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High-precision mapping of protein protein interfaces: an integrated genetic strategy combining en masse mutagenesis and DNA-level parallel analysis on a yeast two-hybrid platform.

Pajunen, Maria; Turakainen, Hilkka; Poussu, Eini; Peränen, Johan; Vihinen, Mauno LU and Savilahti, Harri (2007) In Nucleic Acids Research 35(16). p.103-103
Abstract
Understanding networks of protein-protein interactions constitutes an essential component on a path towards comprehensive description of cell function. Whereas efficient techniques are readily available for the initial identification of interacting protein partners, practical strategies are lacking for the subsequent high-resolution mapping of regions involved in protein-protein interfaces. We present here a genetic strategy to accurately map interacting protein regions at amino acid precision. The system is based on parallel construction, sampling and analysis of a comprehensive insertion mutant library. The methodology integrates Mu in vitro transposition-based random pentapeptide mutagenesis of proteins, yeast two-hybrid screening and... (More)
Understanding networks of protein-protein interactions constitutes an essential component on a path towards comprehensive description of cell function. Whereas efficient techniques are readily available for the initial identification of interacting protein partners, practical strategies are lacking for the subsequent high-resolution mapping of regions involved in protein-protein interfaces. We present here a genetic strategy to accurately map interacting protein regions at amino acid precision. The system is based on parallel construction, sampling and analysis of a comprehensive insertion mutant library. The methodology integrates Mu in vitro transposition-based random pentapeptide mutagenesis of proteins, yeast two-hybrid screening and high-resolution genetic footprinting. The strategy is general and applicable to any interacting protein pair. We demonstrate the feasibility of the methodology by mapping the region in human JFC1 that interacts with Rab8A, and we show that the association is mediated by the Slp homology domain 1. (Less)
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author
publishing date
type
Contribution to journal
publication status
published
subject
keywords
Saccharomyces cerevisiae: genetics, Insertional: methods, Mutagenesis, Membrane Proteins: metabolism, Membrane Proteins: chemistry, Membrane Proteins: genetics, rab GTP-Binding Proteins: chemistry, rab GTP-Binding Proteins: metabolism
in
Nucleic Acids Research
volume
35
issue
16
pages
103 - 103
publisher
Oxford University Press
external identifiers
  • pmid:17702760
  • scopus:34548743990
ISSN
1362-4962
DOI
10.1093/nar/gkm563
language
English
LU publication?
no
id
bef520fc-d4bb-48c8-8492-057ddf6f8097 (old id 3635266)
alternative location
http://www.ncbi.nlm.nih.gov/pubmed/17702760?dopt=Abstract
date added to LUP
2013-06-12 16:23:53
date last changed
2017-08-20 04:37:44
@article{bef520fc-d4bb-48c8-8492-057ddf6f8097,
  abstract     = {Understanding networks of protein-protein interactions constitutes an essential component on a path towards comprehensive description of cell function. Whereas efficient techniques are readily available for the initial identification of interacting protein partners, practical strategies are lacking for the subsequent high-resolution mapping of regions involved in protein-protein interfaces. We present here a genetic strategy to accurately map interacting protein regions at amino acid precision. The system is based on parallel construction, sampling and analysis of a comprehensive insertion mutant library. The methodology integrates Mu in vitro transposition-based random pentapeptide mutagenesis of proteins, yeast two-hybrid screening and high-resolution genetic footprinting. The strategy is general and applicable to any interacting protein pair. We demonstrate the feasibility of the methodology by mapping the region in human JFC1 that interacts with Rab8A, and we show that the association is mediated by the Slp homology domain 1.},
  author       = {Pajunen, Maria and Turakainen, Hilkka and Poussu, Eini and Peränen, Johan and Vihinen, Mauno and Savilahti, Harri},
  issn         = {1362-4962},
  keyword      = {Saccharomyces cerevisiae: genetics,Insertional: methods,Mutagenesis,Membrane Proteins: metabolism,Membrane Proteins: chemistry,Membrane Proteins: genetics,rab GTP-Binding Proteins: chemistry,rab GTP-Binding Proteins: metabolism},
  language     = {eng},
  number       = {16},
  pages        = {103--103},
  publisher    = {Oxford University Press},
  series       = {Nucleic Acids Research},
  title        = {High-precision mapping of protein protein interfaces: an integrated genetic strategy combining en masse mutagenesis and DNA-level parallel analysis on a yeast two-hybrid platform.},
  url          = {http://dx.doi.org/10.1093/nar/gkm563},
  volume       = {35},
  year         = {2007},
}