these questions, we designed a 3888-member combinatorial peptide library based on the TM domain of Neu (ErbB2) as a model RTK. The library contains many closely related, Neu-like sequences, including thousands of sequences with known dimerization motifs. We used an SDS-PAGE-based screen to select peptides that dimerize better than the native Neu sequence, and we assayed the activation of chimeric Neu receptors in mammalian cells with TM sequences selected in the screen. Despite the very high abundance of known dimerization motifs in the library, only a very few dimerizing sequences were identified by SDS-PAGE. About half of those sequences activated the Neu kinase significantly more than did the wild-type TM sequence. This work furthers our knowledge about the requirements for membrane protein interactions and the requirements for RTK activation in cells.

Original languageEnglish (US)
Pages (from-to)43-54
Number of pages12
JournalJournal of Molecular Biology
Volume412
Issue number1
DOIs
StatePublished - Sep 9 2011

Fingerprint

Receptor Protein-Tyrosine Kinases
Dimerization
Libraries
Polyacrylamide Gel Electrophoresis
Phosphotransferases
ErbB-2 Receptor
Peptide Library
Membrane Proteins
Peptides
Membranes

Keywords

  • high-throughput screening
  • neu
  • receptor tyrosine kinase
  • RTK activation
  • SDS PAGE

ASJC Scopus subject areas

  • Molecular Biology

Cite this

High-throughput selection of transmembrane sequences that enhance receptor tyrosine kinase activation. / He, Lijuan; Hoffmann, Andrew R.; Serrano, Christopher; Hristova, Kalina A; Wimley, William C.

In: Journal of Molecular Biology, Vol. 412, No. 1, 09.09.2011, p. 43-54.

Research output: Contribution to journalArticle

He, Lijuan ; Hoffmann, Andrew R. ; Serrano, Christopher ; Hristova, Kalina A ; Wimley, William C. / High-throughput selection of transmembrane sequences that enhance receptor tyrosine kinase activation. In: Journal of Molecular Biology. 2011 ; Vol. 412, No. 1. pp. 43-54.
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